Photoswitchable probe with distinctive characteristics for selective fluorescence imaging and long-term tracing

A photo-switchable and high-contrast bio-imaging indicator 4,4′-(1E,1′E)-(4,4′-(cyclopentene-1,2-diyl)bis(5-methylthiophene-4,2-diyl))bis(methan-1-yl-1-ylidene)bis(azan-1-yl-1-ylidene)bis(2-(benzo[d]thiazol-2-yl)phenol) (BMBT) has been demonstrated, by integrating photochromophore with excited-state intramolecular proton transfer (ESIPT) moiety. The ability of reversible emission switching enables arbitrarily selective labeling or concealing of cells simply by controlling light irradiation. Besides, when the emission was switched on, BMBT is demonstrated to exhibit unique characteristics of aggregation induced emission (AIE), providing a high on–off ratio for favorable bio-imaging. Thus, the non-labeling and easily-controlled selective imaging, as well as good biocompatibility indicates BMBT to be a favorable cell probe with great potentials for functional bio-imaging fluorophore.

In this work, a photoswitchable probe was synthesized by integrating a photochromophore with an excited-state intramolecular proton transfer (ESIPT) moiety. It was explored to be a favorable fluorophore for selective fluorescence imaging and long-term tracing.

Although there are many conventional fluorescent probes used for fluorescent imaging in the past few years,^1–5 such as rhodamine,⁶ cyanine dye,⁷ quantum dots,^8,9 and lanthanide probes,¹⁰ these fluorescent probes can only respond irreversibly to one event.^11,12 In comparison, photochromophores,¹³ which can reversibly response with UV and visible light,¹⁴ are more valuable fluorescent probes for regional optical marking of interested cells.^15,16 Because of their favourable characteristics, such as excellent thermal stability, good fatigue resistance and fast response time, diarylethenes derivatives have drawn wide spread concern of researchers.^17–20 Furthermore, the special optical properties of diarylethenes enable them to be suitable for long time and real time monitoring in bioimaging.The significance of targeted imaging of fluorescence probe for bio-samples in vitro or in vivo is well-known for their high sensitivity, non-invasiveness, real-time detection and especially selectivity.^21–24 However, the synthesis of these targeting materials is usually complicated. Moreover, for the same type of cells, no selectivity is demonstrated by these complex targeting agents.^21–24 Therefore, easy-prepared and easy-controlled non-labeling fluorescence imaging agents have always been pursued by both the industry and scientific communities. Among the above-mentioned photochromophores, diarylethenes are expected to be promising photochromic imaging candidates due to their favorable characteristics described in preceding paragraph. Prospectively, they can be used for selective long-term tracing if the photostability can be ensured.In our previous work, a series of AIE-active excited-state intramolecular proton transfer (ESIPT) complexes have been demonstrated to be good bio-imaging candidate with many advantages such as simple preparation, good biocompatibility, high quantum yields, fast cell staining as well as long-term anti-photobleaching.^25–27 Sometimes, ESIPT compounds exhibit dual emission, originated from keto and enol state, respectively. This caused extremely fast four-level photophysical cycle (E–E*–K*–K–E), mediated by intramolecular H-bonds immediately after photoexcitation, enables two emissions.^28,29 Herein, we have demonstrated a new type of multifunctional bio-imaging materials based on facile synthesis design concept, by introducing a photochromic diarylethene moiety to enable regional emission turn-on, and introducing an ESIPT moiety to allow good photo stability for long-term tracing. To the best of our knowledge, this is the first example with integrating abilities of non-labeling selectively long-term regional tracing.The new diarylethene derivative (BMBT, see the molecular structure in Scheme 1) has been synthesized (Fig. S1^†) via condensation reaction of 5-amino-2-(benzo[d]thiazol-2-yl)phenol with 4,4-(cyclopentene-1,2-yl)-bis(5-methyl-thiophene-2-formaldehyde), according to a previous reported procedure.^30,31 The molecular structures and purities were confirmed by ¹H NMR spectroscopy and mass spectroscopy (Fig. S6 and S7,^† see the synthesis details and full molecular characterizations in the ESI^†).Open in a separate windowScheme 1Structure and photochromic process of BMBT.The photoirradiation-induced changes in absorption and fluorescence spectra at room temperature are investigated in THF/DMEM mixture (1.0 × 10⁻⁵ mol L⁻¹). The open-ring isomer mainly exhibits two absorption peaked at 304 and 380 nm, respectively (Fig. S2^†). This is ascribed to the internal charge transfer and π–π transition of 2-(2′-hydroxy-phenyl)benzothiazole (HBT),^10,30 coupled with the CT inside the HBT unit from the hydroxyphenyl ring to the benzothiazole ring (Scheme 1). BMBT exhibits two emissions; one blue emission peak around 458 nm and a red emission peak around 600 nm, corresponding to the enol and keto emission, respectively (Fig. 1). Upon irradiation with ultraviolet light at 365 nm, the absorption band at longer wavelength region centered at 595 nm increased obviously with irradiation time (Fig. S2^†). This is caused by the formation of the closed-ring isomer (see the photochromic reaction in Scheme 1). Correspondingly, due to spectroscopic overlap between this longer-wavelength absorbance with the red emission ranged from 560–700 nm, the relative intensity of the red emission substantially decreases with the UV irradiation (Fig. 1), because of efficient energy transfer. Upon further visible light (λ = 520 nm) irradiation, the closed-ring isomer transfers back to the initial open-ring isomer, and thus the longer-wavelength absorption decreases and the red emission restores. This indicates good reversibility of the photochromic reaction.Open in a separate windowFig. 1Fluorescence emission changes of BMBT in THF/DMEM (1 : 200, vol : vol) mixture upon irradiation with 365 nm light (λ_ex = 385 nm) and visible light (λ_ex = 520 nm).Excitingly, BMBT exhibit prominent characteristics of Aggregation-Induced Emission (AIE).^31–33 The fluorescence intensity of BMBT in THF solution was relatively weak, while the powder or nanoparticles of the material dispersed in DMEM buffer exhibited a significantly enhanced emission (Fig. 2). When the DMEM buffer fraction was increased gradually from 0% to 20%, the fluorescence intensity only slightly enhanced. When the DMEM fraction was further increased to 40%, the emission exhibits a significant enhancement. The total intensity at the blue enol emission increased more than 10 times at 100% DMEM fraction as compared with that in THF solution. This AIE property enables high signal-to-noise ratios for favorable bioimaging (Fig. S3^†).Open in a separate windowFig. 2Fluorescence spectra of 1 × 10⁻⁵ M BMBT in the THF/DMEM mixture at different water fractions (λ_ex = 385 nm).Based on the good reversibility of light response and AIE characteristics, the practical application of the BMBT as bioprobe was further investigated. The biological imaging of BMBT was observed by using confocal laser scanning microscopy (CLSM). Blue luminescence in the cytoplasm of HeLa cells was observed after incubation with a THF/DMEM (1 : 200, vol : vol) solution of BMBT (20 μM) for 30 min at 37 °C (Fig. 2 inset). The overlay of luminescent images and bright-field images confirmed that BMBT was located mainly in the cytoplasm of cells rather than the membrane and nucleus (Fig. S3^†). Intense intracellular luminescence with a high signal-to-noise ratio (I₁/I₂ > 7) was detected between the cytoplasm (regions 3 and 1) and nucleus (region 2), also implying weak even few nuclear uptake of BMBT (Fig. S4^†). Besides, BMBT has a low cytotoxicity with the cellular viabilities estimated to be greater than 85% after 24 h incubation with the highest cultural concentration of 50 μM BMBT (Fig. S5^†).The luminescence switching of BMBT can also be achieved while alternating UV and visible light illumination in fixed HeLa cells. Cells (shown in red circle, Fig. 3a) were irradiated with 488 nm light (0.5 mW) for 3 min, the blue fluorescence of the irradiated cells was lamped off while the surrounding cells remained almost unchanged. Such fluorescence quenching is likely ascribed to the intramolecular fluorescence resonance energy transfer of BMBT, due to the intensified short-wavelength absorption band (270–450 nm) of the closed-open form of BMBT with the blue emission band (420–560 nm).³⁴ Upon irradiation with 405 nm light (1.25 mW) the fluorescence of all the selected cell was rapidly recovered within 1 min, caused by decrease of the relative intensity of the 270–450 nm absorption. The fluorescence can be repeatedly erased and recovered many rounds without significant fluorescence quenching, which was hardly achieved by conventional fluorophores (Fig. 3b).^35,36Open in a separate windowFig. 3(a) CLSM image (above) and the overlay image (bottom) of fixed HeLa cells incubated with 20 μM BMBT for 30 min at 37 °C (1) and (5) in original state; (2) and (6) irradiated by 488 nm light (0.5 mW) for a single cell; (3) and (7) all cells, and; (4) and (8) recovered by 405 nm light (1.25 mW). (b) Fluorescence switching of fixed HeLa cells by alternating UV (405 nm, 1.25 mW, 10 s/time) and visible (488 nm, 0.5 mW, 3 min/time) light illumination (λ_ex = 405 nm).This characteristic of selectively opto-marking or de-marking of cells may be used for non-invasive and dynamic tracing of the interested objects in vitro. That is, BMBT can arbitrarily opto-label or de-label interested cells without affecting cell proliferation. As shown in Fig. 4, the cell marked in the red circle was treated with visible light illumination (488 nm, 0.5 mW, 3 min), and its fluorescence was effectively quenched. Cells division of the remaining “bright” cells was observed under the microscope field of vision for a long-term tracing with time up to 36 h. Multiple new cells were produced, as indicated by upper white arrow. Even, as indicated by bottom white arrow, the tacked cells were observed to be doubled. The high brightness of the fluorescence in the proliferated cells indicated the good photo stability of BMBT. This indicated that BMBT can be used as a cell marker for arbitrarily selective erasing the fluorescence of the designated cell. It can also be used for selective lighting their emission by making them as the remaining bright cells after selective photo-erasing or selective photo-recovering after full erasing. This long-term tracking with non-labeling and selective optically marking or de-marking is seldom reported by other photochromophore-based bioimaging agents.³⁷ Herein, the excellent anti-photo bleaching characteristic in the long-term tracing is attributed to high photo stability of the HBT moiety.^26,27Open in a separate windowFig. 4Cells was treated with visible (488 nm, 0.5 mW, 3 min) light illumination for erasing the fluorescence. The remaining bright cells were incubated for another 36 hours, which were observed to be amplified normally (λ_ex = 405 nm).     

A novel fluorescent probe for imaging the process of HOCl oxidation and Cys/Hcy reduction in living cells

A new on-off-on fluorescent probe, CMOS, based on coumarin was developed to detect the process of hypochlorous acid (HOCl) oxidative stress and cysteine/homocysteine (Cys/Hcy) reduction. The probe exhibited a fast response, good sensitivity and selectivity. Moreover, it was applied for monitoring the redox process in living cells.

A new on–off–on fluorescent probe, CMOS, was designed and applied to detect the process of HOCl oxidation and Cys/Hcy reduction.

Reactive oxygen species (ROS) are indispensable products and are closely connected to various physiological processes and diseases.¹ For instance, endogenous hypochlorous acid (HOCl) as one of the most important ROS, which is mainly produced from the reaction of hydrogen peroxide with chloride catalyzed by myeloperoxidase (MPO), is a potent weapon against invading pathogens of the immune system.^2,3 However, excess production of HOCl may also give rise to oxidative damage via oxidizing or chlorinating the biomolecules.⁴ The imbalance of cellular homostasis will cause a serious pathogenic mechanism in numerous diseases, including neurodegenerative disorders,⁵ renal diseases,⁶ cardiovascular disease,⁷ and even cancer.⁸ Fortunately, cells possess an elaborate antioxidant defense system to cope with the oxidative stress.⁹ Therefore, it is necessary and urgent to study the redox process between ROS and antioxidants biosystems.Fluorescence imaging has been regarded as a powerful visual methodology for researching various biological components as its advantages of high sensitivity, good selectivity, little invasiveness and real-time detection.^10,11 To date, amounts of small molecular fluorescent probes have been reported for detection and visualization of HOCl in vivo and in vitro.^12–22,29 The designed strategies of HOCl sensitive probes are based on various HOCl-reactive functional groups, such as p-methoxyphenol,¹³p-alkoxyaniline,¹⁴ dibenzoyl-hydrazine,¹⁵ selenide,¹⁶ thioether,¹⁷ oxime,¹⁸ hydrazide,¹⁹ hydrazone.²⁰ But, many of these probes display a delayed response time and low sensitivity. And, only few fluorescent probes can be applied for investigating the changes of intracellular redox status.²¹ Besides, it''s worth noting that most of the redox fluorescent probes rely on the organoselenium compounds.²² Even though these probes are well applied for detection of cellular redox changes, excessive organic selenium is harmful to organisms and the synthesis of organoselenium compounds is high requirement and costly. Additionally, almost all the reports have only investigated the reduction effects of glutathione (GSH) as an antioxidant in the redox events. While, there are the other two important biothiols, cysteine (Cys) and homocysteine (Hcy), which not only present vital antioxidants, but also are tightly related to a wide variety of pathological effects in biosystem, such as slowed growth, liver damage, skin lesions,²³ cardiovascular,²⁴ and Alzheimer''s diseases.²⁵ However, the fluorescent probes for specially studying internal redox changes between HOCl and Cys/Hcy are rarely reported. In this respect, a novel redox-responsive fluorescent probe, CMOS, was designed and synthesized in this work, and we hope that it can be a potential tool for studying their biological relevance in living cells.Based on literature research, the aldehyde group has excellent selectivity in identification of Cys/Hcy, and the thiol atom in methionine can be easily oxidized to sulfoxide and sulfone by HOCl.^26,27 Considering these two points, we utilized 2-mercaptoethanol to protect the 3-aldehyde of 7-diethylamino-coumarin as the recognition part of HOCl, meaning that two kinds of potential recognition moieties are merged into one site. Fluorescent probe CMOS can be easily synthesized by the acetal reaction in one step (Scheme S1^†). A control molecule CMOS-2 was also prepared by 3-acetyl-7-diethylaminocoumarin (CMAC) similarly. The structure of all these compounds have been convinced by ¹H NMR, ¹³C NMR, and HR-MS (see ESI^†).As shown in Scheme 1a, we estimated that both CMOS and CMOS-2 can be rapidly oxidized in the appearance of HOCl. The oxidation product CMCHO of CMOS, which has the aldehyde moiety, can further react with Cys/Hcy to obtain the final product CMCys and CMHcy, respectively. In contrast, the oxidation product CMAC of CMOS-2 cannot combine with Cys/Hcy or other biothiols anymore (Scheme 1b).Open in a separate windowScheme 1Proposed reaction mechanism of CMOS and CMOS-2 to HOCl and Cys/Hcy.In order to confirm our design concept, the basic photo-physical characteristics of CMOS, CMCHO, CMOS-2 and CMAC were tested (Table S1, Fig. S1^†). Under the excitation wavelength 405 nm, CMOS and CMOS-2 exhibited strong fluorescence centred at 480 nm in PBS buffer solution, while the fluorescence of CMCHO and CMAC was weak around this band. The emission properties of CMOS and CMCHO were also investigated at the excitation wavelength 448 nm under the same experimental conditions as well (Fig. S2^†). After careful consideration, we chose 405 nm as the excitation wavelength in the follow-up experiments in vitro and in vivo.Next, the sensitivity of CMOS and CMOS-2 to HOCl and Cys/Hcy were investigated. As we expected, both the CMOS and CMOS-2 exhibited good response to HOCl. The fluorescence intensity of CMOS and CMOS-2 decreased gradually with addition of NaOCl (Fig. 1a, S3a^†), indicating that the fluorescence was switched off obviously in the presence of HOCl. The variation of intensity displayed good linearity with concentration of HOCl in the range of 0–20 μM (R² = 0.993, Fig. S4^†), and the detection limit of CMOS to HOCl was calculated to be 21 nM (S/N = 3). Subsequently, when Cys/Hcy was added to the final solution in Fig. 1a, the fluorescence intensity increased gradually within 180 min (Fig. 1b, S5^†). However, the fluorescence cannot be recovered by addition thiols to the CMOS-2 solution with excess HOCl (Fig. S3b^†). These results indicate that the probe CMOS can response to HOCl and Cys/Hcy in a fluorescence on-off-on manner, and can be used for monitoring the redox process with high sensitivity.Open in a separate windowFig. 1(a) Fluorescence responses of CMOS (2 μM) to different concentrations of NaOCl (0–200 μM). (b) Fluorescence responses of the CMOS solution (2 μM) with HOCl (200 μM) to Cys/Hcy (5 mM). (20 mM PBS buffer/CH₃CN, 7 : 3, v/v, pH = 7.4, λ_ex = 405 nm).To further identify the recognizing mechanism of probe CMOS, high performance liquid chromatography (HPLC) and mass spectral (MS) analysis were used to detect the redox process. Initially, probe CMOS displayed a single peak with a retention time at 3.7 min (Fig. 2a, S6^†) while reference compound CMCHO produced a single peak with a retention time at 2.5 min (Fig. 2b, S7^†). Upon the addition of HOCl to the solution of CMOS, the peak at 3.7 min weakened while 2.5 min and 2.2 min appeared (Fig. 2c). According to corresponding mass spectra, the new main peak at 2.5 min is related to compound CMCHO (Fig. S8^†). The other new peak of 2.2 min corresponds to the compound C3, which can be predicted as an intermediate in the oxidation process (Fig. S8^†).²⁸ The addition of Cys to the solution of CMCHO also caused a new peak with a retention time at 2.1 min, which has been confirmed to be the thioacetal product CMCys (Fig. S9^†). The possible sensing mechanism is depicted in Fig. S10.^†Open in a separate windowFig. 2The reversed-phase HPLC with absorption (400 nm) detection. (a) 10 μM CMOS. (b) 10 μM CMCHO. (c) 10 μM CMOS in the presence of 50 μM HOCl for 30 s. (d) 10 μM CMCHO in the presence of 1 mM Cys for 30 min. (Eluent: CH₃CN containing 0.5% CH₃COOH; 100% CH₃CN (0–7 min), 0.5 ml min⁻¹, 25 °C; injection volume, 5.0 μL).To study the selectivity of CMOS towards HOCl, we performed fluorescence response to different reactive oxygen species (ROS), reactive nitrogen species (RNS) and reactive sulfur species (RSS). As shown in Fig. 3a, CMOS exhibited significant change of fluorescence intensity only in the presence of HOCl, while other ROS and RNS, such as singlet oxygen (¹O₂), hydrogen peroxide (H₂O₂), hydroxyl radical (HO·), superoxide anion (O₂⁻), nitric oxide (NO), tert-butylhydroperoxide (t-BuOOH) and tert-butoxy radical (t-BuOO·) had no obvious fluorescence emission changes. Additionally, RSS which are abundant in biological samples, showed no influence in this process under the identical condition. The detection of reducing process was also investigated. As displayed in Fig. 3b, only cysteine and homocysteine induced excellent fluorescence recovery towards other reducing materials, such as RSS and various amino acids. Furthermore, the selectivity of CMOS-2 was also studied in the same condition. As expected, CMOS-2 could selectively detect HOCl, and not alter fluorescence intensity under various kinds of biothiols (Fig. S11^†). Therefore, our design strategy for the on–off–on probe is confirmed by results obtained above, with which CMOS can be utilized for detecting the redox process between HOCl and Cys/Hcy with high selectivity.Open in a separate windowFig. 3(a) Fluorescence response of CMOS (2 μM) to different ROS, RNS and RSS (200 μM). Bars represent emission intensity ratios before (F₀) and after (F₁) addition of each analytes. (a) HOCl; (b) KO₂; (c) H₂O₂; (d) ¹O₂; (e) HO·; (f) t-BuOOH; (g) t-BuOO·; (h) NO₂⁻; (i) NO₃⁻; (j) NO; (k) GSH; (l) Cys; (m) Hcy; (n) Na₂S; (o) Na₂S₂O₃; (p) Na₂S₂O₈; (q) NaSCN; (r) DTT; (s) Na₂SO₃. (b) Fluorescence response of the solution added HOCl in (a) to different RSS and amino acids. Bars represent emission intensity ratios before (F₂) and after (F₃) addition of each analytes (5 mM). (a) Cys; (b) Hcy; (c) Na₂S; (d) Na₂S₂O₃; (e) Na₂S₂O₈; (f) NaSCN; (g) DTT; (h) Na₂SO₃; (i) Ala; (j) Glu; (k) Gly; (l) His; (m) Ile; (n) Leu; (o) Met; (p) Phe; (q) Pro; (r) Ser; (s) Trp; (t) Vc; (u) GSH. (20 mM PBS buffer/CH₃CN, 7 : 3, v/v, pH = 7.4, λ_ex/λ_em = 405/480 nm).Subsequently, the influence of pH on probe CMOS was measured. The fluorescence intensity of CMOS and CMCHO perform no significant variances in wide pH ranges (pH = 4–11, Fig. S12a^†). Fluorescence intensity changes could be observed immediately when HOCl was added into the solution of probe CMOS, especially in alkaline condition (Fig. 4a). Considering the pK_a of HOCl is 7.6,²⁹CMOS is responsive to both HOCl and OCl⁻. Alkaline condition was also benefit for the fluorescence recovery of CMOS from Cys/Hcy (Fig. S12b^†). It is reasonable to consider that thiol atom displays higher nucleophilicity in alkaline condition. From the stop-flow test, the UV-visible absorbance of probe CMOS sharply decreased at the wavelength of 400 nm (Fig. 4b). The response time was within 10 s and the kinetic of the reaction was fitted to a single exponential function (k_obs = 0.67 s⁻¹). The ability of instantaneous response is extremely necessary to intracellular HOCl detection.Open in a separate windowFig. 4(a) Fluorescence responses of CMOS to HOCl under different pH values. Squares represent emission intensity ratios after (F₁) and before (F₀) addition of 200 μM HOCl (λ_ex/λ_em = 405/480 nm). (b) Time-dependent changes in the absorption intensity of CMOS (1 μM) before and after addition of HOCl. (20 mM PBS buffer/CH₃CN, 7 : 3, v/v, pH = 7.4, λ_abs = 400 nm).With these data in hand, we next applied CMOS for fluorescence imaging of the redox changes with HOCl and Cys/Hcy in living cells. After incubation with 5 μM CMOS at 37 °C for 30 min, intense fluorescence was observed of the SKVO-3 cells in the optical window 425–525 nm (Fig. 5a and d), indicating the probe can easily penetrate into cells. Treating the cells with 100 μM NaOCl led to remarkable fluorescence quenching as the probe sensed the HOCl-induced oxidative stress (Fig. 5b and e). After 3 min, the cells were washed with PBS buffer three times, and added 5 mM Cys/Hcy for 1 h, respectively. Then the fluorescence was recovered obviously (Fig. 5c and f). Experimental results clearly declare that the probe CMOS was successfully used to detect the process of HOCl oxidative stress and Cys/Hcy reducing repair in living cells.Open in a separate windowFig. 5Fluorescence imaging of the process of HOCl oxidative stress and thiols repair in CMOS-labeled SKVO-3 cells. Fluorescence images of SKVO-3 cells loaded with 5 μM CMOS at 37 °C for 30 min (a and d). Dye-loaded cells treated with 100 μM NaOCl at 25 °C for 3 min (b and e). Dye-loaded, NaOCl-treated cell incubated with 5 mM Cys (c), 5 mM Hcy (f) for 1 h. Emission intensities were collected in an optical window 425–525 nm, λ_ex = 405 nm, intensity bar: 0–3900.     

Retracted Article: Facile preparation of bithiazole-based material for inkjet printed light-emitting electrochemical cell

Light-emitting electrochemical cell of bithiazole-based material was fabricated by solution processing rendered high external quantum efficiency over 12.8% and luminance of 1.8 10⁴ cd m⁻².

Light-emitting electrochemical cell of bithiazole-based material was fabricated by solution processing rendered high external quantum efficiency over 12.8% and luminance of 1.8 10⁴ cd m^−2.

It is well known that inkjet printing works as precise and versatile patterning method for printed electronics.^1,2 As for its advantages, it is already being exploited widespread for printing electronics.^3,4 Those merits are easily processed from solutions and conveniently used for air-stable electrodes.^5,6Light-emitting electrochemical cells (LECs) have emerged as an active layer,^7,8 arousing tremendous attention over the years.^9–11 Nevertheless, LECs are simpler than organic light-emitting diodes (OLEDs).^12,13 Due to its single layer architecture, low fabrication price and operating voltages, LECs are considered as a promising, next-generation, emissive thin-film technology.^14–16The most efficient device used to date for LECs are biscyclometalated iridium(iii) (Ir) complexes, because they have highly efficient and stable devices spanning the whole visible range.^17–19 However, avoiding the use of Ir is strongly desired because of its high cost and limited supply.^20–22 Till now, thiazole has worked as active component in a LECs system with a long-lived charge-separation molecule, without additional ions in its active layer.^23–26 As for its advantages, it can reduce recombination of charge carriers and facilitate carrier transfer.^27–29 More and more importance has been attached to thiazole compounds, especially bithiazole-based ones.^30–33 Therefore, these have aroused interest in the syntheses of bithiazoles.^34–36Based on our previous research,^37–41 the use of bithiazole ligands already explored by our group for photocatalytic technology has the advantage of an easy transformation of neutral complexes into charged ones by substitution on the N-atom of the thiazole moiety.^42,43 Furthermore, we demonstrate the fabrication of LEC devices by a combination of inkjet printing and spin coating, on A4 paper substrates for low cost, disposable and flexible conductive pattern.^44,45 Their corresponding material characteristics were closely investigated, a detailed report of the material properties of the resulting coatings, and an envisaged proof of concept application are disclosed in this paper. These fully printed devices demonstrate the potential upscaling of the fabrication of optoelectronic devices.Herein we report the design for a range of bithiazole derivatives 1a–1c (Fig. 1, their synthesis is shown in Scheme S1^†), and explore their properties. First of all, the UV-visible diffuse-reflectance (UV-vis DRS) spectra of those specimens at room temperature are displayed in Fig. 2 and the data are collected in Table S1.^† This enhancement was ascribed to the increase of aromatic rings, which has an intense absorption in the visible-light region. And their bandgap energies (E_g) are between 2.97–3.01 eV (Table S1^†), and are estimated from these absorption spectra according to the Kubelka–Munk method. Expanding the conjugated system and electron density with these donor groups, it can lead to a larger bathochromic shift of the absorption maximum.^46,47Open in a separate windowFig. 1Chemical structures of compounds 1a–1c described in this work.Open in a separate windowFig. 2UV-vis DRS spectra of 1a–1c.All of the three compounds are photoluminescent at room temperature, the relevant fluorescence peak maximum ranges from 360 to 398 nm (Fig. 3). They emit blue light when they are being excited. In agreement with Fig. 3, the presence of electron-releasing groups would shift the emission maximum. Among them, the most electron-donating carbazole unit would give the most red-shifted peak at 398 nm for 1c (Table S1^†). The PL quantum yield (ϕ_PL) studies show that both 1a and 1b are as high as 62 and 78%, while another one is 85% (Table S1^†). The data for their fluorescent quantum yields largely depends on interaction effect with molecules 1a–1c in the crystal packing. In this context, π–π intermolecular interactions can inhibit the fluorescence.⁴⁸Open in a separate windowFig. 3Typical PL spectra of 1a–1c.It may indicate that enlarging the conjugation length and electron density with these donor groups plays an important role in increasing the ϕ_PL.⁴⁹ Moreover, those with high ϕ_PL may be suitable for application as efficient light-emitting material in LECs.⁵⁰In addition, time-resolved measurements of the donor lifetime (τ) in 1a–1c were carried out, and their corresponding values are given in Table S1.^† The bithiazole derivatives 1a–1c showed long-lived singlet fluorescence lifetimes (τ_F), ranging between 1.04 and 9.54 ns (Table S1^†). This was on average more than double the fluorescence lifetime of the bithiazole starting material.⁵¹ These three samples possess relatively high decay time, as a result of their inhomogeneity.⁵² Compound 1c exhibited the longest fluorescence lifetime decay τ_F = 9.54 ns. The sensitivity of the emission to the polarity of the solvent is beneficial for an intramolecular charge transfer (ICT)-like emission.⁵³ This difference probably accounts for the high concentration of donor units in the bithiazole backbone, low rates of intersystem crossing to reactive triplet states.^48,54Consequently, the composite 1c was the best performing LEC and chosen for the below test. The EL spectra obtained for both devices were almost identical, with a main peak at 498 nm and other peak at 504 nm wavelength as illustrated in Fig. 4a. It may account for trapping, cavity, and self-absorption effects from within the LEC device 1c multilayer-structure. The spin-coated emission is quantified by the commission Internationale de L’Eclairage (CIE) coordinates of (0.28, 0.42), and a colour rendering index (CRI) of 65 (Table S2^†). While inkjet-printed light emission with CIE coordinates of (0.34, 0.43) and a CRI value of 83 was achieved for device 1c (Table S2^†). It exhibits good colour rendering indices (CRI > 70), this device exhibited a warm-white appearance, and the colour rendering is considered sufficient for indoor lighting applications.Open in a separate windowFig. 4Comparison of device characteristics with spin-cast and inkjet printer emitting layer for 1c: (a) electroluminescent spectra, (b) luminance vs. voltage, (c) current density vs. voltage, (d) efficiency vs. luminance behaviour.The time-dependent brightness and current density under constant biases of 2.9–3.3 V for device 1c are shown in Fig. 4b and c. Interestingly, the device with the inkjet-printed emitting layer (EML) produced a light output of around 3600 cd m⁻², whereas 4600 cd m⁻² was achieved with the spin-coated EML. In other words, the device with the inkjet printed EML obtained about 78.3% of brightness compared with the reference device. The differences between the two devices regarding current efficiency (J) and power efficiency (PE) were rather closed, as listed in the Table S2.^† A luminous efficiency much larger than 10 cd A⁻¹ was achieved for the inkjet-printed EML, which tended to be 73.9%, and is similar to the spin-coated emitting layer.The performance gap between the devices (Fig. 4d) can be attributed to differences in the height and surface roughness of the emitting layer.⁵⁵ Lateral sizing histograms (Fig. S11^†) show that the spin-coated EML possessed a thickness of 35 nm and a very smooth surface. The inferior performance of the inkjet-printed EML is caused by a less homogeneous surface morphology and an overall fatter layer.⁵⁶ The depth of the inkjet-printed EML ranged from 25 nm (pixel centre) to 35 nm (pixel edges). As a consequence, it is an inkjet deposition process and the related evaporation dynamics reduces the light output and efficiency, but also postpones the stability of the device in the system.^10,57We then shifted our attention to the turn-on kinetics, efficiency and long-term stability of the LEC device for 1c; the typical evolution of current density and light emission is presented in Fig. 5a and b, showing a slow turn-on followed by a progressive decay over time.Open in a separate windowFig. 5(a) Current density (closed symbols) and brightness (open symbols) versus time at 3 V for a device with the representative sample 1c; (b) EQE versus time at an applied voltage of 3 V.The build-up of the light output is synchronous with that of the current density. This time-delayed response is one of the striking features of the operation of an electrochemical cell and reflects the mechanism of device operation.²² The champion device in this set exhibited a peak power efficiency (PE) of 17.4 lm W⁻¹ at a luminance of 18 000 cd m⁻². Meanwhile, the current density began to decrease, possibly due to the electrochemical oxidation or electro corrosion that occurred on component 1c through accumulated electroinduced holes, and then it kept up a durative datum.⁵⁸ Apparently it exhibited outstanding long-term operation stability beyond 40 h (Fig. 5b).The time-dependent external quantum efficiency (EQE) of LEC for the representative substrate 1c are shown in Fig. 5b. It also exhibited similar temporal tendency in device efficiency. When a bias was applied on the LECs, the EQE quickly increased since balanced carrier injection was achieved by the formation of the doped layers.⁵⁹ After attaining the crest value, the device current was still rising while the EQE reduced little by little. It implied that both growing the doped layers and weakening of the EQE was resulting from exciton quenching near the bithiazlole core in persistently extended doped layers.⁶⁰ Doping-induced self-absorption was rather adaptable to the temporal roll-off in device efficiency. This configuration shows the best performance (EQE = 12.8%) compared to those devices prepared with the single compounds, which is given in Table S3.^† The reason may be that the steady-state recombination zone in 1c device was close to the central active layer.⁶¹ Besides it contains fluorenes on its side chains, constructing plane structure, benefiting from good electron injection abilities, leading to enhancing the EQE itself.⁶²The fabrication of inkjet-printed bithiazole interdigitated electrode (IDE) is illustrated in Fig. 6. All the prepared varieties of bithiazole-based ink have been found to be highly stable with nil or miniscule precipitation in over 180 days being stored on the shelf (Fig. 6a). After HCl treatment of the paper substrate, the prepared inks have filled within ink cartridges of a low-cost desktop printer Canon iP1188 (Fig. 6b). Fig. 6c shows the photograph of inkjet-printed conductive patterns.Open in a separate windowFig. 6(a) The representative ink based on sample 1c was stable for 180 days; (b) electrode printing using Canon iP1188 printer; (c) scale showing size of printed electrodes; (d) interdigitated electrode (inset shows small size printed pattern).In addition, the most attractive prospect of the bilayer device structure at this stage is the possibility for patterned emission for the creation of a static display. Fig. 7 presents a photograph of a small portion of a larger static display, with a resolution of 170 PPI, which repeatedly exhibits a message in the form of the word “LEC”. As shown in Fig. 7, the pixel array produced are fairly luminous. The EL spectra were collected from each pixel on the substrate, the emission peak wavelength was about 510 nm and the full width at half maximum was about 170 nm. The low variation in emission intensity in the different pixels implies a small variation in thickness of the solution-coated layers. More specifically, if we assume a minimum diameter of 20 μm for an inkjetted electrolyte droplet, and a smallest inter-droplet distance of 10 μm, we attain a pitch of 30 μm, which corresponds to a high display resolution of 850 PPI. Finally, we draw attention to the herein presented static-display LEC that comprises solely air-stabile materials, and that we routinely fabricate the bilayer stack under ambient atmosphere.Open in a separate windowFig. 7(a) The patterned light emission from a bilayer LEC, with the emission pattern defined by the selected positions of the ink jetted electrolyte droplets. (b) The droplet diameter and pitch were 50 and 150 μm, respectively, and the device was driven at V = 3 V. The scale bar measures 300 μm.     

Disproportionation-induced solid-state fluorescence in 6,13-dihydropentacenes

The thermally and photolytically induced disproportionation of 6,13-dihydropentacene derivatives into tetrahydropentacenes and pentacenes results in unique solid-state fluorescence. The fluorescence thereby depends on the molecular structure and the molecular arrangement in the solid state.

The thermally and photolytically induced disproportionation of 6,13-dihydropentacene derivatives into tetrahydropentacenes and pentacenes results in unique solid-state fluorescence.

Disproportionation reactions are arguably one of the most important classes of organic reactions, especially in the context of coal liquefaction and aromatization, as well as Cannizzaro and biochemical reactions. However, reports on disproportionation reactions of aromatic hydrocarbons remain scarce. Stein and co-workers have reported the disproportionation of 9,10-dihydroanthracene (>350 °C; liquid phase pyrolysis) into tetrahydroanthracene and anthracene.¹ Pentacene has also been reported to undergo disproportionation at 320 °C in a horizontal vapor phase deposition furnace to produce 6,13-dihydropentacene and polycondensed aromatic hydrocarbons.² It has been postulated that H-atom-transfer processes play a crucial role in such disproportion reactions. Indeed, H-atom transfer from 9,10-dihydroantharacene,^3,4 xanthene,⁴ fluorene,⁵ acridan,⁶ and other H donors (D–H) with weak C–H bonds⁷ to unsaturated H acceptors has been widely explored.On the other hand, the molecular design of π-conjugated systems is crucial for the development of new functional materials. Nevertheless, reports on π-conjugated systems exhibiting disproportionation-induced photoluminescence changes are relatively rare. Against this background, we decided to explore the utility of disproportionation reactions for the development of fluorescent chromic materials. Herein, we report the synthesis and disproportion of 6,13-dihydropentacene derivatives, as well as a detailed investigation into the fluorescence properties of the obtained compounds in the solid state.The synthesis of N,N′-bisalkyl-6,13-dihydropentacene[2,3:9,10]biscarboxyimides 3a–c is outlined in Scheme 1. We have previously reported the synthesis of key starting material 6,13-dihydropentacene-2,3,9,10-tetracarboxylic acid (1).⁸ Tetracarboxylic acid 1 was converted into the corresponding anhydride (2) in good yield using a modification of the procedure reported by Qian and co-workers.⁹N,N′-Bisalkyl-6,13-dihydropentacene[2,3:9,10]biscarboxyimides 3a–c were then obtained in 20–64% yield by stirring anhydride 2 under reflux in the presence of the corresponding amines. The structure of the obtained dihydropentacene bisimides was confirmed by ¹H and ¹³C NMR spectroscopy, IR spectroscopy, FAB mass spectrometry, and elemental analysis. The ¹H NMR spectrum of 3a in CDCl₃ shows a singlet at 4.36 ppm, which was assigned to the central ring of dihydropentacene. In the aromatic region, two singlets appear at 8.03 and 8.23 ppm, which were attributed to the naphthalene ring. The ¹³C NMR of 3a displays 13 signals, including five aromatic carbon atoms (123.5, 128.0 × 2, 134.4, 138.4, and 168.3 ppm) for the naphthalene ring, one carbonyl carbon atom (186.3 ppm), and one sp³-hybridized carbon atom of the central ring (38.5 ppm). In general, the spectra of 3a–c are very similar.Open in a separate windowScheme 1Synthesis of N,N′-bisalkyl-6,13-dihydropentacene[2,3:9,10]bis-carboxyimides 3a–c.Subsequently, we examined the disproportionation reaction of dihydropentacene 3a in the solid state (Scheme 2). Interestingly, upon heating solid 3a from room temperature to 230 °C, the color of the solid changed from colorless to green. To elucidate the origin of this green material, we investigated the absorption features of a thin film of 3a before and after heating (250 °C; 3 h). In the difference spectrum of 3a (Fig. 1), the positive absorption bands at 523, 560, and 611 nm were attributed to an expansion of the π-conjugated system. A comparison with the absorption spectrum of independently synthesized N,N′-bispentylpentacene[2,3:9,10]biscarboxyimide (4a) revealed very similar features. The formation of pentacene derivative 4a was also confirmed by ¹H NMR spectroscopy (Fig. S1^†). After heating (250 °C; 40 min), the main signals were assigned to the starting material (3a), and the new signals suggested a conversion <7%. The ¹H NMR spectrum in CDCl₃ showed three singlets at 8.50, 8.94, and 9.12 ppm, which were assigned to the protons attached to the pentacene core, respectively. Moreover, three singlets for aromatic protons (8.06, 8.22, and 8.39 ppm) and two singlets for methylene protons (4.27 and 4.81 ppm) were observed for tetrahydropentacene derivative 5a. Upon exposure to air, the signals of 4a disappeared and two singlets at 8.56 and 9.14 ppm appeared, which were assigned to pentacene quinone 7a, i.e., the oxidation product of pentacene 4a (Fig. S2^†).^8a,10 An integral 5a : 7a ratio of 1 : 1 was estimated. These results strongly indicate that the thermal reaction of dihydropentacene 3a affords pentacene 4a and tetrahydropentacene 5a (Scheme 2).Open in a separate windowScheme 2Disproportionation reaction of dihydropentacene 3a and decomposition of pentacene derivative 4a.Open in a separate windowFig. 1The absorbance difference spectrum of a thin film of 3a before and after the heating.Moreover, we discovered that N,N′-bisalky-6,13-dihydropentacene[2,3:9,10]bis-carboxyimide 3a exhibits solid-state fluorescence (Fig. 2); under illumination with UV light (λ_ex = 365 nm), bright blue fluorescence was observed. Interestingly, upon heating 3a in solid state from room temperature to 230 °C, the color of the apparent fluorescence gradually changed from bright blue to yellow and then red (Fig. 2a). Such phosphorchromaticity was observed for both film and crystalline samples (Fig. 2b), and the original color was not recovered even after cooling to room temperature. The differential scanning calorimetry (DSC) cooling and heating curves of 3a showed no peaks between room temperature and 270 °C (Fig. S3^†). In the case of 3b and 3c, similar fluorescence color changes were observed, while the crystal appearance changed from transparent to opaque. Polarized optical microscopy images revealed rough and cracked surfaces of the crystals of 3b and 3c after heating (Fig. S4^†).Open in a separate windowFig. 2(a) Apparent fluorescence color change of a 3a film with the increasing temperature. (b) Apparent fluorescence color change of crystals of 3a.We also observed significant changes in the fluorescence spectra of a thin film of 3a with increasing temperature (Fig. S5^†). The intensity of the fluorescence of the naphthaleneimide chromophore (300–550 nm) decreased with increasing temperature, while the intensity of the new fluorescence band at 638 nm, which was assigned to a pentacene chromophore, increased. The decaying emission of the naphthaleneimide skeleton may be explained in terms of an energy transfer from 3a to 4a.¹¹Fig. S6^† shows the concentration-dependent emission spectra of 3a in CHCl₃. At a concentration of 1.9 × 10⁻⁴ M, a monomer emission band from the naphthaleneimide moiety was observed at 390 and 400 nm (λ_ex = 280 nm). Upon increasing the concentration of 3a beyond 7.7 × 10⁻⁴ M, the fluorescence intensity of the naphthaleneimide moieties decreased and two new excimer emission bands emerged at approximately 440 and 460 nm with growing intensity. These results indicate that the solid-state fluorescence at 440 nm does not correspond to monomer emission, but to aggregate emission.In order to determine the molecular conformation in the crystals and rationalize the observed reactivity, a single-crystal X-ray diffraction analysis was carried out (Fig. 3). Depending on the solvent, two different crystalline structures were identified in the case of 3a. A structural analysis of single crystals of 3a–c grown from benzonitrile solutions revealed a planar central six-membered ring of the dihydropentacenes (Fig. 3a, and S8^†). The bond angle sums of the central six-membered rings are close to 720°. Numerous hitherto reported crystal structures are characterized by flattened boat conformations for dihydroanthracene and dihydropentacene derivatives, whereas the preferred conformation of dihydroaromatics should be a planar according to a theoretical study.¹² In fact, there are a few crystallographic reports on dihydroanthacene¹³ and dihydropentacene¹⁴ skeletons with a planar geometry. In contrast, the structural analysis for crystals of 3a grown from a THF solution revealed a V-shaped dihydropentacene skeleton with a bent central six-membered ring 3a_(v).Open in a separate windowFig. 3ORTEP drawings of (a) 3a_(planer) and (b) 3a_(v) (thermal ellipsoids at 50% probability).The energy difference between the planar and V-shaped conformers was examined using density functional theory (DFT) calculations on model compound 3′, which bears H atoms instead of alkyl chains. The V-shaped conformer 3′_(v) is by 4.0 kcal mol⁻¹ thermodynamically more stable than the planar conformer 3′_(planar). These results suggest a small energy difference between the two conformers. Consequently, the solid-state structure of 3a is most likely determined by packing forces in the crystal.Interestingly, the phosphorchromaticity changes in the solid state were also triggered by photoirradiation (>300 nm); however, the reaction was limited to the surface of the specimen. The phosphorchromaticity switching rate was sensitive to the substituents on the dihydropentacene. Upon photoirradiation of solids 3a_(planar) and 3b (λ_ex > 300 nm; 1 h), the apparent fluorescence color gradually changed from bright blue to red. In the case of 3_(v) and 3c, the apparent fluorescence color did not change, not even after two days of photoirradiation. To clarify the different reactivity of the planar and V-shape conformers of 3a, further DFT calculations were carried out. Fig. 4 shows the relevant Kohn–Sham molecular orbitals for the optimized structures of 3′_(planar) and 3′_(v). Both HOMOs are clearly localized on the C–H bonds of the central six-membered ring and the two naphthalene units. Interestingly, the LUMO of 3′_(planar) is notably less localized around the C–H bond of the central six-membered ring than the LUMO of 3′_(v). These results indicate that the excited state of 3a_(planar) favors the elimination of the C–H protons of the central six-membered ring compared to the case of 3a_(v).Open in a separate windowFig. 4Molecular Kohn–Sham orbitals of 3′_(v) and 3′_(planer), calculated at the M06-2X/6-31G* level of theory.To evaluate the differences in photoreactivity depending on the substituents, the molecular arrangements in the crystals were investigated. None of the crystals contained any solvent in the lattice. The distances between the H (from the C–H bonds of the central six-membered ring) and C (from the naphthalene ring of the nearest-neighboring molecule atom) atoms were ca. 3.0 Å (3a), 2.8 Å (3b), and 3.3 Å (3c). The long distance in the crystals of 3c should thus be unfavorable for hydrogen-atom transfer from the C–H bond of the central six-membered ring to the naphthalene ring of the neighboring molecule.     

Coumarin-based fluorescent ‘AND’ logic gate probes for the detection of homocysteine and a chosen biological analyte

With this research we set out to develop a number of coumarin-based ‘AND’ logic fluorescence probes that were capable of detecting a chosen analyte in the presence of HCys. Probe JEG-CAB was constructed by attaching the ONOO⁻ reactive unit, benzyl boronate ester, to a HCys/Cys reactive fluorescent probe, CAH. Similarly, the core unit CAH was functionalised with the nitroreductase (NTR) reactive p-nitrobenzyl unit to produce probe JEG-CAN. Both, JEG-CAB and JEG-CAN exhibited a significant fluorescence increase when exposed to either HCys and ONOO⁻ (JEG-CAB) or HCys and NTR (JEG-CAN) thus demonstrating their effectiveness to function as AND logic gates for HCys and a chosen analyte.

With this research we set out to develop of a number of coumarin-based ‘AND’ logic fluorescence probes that were capable of detecting a chosen analyte in the presence of HCys.

Homocysteine (HCys) is a non-proteinogenic amino acid, formed from the de-methylation of methionine,¹ which is then converted into cysteine (Cys) via a vitamin B₆ cofactor. Typical physiological concentrations of HCys range between 5–15 μmol L⁻¹.² However, elevated levels of HCys (>15 μmol L⁻¹), which is known as hyperhomocysteinemia (hHCys),³ have been associated with pregnancy disorders, Alzheimer''s disease, cardiovascular disease and neurodegenerative diseases (NDs).^4–6 It is believed that the main cause of HCys induced toxicity is through the non-enzymatic modification of proteins. This is achieved through irreversible covalent attachment of the predominant metabolite of HCys, homocysteine thiolactone (HTL), to lysine residues; a phenomenon known as ‘protein N-homocysteinylation’ that results in the loss of a proteins structural integrity leading to loss of enzymatic function and aggregation.⁷A number of fluorescent sensors have been developed for the detection of HCys to help improve our understanding of its role in biological systems.^8–11 However, these fluorescent probes have focused on the detection of a single biomarker (HCys), however, processes associated with HCys induced toxicity often involve more than one biochemical species. For example, it has been reported that peroxynitrite (ONOO⁻) and nitric oxide (NO˙) play a significant role in HCys-mediated apoptosis in trigeminal sensory neurons¹² and HCys has been reported to induce cardiomyocytes cell death through the generation of ONOO⁻.¹³ The production of ONOO⁻ is believed to be the result of an increased production of superoxide (O₂˙⁻) by HCys activating the enzyme NADPH oxidase.^14–16 This increased production of O₂˙⁻ leads to a reduction in the bioavailability of NO˙ by increasing the formation of ONOO⁻ (NO˙ + O₂˙⁻ → ONOO⁻).¹⁷ The reported ONOO⁻ concentrations in vivo are believed to be approximately 50 μM but, higher concentrations of 500 μM have been found in macrophages.^18,19 Furthermore, hypoxia has been reported to facilitate HCys production in vitamin-deficient diets²⁰ where hypoxia leads to an upregulation of nitroreductase (NTR) – a reductive enzyme upregulated in cells under hypoxic stress.^21,22 Therefore, the development of tools that enable an understanding of the relationship of HCys with these biologically important species would be highly desirable.To achieve this, a number of fluorescent probes have been developed that are capable of detecting two or more analytes.²³ These include AND logic gate based-fluorescence probes, which require both analytes to work in tandem to produce a measurable optical output.^24–28 In our group, we have developed several ‘AND’ reaction-based probes for the detection of ROS/RNS and a second analyte.^29–32 These ‘AND’ logic scaffolds have been used to detect two analytes within the same biological system.^24,33Owing to the pathological role of HCys, we set out to develop the first example of a fluorescent probe for the detection of HCys and biological related analyte. Aiming towards that target, we became interested in a previously reported coumarin-based fluorescent probe developed by Hong et al.CAH, with a salicylaldehyde (Fig. 1).³⁴ Salicylaldehyde is a known reactive unit towards HCys/Cys, therefore we believed CAH could be used as a scaffold for the development of ‘AND’-based systems for the detection of HCys/Cys and a second analyte.³⁴ In the presence of HCys, CAH exhibited a ‘turn-on’ fluorescence response which is attributed to the nucleophilic nature of the nitrogen and sulfur atoms resulting in thiazine ring formation (Scheme S1, Fig. S1 and S2^†).^34–36Open in a separate windowFig. 1(a) CAH – a core fluorescent unit that enables the synthesis of ‘AND’ based fluorescent probe for the detection of HCys/Cys and a second analyte. (b) JEG-CAB enables the detection of HCys/Cys and (ROS/RNS) while (c) JEG-CAN enables the detection of HCys/Cys and NTR.We believed that CAH was a useful core unit that can be used to introduce the chosen reactive chemical trigger on the phenol for the detection of the corresponding analyte with HCys/Cys. Owing to the relationship between HCys and ONOO⁻/NTR, we set out on the development of a HCys AND ONOO⁻ probe and a HCys AND NTR probe.Therefore, we set out to prepare JEG-CAB and JEG-CAN, which are able to detect HCys/Cys and peroxynitrite (ONOO⁻) or nitroreductase (NTR), respectively (Scheme 1). For JEG-CAB, a benzyl boronate ester was introduced as a ONOO⁻ reactive unit.³⁷ For JEG-CAN, a p-nitrobenzyl group was installed as it is known to be an effective substrate for NTR.^38–40Open in a separate windowScheme 1Synthesis of target probe JEG-CAB and JEG-CAN.To afford CAH, compound 2 was synthesized by refluxing umbelliferone and acetic anhydride at 140 °C. Compound 2 was then dissolved in trifluoroacetic acid at 0 °C followed by the addition of hexamethylenetetramine (HMTA). The mixture was heated to reflux overnight and the solvent was then removed. The intermediate was then hydrolyzed in H₂O for 30 min at 60 °C. Upon isolating CAH, it was then alkylated using 4-bromomethylphenylboronic acid pinacol ester and K₂CO₃ in DMF at r.t. to afford JEG-CAB in 51% yield. JEG-CAN was produced by alkylating CAH using 4-nitrobenzyl bromide and K₂CO₃ in DMF at r.t. to give 49% yield (Scheme 1).We then evaluated the ability of JEG-CAB to detect ONOO⁻ ‘AND’ HCys in PBS buffer solution (10 mM, pH 7.40). The maximum absorption of JEG-CAB at 336 nm shifted to 323 nm with the addition of HCys and then slightly shifted to 328 nm following the addition of ONOO⁻ (Fig. S3^†). As shown in Fig. 2, JEG-CAB was initially non-fluorescent, but the addition of HCys (1 mM) led to a small increase in fluorescence intensity, the subsequent additions of ONOO⁻ (0–24 μM) led to a significant increase in fluorescence intensity (>17-fold, see Fig. S5^†). These results demonstrated the requirement for both ONOO⁻ ‘AND’ HCys to obtain a significant turn ‘‘on’’ fluorescence response.Open in a separate windowFig. 2Fluorescence spectra of JEG-CAB (15 μM) with addition of HCys (1 mM) and incubated for 40 min then measured. Followed by incremental additions of ONOO⁻ (0–24 μM). The data was obtained in PBS buffer solution (pH 7.40, 10 mM) at 25 °C. λ_ex = 371 (bandwidth 20) nm. Dashed line represents JEG-CAB and Hcys addition only. Blue line represents highest intensity after addition of ONOO⁻.The addition of HCys and ONOO⁻ were then performed in reverse where JEG-CAB exhibited a negligible increase in fluorescence intensity upon addition of ONOO⁻ (16 μM). However, in a similar manner to that shown in Fig. 2, a large increase in fluorescence intensity was produced after the subsequent addition of HCys (0–5.5 mM) (Fig. 3 and S6^†). LC-MS experiments were carried out to ascertain the reaction mechanism and the results confirmed the sequential formation of the thiazine ring in the presence of HCys followed by boronate ester cleavage in the presence of ONOO⁻ or vice versa (Scheme S2 and Fig. S19–S21^†).Open in a separate windowFig. 3Fluorescence spectra of JEG-CAB (15 μM) with addition of ONOO⁻ (16 μM) and followed by incremental additions of HCys (0–5.5 mM) measurements were taken after 40 min of both additions. The data was obtained in PBS buffer solution (pH 7.40, 10 mM) at 25 °C. λ_ex = 371 (bandwidth 20) nm. Dashed line represents JEG-CAB and ONOO⁻ addition only. Blue line represents highest intensity after addition of HCys.As expected, probe JEG-CAB was shown to have excellent selectivity with ONOO⁻ against other ROS in the presence of HCys (1 mM) (Fig. S9 and S10^†). Furthermore, JEG-CAB exhibited a high degree of selectivity towards a series of amino acids where only HCys and Cys led to a fluorescence response in the presence of ONOO⁻. This is due to the formation of stable five or six-membered thiazine rings (Fig. S7 and S8^†).³⁴We then evaluated the changes in the fluorescence of JEG-CAN with both HCys and NTR in PBS buffer solution (10 mM, pH 7.40, containing 1% DMSO). As shown in Fig. 4, addition of HCys led to a small increase in fluorescence intensity. However, subsequent addition of NTR (4 μg mL⁻¹) led to a large time dependant increase in fluorescence intensity. To ensure both analytes were required, NTR and NADPH was kept constant (4 μg mL⁻¹ and 400 μM respectively) resulting in a 3.4 fold fluorescence increase (Fig. 5). We attribute the large initial increase to background fluorescence of NADPH.⁴¹ NTR then facilitates reduction of the nitro group of JEG-CAN releasing the core probe CAHvia a fragmentation cascade (Scheme S3^†).^38,42 Subsequent addition of HCys (2.0 mM) led to a 2 fold increase in fluorescence intensity. Again, LC-MS experiments confirmed the proposed reaction mechanism (Fig. S22^†).Open in a separate windowFig. 4Fluorescence spectra of JEG-CAN (15 μM) with initial addition of HCys (2 mM) and incubated for 60 min. Followed by addition of nitroreductase (4 μg mL⁻¹) and NADPH (400 μM) and measured over 90 min in PBS buffer solution (pH = 7.40, 10 mM, containing 1% DMSO). λ_ex = 363 nm. Ex slit: 5 nm and em slit: 5 nm. Dashed line represents JEG-CAN and HCys addition only. Blue line represents highest intensity after addition of NTR.Open in a separate windowFig. 5Fluorescence spectra of JEG-CAN (15 μM) with initial addition of nitroreductase (4 μg mL⁻¹) and NADPH (400 μM) and incubated for 60 min. Followed by addition HCys (2 mM) and measured over 90 min in PBS buffer solution (pH = 7.40, 10 mM, containing 1% DMSO). λ_ex = 363 nm. Ex slit: 5 nm and em slit: 5 nm. Dashed line represents JEG-CAN and NTR addition only. Blue line represents highest intensity after addition of HCys.Kinetic studies for JEG-CAN with both NTR and HCys were carried out (Fig. S11–S18^†) where it is clear that JEG-CAN exhibits a dose dependant fluorescence increase in response of both HCys and NTR.Unfortunately, the probes failed to give good data in cells, which could be due to their short excitation wavelengths or the extremely low intracellular HCys concentrations (5–15 μM). We are now pursuing the development ‘AND’ logic fluorescence probes with longer excitation and emission wavelengths suitable for in vitro and in vivo applications.In summary, we have developed two coumarin-based ‘AND’ logic fluorescence probes (JEG-CAB and JEG-CAN) for the detection of HCys/Cys and ONOO⁻ or NTR, respectively. CAH is a useful platform that enables easy modification for the development of ‘AND’-based fluorescent probes for the detection of HCys/Cys and a second analyte. Both JEG-CAB and JEG-CAN were shown to be ‘AND’-based fluorescent probes.     

Coumarin-based fluorescent probe for the rapid detection of peroxynitrite ‘AND’ biological thiols

A coumarin-based novel ‘AND’ logic fluorescent probe ROS-AHC has been developed for the simultaneous detection of ONOO⁻ and biological thiols. ROS-AHC was shown to exhibit only a very small fluorescence response upon addition of a single GSH or ONOO⁻ analyte. Exposure to both analytes, however, resulted in a significant fluorescence enhancement.

A coumarin-based novel ‘AND’ logic fluorescent probe ROS-AHC has been developed for the simultaneous detection of ONOO⁻ and biological thiols.

Peroxynitrite (ONOO⁻) is a short-lived reactive oxygen and reactive nitrogen species (ROS and RNS) produced intracellularly by the diffusion-controlled reaction of nitric oxide (NO˙) with superoxide (O₂˙⁻).^1–3 Despite playing a key role as a physiological regulator,⁴ it is commonly known for its high reactivity towards most types of biomolecules, causing deleterious effects and irreversible damage to proteins, nucleic acids, and cell membranes.^5,6 ONOO⁻ is therefore a central biological pathogenic factor in a variety of health conditions such as strokes, reperfusion injuries or inflammatory and neurodegenerative diseases (Parkinson''s disease, Alzheimer''s disease).^7–9 Conversely, biothiols such as glutathione and cysteine are endogenous reducing agents, playing a central role in the intracellular antioxidant defence systems.^10–12 Glutathione (GSH), in particular, is the most abundant biothiol in mammalian cells, and exists as both its reduced GSH form, and as the oxidised disulphide form GSSG.^13–15 Peroxynitrite and biothiols such as GSH are intimately linked, as abnormal levels of highly oxidising ONOO⁻ can perturb the delicate GSH/GSSG balance, causing irreversible damage to key processes such as mitochondrial respiration.¹⁶ Thus, abnormal levels of GSH are common in cells undergoing oxidative stress, in which the regulation of and interplay between ONOO⁻ and GSH is closely associated with physiological and pathological processes.^17,18 One such example is drug-induced liver injury (DILI), in which upregulation of ONOO⁻ occurs in hepatotoxicity. Treatment with GSH could be used to remediate this type of cell injury by depletion of ONOO⁻.^19–22One of our core research interests lies in the development of dual analyte chemosensors capable of detecting two distinct analytes such as biological reactive oxygen species and biothiols.^23–26 Although a wide range of single-analyte probes exist for the detection of ROS and thiols separately,^27–30 ‘AND’ logic sensors for their simultaneous detection are still rare.^31–33 We are therefore interested in developing such probes, containing two distinct sensing units, one for each analyte, working simultaneously or in tandem to elicit a fluorescence response.³⁴ This approach allows the monitoring of multiple biomolecular events and factors involved in specific disease pathologies, in order to achieve optimal predictive accuracy for diagnosis and prognostication.³⁵Using these principles, our group has recently focused on developing a range of ‘AND’ logic based sensors exploiting a variety of sensing units and mechanisms of fluorescence. Two such probes are shown below: GSH-ABAH (Fig. 1a), an ESIPT probe with a 4-amino-2-(benzo[d]thiazol-2-yl)phenol (ABAH) core, employing a maleic anhydride thiol-acceptor group;³¹ and JEG-CAB (Fig. 1b), a coumarin-based probe, this time with a salicylaldehyde homocysteine-reactive unit.²⁴ Both of these sensors employ a benzyl boronate ester as their peroxynitrite-reactive unit.Open in a separate windowFig. 1(a) GSH-ABAH, previously reported probe for simultaneous detection of ONOO⁻ and GSH. (b) JEG-CAB, previously reported probe for simultaneous detection of ONOO⁻ and GSH. (c) AHC – a core fluorescent unit that enables the synthesis of ‘AND’ based fluorescent probe for the detection of ONOO⁻ and GSH (d) ROS-AHC, a novel probe detailed in this work for simultaneous detection of ONOO⁻ and GSH.Herein, we set out to develop an ‘AND’ logic gate based fluorescence probe for simultaneous detection of ONOO⁻ and GSH. 3-Amino-7-hydroxy-2H-chromen-2-one (AHC) was chosen as a suitable coumarin fluorophore core for the development of an ‘AND’ logic based sensor, as its free phenol and amine functional groups provided a good opportunity for independent derivatization (Fig. 1).^36–39Previous literature reports show that protection of AHC with a maleic anhydride group results in quenching of the coumarin''s fluorescence intensity due to photoinduced electron transfer (PeT) processes. This fluorescence is rapidly restored in the presence of biological thiols, however, due to their fast addition to this functional group.⁴⁰ Therefore, we suggested that functionalization of the free phenol of this sensor using a benzyl boronic ester should further block the fluorescence, whilst serving as reporter unit for ONOO⁻. The greatly increased reactivity of peroxynitrite over other ROS towards boronate esters^41–43 should allow this functionality to act as a peroxynitrite-selective reporter, leading to an ‘AND’ logic based probe for the detection of ONOO⁻ and biological thiols (Fig. 1, Scheme 1). ROS-AHC was synthesized in 5 steps, starting with a 4-step synthesis of compound 1 adapted from literature procedures,^40,44 followed by the addition of the benzyl boronic pinacol ester (see Scheme S1 ESI^†).Open in a separate windowScheme 1Fluorescence ‘turn on’ mechanism of ROS-AHC in the presence of ONOO⁻ and GSH.The UV-Vis behaviour of ROS-AHC before and after exposure to both GSH and ONOO⁻ was evaluated in pH 7.40 buffer solution, showing a maximum absorption peak at 340 nm for both the unreacted probe and the probe following exposure to GSH, shifting to 350 nm with the addition of ONOO⁻ to the probe and 365 nm after sequential additions of GSH and ONOO⁻ to the probe (Fig. S1 ESI^†). Fluorescence experiments were then carried out. As expected, ROS-AHC was initially non-fluorescent, with a small fluorescence increase upon addition of ONOO⁻ (6 µM) (Fig. 2 and S2 ESI^†). Incremental additions of GSH (0–4.5 µM) resulted in a much larger increase in fluorescence intensity (>69-fold, see Fig. 2 and S3 ESI^†), demonstrating the need for both GSH and ONOO⁻ in order to achieve a significant ‘turn on’ fluorescence response.Open in a separate windowFig. 2Fluorescence spectra of ROS-AHC (5 µM) with addition of ONOO⁻ (6 µM), wait 5 min then incremental addition of GSH (0–4.5 µM), 5 min incubation before measurements in PBS buffer solution (10 mM, pH = 7.40). Fluorescence intensities were measured with λ_ex = 400 nm (bandwidth 8 nm). The green line represents the highest intensity after addition of GSH (4 µM).Similar fluorescence experiments were then carried out in reverse order, with the addition of GSH (6 µM) to ROS-AHC resulting in only a small increase in fluorescence intensity (Fig. 3 and S4 ESI^†). As before, incremental addition of the second analyte, in this case ONOO⁻ (0–5.5 µM), resulted in a large increase in fluorescence intensity (>46-fold, Fig. 3 and S5 ESI^†), confirming that ROS-AHC requires both GSH and ONOO⁻ for a full fluorescence ‘turn on’ response.Open in a separate windowFig. 3Fluorescence spectra of ROS-AHC (5 µM) with addition of GSH (6 µM), wait 5 min then incremental addition of ONOO⁻ (0–5.5 µM) with 5 min incubation before measurements in PBS buffer solution (10 mM, pH = 7.40). Fluorescence intensities were measured with λ_ex = 400 nm (bandwidth 8 nm). The orange line shows the highest intensity after addition of ONOO⁻ (5 µM).Subsequently, the selectivity of this probe towards both analytes was evaluated. A range of amino acids were evaluated (Fig. S6 ESI^†), with only thiol-containing analytes (glutathione, cysteine and homocysteine) eliciting significant fluorescence response, whilst non-thiol amino acids led to no changes in fluorescence intensity. A broad screen of ROS analytes was also carried out, demonstrating excellent selectivity for ONOO⁻, even over H₂O₂ (Fig. S7 ESI^†).The time-dependent response of ROS-AHC with both ONOO⁻ and GSH was also examined (Fig. S8 and S9 ESI^†). After initial addition of GSH or ONOO⁻ to the probe, subsequent addition of the second analyte triggered a rapid and significant increase in fluorescence, achieving maximum fluorescence intensity within 78 s in both cases. Furthermore, LC-MS experiments confirmed the formation of the suggested non-fluorescent intermediates, as well as the final fluorescent species shown in Scheme 1 (Fig S10, S11 and S12^†).In summary, we have developed a coumarin-based dual-analyte ‘AND’ logic fluorescent sensor, ROS-AHC, for the simultaneous detection of ONOO⁻ and biological thiols. ROS-AHC has shown high sensitivity and selectivity towards both ONOO⁻ and biological thiols.     

Size effect studies in catalysis: a simple surfactant-free synthesis of sub 3 nm Pd nanocatalysts supported on carbon

Supported Pd nanoparticles are prepared under ambient conditions via a surfactant-free synthesis. Pd(NO₃)₂ is reduced in the presence of a carbon support in alkaline methanol to obtain sub 3 nm nanoparticles. The preparation method is relevant to the study of size effects in catalytic reactions like ethanol electro-oxidation.

A simple surfactant-free synthesis of sub 3 nm carbon-supported Pd nanocatalysts is introduced to study size effects in catalysis.

A key achievement in the design of catalytic materials is to optimise the use of resources. This can be done by designing nanomaterials with high surface area due to their nanometre scale. A second achievement is to control and improve catalytic activity, stability and selectivity. These properties are also strongly influenced by size.^1–3 To investigate ‘size effects’ it is then important to develop synthesis routes that ensure well-defined particle size distribution, especially towards smaller sizes (1–10 nm).Metal nanoparticles are widely studied catalysts. In several wet chemical syntheses, NP size can be controlled using surfactants. These additives are, however, undesirable for many applications^4,5 since they can block active sites and impair the catalytic activity. They need to be removed in ‘activation’ steps which can negatively alter the physical and catalytic properties of the as-produced NPs. Surfactant-free syntheses are well suited to design catalysts with optimal catalytic activity⁶ but their widespread use is limited by a challenging size control.³Palladium (Pd) NPs are important catalysts for a range of chemical transformations like selective hydrogenation reactions and energy applications.^7–9 It is however challenging to obtain sub 3 nm Pd NPs, in particular without using surfactants.² Surfactant-free syntheses are nevertheless attracting a growing interest due to the need for catalysts with higher performances.^10–14Promising surfactant-free syntheses of Pd NPs were recently reported.^8,15 The NPs obtained in these approaches are in the size range of 1–2 nm and show enhanced activity for acetylene hydrogenation⁸ and dehydrogenation of formic acid.¹⁵ Enhanced properties are attributed to the absence of capping agents leading to readily active Pd NPs. The reported syntheses consist in mixing palladium acetate, Pd(OAc)₂, in methanol and the reduction of the metal complex to NPs occurs at room temperature. The synthesis is better controlled in anhydrous conditions to achieve a fast reaction in ca. 1 hour. Another drawback is that the synthesis must be stopped to avoid overgrowth of the particles. Therefore, a support material needs to be added after the synthesis has been initiated and no simple control over the NP size is achieved.^8,15In this communication a more straightforward surfactant-free synthesis leading to sub 3 nm carbon-supported Pd NPs in alkaline methanol at ambient conditions is presented. A solution of Pd(OAc)₂ in methanol undergoes a colour change from orange to dark, indicative of a reduction to metallic Pd, after ca. a day. However, only ca. 1 hour is needed with Pd(NO₃)₂, Fig. 1 and UV-vis data in Fig. S1.^† The fast reduction of the Pd(NO₃)₂ complex in non-anhydrous conditions is a first benefit of the synthesis presented as compared to previous approaches.Open in a separate windowFig. 1Pictures of 4 mM Pd metal complexes in methanol without or with a base (as indicated).For particle suspensions prepared with Pd(OAc)₂ or Pd(NO₃)₂ the NPs agglomerate and quickly sediment leading to large ‘flake-like’ materials. When the reduction of Pd(NO₃)₂ in methanol is performed in presence of a carbon support and after reduction the solution is centrifuged and washed in methanol, a clear supernatant is observed indicating that no significant amount of NPs are left in methanol. Transmission electron microscope (TEM) analysis confirms that NPs are formed and well-dispersed on the carbon support surface and no unsupported NPs are observed, Fig. 2a. Likely, the reduction of the NPs proceeds directly on the carbon support. However, the size of the NPs is in the range 5–25 nm, which is still a relatively large particle size and broad size distribution.Open in a separate windowFig. 2TEM micrographs of Pd NPs obtained by stirring 4 mM Pd(NO₃)₂ in methanol and a carbon support for 3 hours, (a) without NaOH and (b) with 20 mM NaOH. Size distribution histograms are reported in Fig. S4.^† The same samples after electrochemical treatments are characterised in (c) and (d) respectively. Size distribution histograms are reported in Fig. S7.^†Assuming a ‘nucleation and growth’ mechanism, the NPs should become larger over time.¹⁶ But the reaction is so fast that by stopping the reaction before completion, size control is not achieved and unreacted precious metal is observed, Fig. S2.^† To achieve a finer size control and more efficient use of the Pd resources, a base was added to the reaction mixture, e.g. NaOH.³ In alkaline media, the formation of Pd NPs is slower; it takes ca. 60 minutes to observe a dark colour for a 5 mM Pd(NO₃)₂ solution with a base/Pd molar ratio of 10 in absence of a support, Fig. 1.Also in alkaline methanol, the NPs agglomerate over time in absence of a support material. However, if the alkaline solution of Pd(NO₃)₂ is left to stir in presence of a carbon support the desired result is achieved, i.e. Pd NPs with a significantly smaller size and size distribution of ca. 2.5 ± 1.0 nm, Fig. 2b. The NPs homogeneously cover the carbon support and no unsupported NPs are observed by TEM suggesting that the NPs nucleate directly on the carbon surface. Furthermore, the supernatant after centrifugation is clear, indicating an efficient conversion of the Pd(NO₃)₂ complex to NPs, Fig. S3.^† Furthermore, there is no need for an extra reducing agent as in other approaches, for instance in alkaline aqueous solutions.⁹The benefits of surfactant-free syntheses of Pd NPs for achieving improved catalytic activity have been demonstrated for heterogeneous catalysis.^8,15 Surfactant-free syntheses are also well suited for electrochemical applications where fully accessible surfaces are required for fast and efficient electron transfer. Several reactions for energy conversion benefit from Pd NPs. An example is the electro-oxidation of alcohols,⁷ in particular ethanol¹⁷ (see also Table S1^†).Previous studies optimised the activity of Pd electrocatalysts by alloying,^18–20 by using different supports^17,21–23 or crystal structures.^24,25 Investigating NPs with a diameter less than 3 nm was challenging.^2,26,27 The surfactant-free synthesis method presented here allows to further study the size effect on Pd NPs supported on carbon (Pd/C) for electrocatalytic reactions.In Fig. 3, results for ethanol electro-oxidation in 1 M ethanol solution mixed with 1 M KOH aqueous electrolyte are reported based on cyclic voltammetry (CV) and chronoamperometry (CA) with Pd/C catalysts exhibiting 2 significantly different size distributions. The electrode preparation, the measurement procedure and the sequence of electrochemical treatments are detailed in the ESI.^† In order to highlight size effects, we compare geometric and Pd mass normalized currents (Fig. 3a and c) as well as the oxidation currents normalized to the Pd surface area (Fig. 3b).Open in a separate windowFig. 3Electrochemical characterisation of carbon supported Pd NPs with 5–25 nm (grey) and 2.5 nm (dark) size in 1 M KOH + 1 M ethanol aqueous electrolyte. (a) 2^nd CV before chronoamperometry (CA), (b) current normalised by the electrochemically active surface area of Pd, (c) CA recorded at 0.71 V vs. RHE after 50 cycles between 0.27 and 1.27 V.It is clearly seen that based on the geometric current density, the smaller Pd NPs exhibit significantly higher currents for ethanol oxidation than the larger NPs. To differentiate if this observation is a sole consequence of the different surface area, the electrochemically active surface area (ECSA) has been estimated based on “blank” CVs (without ethanol) recorded between 0.27 and 1.27 V vs. RHE in pure 1 M KOH aqueous electrolyte and integrating the area of the reduction peak at ca. 0.68 V, Fig. S5.^† As conversion factor, 424 μC cm⁻² was used.²⁸Using this method, the smaller NPs with a size around 2.5 nm exhibit an ECSA of 92 m² g⁻¹ whereas the larger NPs with a size in the range 5–25 nm exhibit an ECSA of 47 m² g⁻¹, consistent with a larger size. Normalising the ethanol electro-oxidation to these ECSA values instead of the geometric surface area, Fig. 3b, still indicates a size effect. It is clearly seen that the smaller Pd NPs exhibit higher surface specific ethanol oxidation currents, in particular at low electrode potentials. Furthermore, a clear difference in the peak ratios in the CVs is observed. The ratio in current density of the forward anodic peak (j_f, around 0.9 V) and the backward cathodic peak (j_b, around 0.7 V vs. SCE) is around one for the smaller NPs, whereas it is about 0.5 for the larger NPs. The forward scan corresponds to the oxidation of chemisorbed species from ethanol adsorption. The backward scan is related to the removal of carbonaceous species not fully oxidised in the forward scan. The higher j_f/j_b ratio therefore confirms that the smaller NPs are more active for ethanol electro-oxidation and less prone to poisoning, e.g. by formation of carbonaceous species that accumulate on the catalyst surface.^29,30 This observation is further supported by a chronoamperometry (CA) experiment, Fig. 3c, at 0.71 V performed after continuous cycling (50 cycles between 0.27 and 1.27 V at a scan rate of 50 mV s⁻¹). In the CA testing of the thus aged catalysts at 0.71 V, the ethanol oxidation current on the two catalysts starts at around the same values, however, its decay rate is significantly different. The Pd mass related oxidation currents for the smaller NPs are after 30 minutes almost twice as high (ca. 200 A g_Pd⁻¹) as for the larger ones (ca. 130 A g_Pd⁻¹), confirming that the small Pd NPs are less prone to poisoning. In particular a factor up to 4 in the Pd mass related ethanol oxidation currents after 1800 s of continuous operation is achieved compared to a recently characterised commercial Pd catalyst on carbon,²⁰ Table S1.^† Despite different testing procedure reported in the literature, it can be concluded from these investigations that the surfactant-free synthesis presented shows promising properties for electrocatalytic ethanol oxidation even after extended cycling.The extended cycling, however, has different consequences for the two catalysts. For the small (2.5 nm) NPs of the Pd/C catalyst, a massive particle loss, but only moderate particle growth is observed as highlighted in Fig. 2 (see also Fig. S6^†). TEM micrographs of the two Pd/C samples recorded before and after the complete testing (CVs and CAs, for details see Fig. S7^†) show that the catalyst with small Pd NPs exhibits a pronounced particle loss as well as a particle growth to ca. 6 nm probably due to sintering. By comparison, for the Pd/C catalyst with the large Pd NPs, no significant influence of the testing on particle size or particle density is apparent.     

A simple two-photon turn-on fluorescent probe for the selective detection of cysteine based on a dual PeT/ICT mechanism

Herein, a simple two-photon turn-on fluorescent probe, N-(6-acyl-2-naphthayl)-maleimide (1), based on a dual PeT/ICT quenching mechanism is reported for the highly sensitive and selective detection of cysteine (Cys) over other biothiols. The probe was applied in the two-photon imaging of Cys in cultured HeLa cells, excited by a near-infrared laser at 690 nm.

N-(6-acyl-2-naphthayl)-maleimide (1) is a simple two-photon fluorescent probe with selectivity for cysteine, based on a thiol-Michael-addition-transcyclization cascade and dual PeT/ICT quenching mechanism.

Cysteine (Cys), homocysteine (Hcy), and glutathione (GSH) are structurally similar biothiols, but their biological functions are quite different from one another.^1–6 Among these biothiols, Cys functions as one of the twenty-one amino acids for peptide and protein synthesis, and Cys deficiency is also associated with certain disease symptoms.^7–10 Methods for the selective detection and differentiation of Cys among different biothiols, including high performance liquid chromatography (HPLC),¹¹ capillary electrophoresis,¹² electrochemical assay,¹³ UV-vis spectroscopy,¹⁴ and fluorescence-based methods,^15–17 are important for its biological studies. Recently, fluorescent probes have attracted much attention as vital chemical biology tools due to their high sensitivity, convenient operation, and real-time imaging capabilities.^18–20 A number of Cys-selective fluorescent probes have been reported,²¹ which utilize Cys-selective recognition groups such as aldehydes,^11,22 acrylates,²³ thioesters,²⁴ and electron-deficient aromatic halides^25–27 in their structures. However, many of them have relatively long response times and low sensitivity due to a slow cyclization process. In addition, fluorescent probes with high selectivity for Cys over Hcy are difficult to achieve because they differ by only one methylene group.²⁸ Recently, we reported that N-(N′-butyl-1,8-naphthalimide-4-yl)-maleimide, containing a single maleimide group as the recognition group, is a fast, sensitive, and selective fluorescent probe for Cys based on a dual photo-induced electron transfer (PeT) and photo-induced intramolecular charge transfer (ICT) quenching mechanism.²⁸ Different from many other maleimide-based fluorescent probes that only undergo a PeT mechanism,¹⁵ the additional ICT quenching mechanism keeps the 1,8-naphthalimide (NAP) fluorophore in the thiol-Michael adduct in a low fluorescence emission state due to the strong electron-withdrawing effects of the succinimide group at its 4-position. Then, a subsequent transcyclization step, involving the formation of a six-membered thiomorpholinone ring and cleavage of a five-membered succinimide ring, converts the non-fluorescent thiol-Michael adduct into the major fluorescent product, in which the ICT quenching is removed, resulting in a drastic fluorescence turn-on response.²⁸ A similar transcyclization process and the simultaneous removal of ICT quenching allowed us to design a NAP-based turn-on fluorescent probe for γ-glutamyltranspeptidase²⁹ and a coumarin-based turn-on fluorescent probe with dual recognition groups and dual cyclization for the selective detection of Cys.³⁰ In addition, another NAP-based dual PeT/ICT probe was recently reported by Meka and Heagy for the detection of hydrogen sulfide, although two recognition groups instead of one were adopted in their probe to achieve the dual quenching mechanism.³¹Our previous work and that of other groups has demonstrated that the combination of PeT and ICT mechanisms is particularly suitable for the design of fluorescent probes with a significant fluorescence turn-on response.^30–33 However, many of these probes have a short excitation wavelength in the UV or visible range, which is not optimal for biological applications due to enhanced phototoxicity and/or autofluorescence.^34,35 Considering that two-photon fluorescence imaging has advantages such as the excitation process being carried out by a near-infrared (NIR) laser that has a reduced cell toxicity and low fluorescence background,³⁶ in this work, we aimed to introduce a similar dual PeT/ICT quenching mechanism to the known two-photon fluorophore 6-acyl-2-naphthylamine^37–39 in order to design a simple maleimide-based two-photon fluorescent probe, 1, for the selective detection of Cys over Hcy and GSH. It was also tested to determine whether it is a turn-on fluorescent probe with high sensitivity and selectivity, which reacts with Cys via a fast two-step thiol-Michael addition and transcyclization cascade reaction.²⁸ The structure of probe 1 is shown in Fig. 1. It has a maleimide group at its 2-position, which promotes the PeT quenching effect. It also has an additional electron-withdrawing methylcarbonyl group at its 6-position to ensure a pull–pull ICT quenching effect.Open in a separate windowFig. 1Design rationale of the fluorescent probe 1 for the selective turn-on detection of Cys over Hcy and GSH.Probe 1 was conveniently synthesized from 6-acyl-2-naphthylamine (3)³⁹ in a two-step process with a total yield of 38% (see Scheme S1 in the ESI^†). First, the amine 3 was reacted with maleic anhydride to form the maleic amide acid 4. Then, the amide acid 4 was cyclized to afford the maleimide 1 in the presence of acetic anhydride (see the ESI^† for more details).We then investigated the absorption and fluorescence emission response of the probe towards just 1 equiv. of Cys. The time-dependent absorption spectra upon the addition of 1 equiv. of Cys are shown in Fig. 2a. Probe 1 has a maximum absorption peak at 292 nm. Upon addition of Cys, the maximum absorption peak shifts to 314 nm, a red-shift of 22 nm. Notably, an isosbestic point can be seen at 295 nm after 2 min, indicating the formation of an intermediate within 2 min, which is then converted into the final product. The UV spectral changes supported the presence of a proposed cascade reaction sequence for the fast formation of a thiol-Michael adduct intermediate, which then underwent a relatively slow intramolecular transcyclization process to give the final product. From time-dependent fluorescence emission studies (Fig. 2b), probe 1 was found to have almost no fluorescence emission due to dual PeT and ICT quenching effects. Upon the addition of 1 equiv. of Cys, a drastic turn-on fluorescence response (a >3000 fold increase) was observed at 446 nm (see Fig. S1b in the ESI^†). The fluorescence intensity at 446 nm reached its maximum value after around 30 min indicating that the cascade reaction finished in about 30 min (Fig. 2b, and S2a in the ESI^†). The pseudo-first-order reaction kinetic constant based on the fluorescence enhancement was calculated as 0.123 min⁻¹ (half-time = 5.64 min, Fig. S2b in the ESI^†), indicating an overall fast cascade reaction. Fluorescence titration experiments using an increasing amount of Cys from 0 to 4.0 equiv. over 30 min showed a steady increase in the fluorescence intensity and the maximum intensity was reached at exactly 1.0 equiv. of Cys. Further Cys addition did not increase the fluorescence intensity, indicating that probe 1 reacts with Cys in a 1 : 1 molar ratio (Fig. 2c and S3 in the ESI^†), which was also supported by the Job plot (see Fig. S4 in the ESI^†). From the linear relationship of the fluorescence intensity at 446 nm versus the Cys concentrations, the detection limit of probe 1 (2 μM) for Cys was calculated as 1.4 nM (S/N = 3, Fig. 3d), indicating that 1 is a highly sensitive probe for Cys. Moreover, the probe showed excellent selectivity for the detection of Cys over many other species (Fig. 2e, and S5 in the ESI^†), including the structurally similar thiols Hcy, GSH, and N-acetylcysteine (NAC). The fluorescence intensity at 446 nm for 1 equiv. of Cys was significantly higher (12.2-fold, 9.1-fold, and 17.7-fold, respectively) than that of 10 equiv. of Hcy, GSH, or NAC. To further confirm the reaction mechanism, the reaction product, 2, from the reaction of probe 1 with Cys, was isolated and its structure was confirmed using ¹H nuclear magnetic resonance (NMR) spectroscopy, ¹³C NMR spectroscopy, 2D-rotating-frame nuclear Overhauser effect spectroscopy (ROESY), and high-resolution mass spectrometry (HRMS) (see the ESI for more details^†). The fluorescence quantum yields of probe 1 and product 2 were measured as 0.002 and 0.782, respectively (see the ESI for more details^†). Therefore, the formation of the transcyclization product 2 was determined to be responsible for the observed fluorescence turn-on response. For the other thiols, the transcyclization steps of the thiol-Michael adducts were much slower, resulting in the observed high selectivity. Overall, we have shown here that probe 1 is a highly sensitive and selective turn-on fluorescent probe for Cys.Open in a separate windowFig. 2(a) Time-dependent UV-vis spectra of probe 1 (10 μM) upon the addition of 1 equiv. of Cys (a spectrum was recorded every 2 minutes); (b) time-dependent fluorescence emission spectra of probe 1 (2 μM) upon the addition of 1 equiv. of Cys (a spectrum was recorded every 3 minutes); (c) time-dependent fluorescence emission intensity at 446 nm of probe 1 (2 μM) upon addition of Cys (0 to 4 equiv.); (d) a linear relationship of the fluorescence intensity at 446 nm versus the Cys concentration (0.2–2.0 μM); (e) fluorescence response of probe 1 (2 μM) at 446 nm toward various species in PBS buffer (10 mM, pH 7.4): (1) blank; (2) Cys; (3) Hcy; (4) GSH; (5) NAC; (6) valine; (7) glycine; (8) isoleucine; (9) lysine; (10) leucine; (11) histidine; (12) asparagine; (13) methionine; (14) proline; (15) serine; (16) alanine; (17) threonine; (18) arginine; (19) glutamine; (20) aspartic acid; (21) glutamic acid; (22) tyrosine; (23) tryptophan; (24) phenylalanine; (25) glucose; (26) H₂O₂; (27) Na⁺; (28) K⁺; (29) Ca²⁺; (30) Mg²⁺; (31) Fe³⁺; (32) Fe²⁺; (33); Cu²⁺; (34) Zn²⁺ (All measurements were made in 10 mM PBS buffer, pH 7.4, 25 °C, and λ_ex = 314 nm).Open in a separate windowFig. 3Two-photon fluorescence images (b, e, h, k) of HeLa cells collected at 410–510 nm (blue to cyan-blue, λ_ex = 690 nm), the corresponding bright field view (a, d, g, j), and overlap of the fluorescence channel and the bright field view (c, f, i, l) after different treatments: (a–c) the cells were pretreated with 0.5 mM of N-ethylmaleimide (NEM) for 30 min and then incubated with 10 μM of probe 1 for 30 min; (d–f) cells were first pretreated with 0.5 mM of NEM for 30 min, then after addition of 1 mM of Cys were incubated for 30 min, and finally, incubated with 10 μM of probe 1 for 30 min (scale bar = 10 μm); the conditions for (g–i) and (j–l) were similar to those of (d–f), except that 10 μM of Hcy and 10 μM of GSH were used instead of 10 μM of Cys.Encouraged by the fast, selective, and sensitive in vitro fluorescence response of probe 1 for the detection of Cys, we further evaluated its potential use as a two-photon imaging agent for Cys in biological systems, such as in living cells. The fluorescence response of probe 1 towards Cys at different pH values was evaluated and a suitable pH range for Cys detection was determined to be 7.0 to 10.0, which is a good range for cell imaging applications because physiological conditions have a pH of around 7.4 (see Fig. S7 in the ESI^†). HeLa cells were then pretreated with N-ethylmaleimide (NEM, 0.5 mM) for 30 min to remove the endogenous cellular thiols, and incubated with Cys (1 mM), Hcy (1 mM), or GSH (1 mM), respectively for 30 min to increase the specific thiol levels. The samples were then further incubated with probe 1 (10 μM) for 30 min and were then washed with PBS buffer before two-photon fluorescence cell images and the corresponding bright-field view images were taken (Fig. 3(d–l)). Control images were also taken for samples pretreated with NEM (0.5 mM) and then incubated with probe 1 (10 μM) (Fig. 3a–c). Only cells pretreated with NEM and then Cys showed a distinctive blue fluorescence (Fig. 3e). The above cell imaging studies clearly demonstrated that probe 1 is capable of the selective detection and imaging of intracellular Cys over Hcy and GSH in living cells by two-photon fluorescence imaging with low background fluorescence interference.     

Insight into trifluoromethylation – experimental electron density for Togni reagent I

3,3-Dimethyl-1-(trifluoromethyl)-1,3-dihydro-1-λ3,2-benziodoxole represents a popular reagent for trifluoromethylation. The σ hole on the hypervalent iodine atom in this “Togni reagent” is crucial for adduct formation between the reagent and a nucleophilic substrate. The electronic situation may be probed by high resolution X-ray diffraction: the experimental charge density thus derived shows that the short intermolecular contact of 3.0 Å between the iodine and a neighbouring oxygen atom is associated with a local charge depletion on the heavy halogen in the direction of the nucleophile and visible polarization of the O valence shell towards the iodine atom. In agreement with the expectation for λ3-iodanes, the intermolecular O⋯I–C_aryl halogen bond deviates significantly from linearity.

The experimentally observed electron density for the “Togni reagent” explains the interaction of the hypervalent iodine atom with a nucleophile.

A “halogen bond” denotes a short contact between a nucleophile acting as electron density donor and a (mostly heavy) halogen atom as electrophile;^1,2 halogen bonds are a special of σ hole interactions.^3–5 Such interactions do not only play an important role in crystal engineering;^6–11 rather, the concept of a nucleophile approaching the σ hole of a neighbouring atom may also prove helpful for understanding chemical reactivity.The title compound provides an example for such a σ hole based reactivity: 3,3-dimethyl-1-(trifluoromethyl)-1,3-dihydro-1-λ3,2-benziodoxole, 1, (Scheme 1) commonly known as “Togni reagent I”, is used for the electrophilic transfer of a trifluoromethyl group by reductive elimination. The original articles in which the application of 1¹² and other closely related “Togni reagents”¹³ were communicated have been and still are highly cited. Trifluoromethylation is not the only application for hypervalent iodine compounds; they have also been used as alkynylating¹⁴ or azide transfer reagents.^15,16 The syntheses of hypervalent iodanes and their application in organofluorine chemistry have been reviewed,¹⁷ and a special issues of the Journal of Organic Chemistry has been dedicated to Hypervalent Iodine Catalysis and Reagents.¹⁸ Recently, Pietrasiak and Togni have expanded the concept of hypervalent reagents to tellurium.¹⁹Open in a separate windowScheme 1Lewis structure of the Togni reagent, 1.Results from theory link σ hole interactions and chemical properties and indicate that the electron density distribution associated with the hypervalent iodine atom in 1 is essential for the reactivity of the molecule in trifluoromethylation.²⁰ Lüthi and coworkers have studied solvent effects and shown that activation entropy and volume play relevant roles for assigning the correct reaction mechanism to trifluoromethylation via1. These authors have confirmed the dominant role of reductive elimination and hence the relevance of the σ hole interaction for the reactivity of 1 in solution by ab initio molecular dynamics (AIMD) simulations.^21,22 An experimental approach to the electron density may complement theoretical calculations: low temperature X-ray diffraction data of sufficient resolution allow to obtain the experimental charge density and associate it with intra- and inter-molecular interactions.^23–25 Such advanced structure models based on aspherical scattering factors have also been applied in the study of halogen bonds.^26–30 In this contribution, we provide direct experimental information for the electronic situation in Togni reagent I, 1; in particular, we analyze the charge distribution around the hypervalent iodine atom.Excellent single crystals of the title compound were grown by sublimation.^§ The so-called independent atom model (IAM), i.e. the structure model based on conventional spherical scattering factors for neutral atoms, confirms the solid state structure reported by the original authors,¹² albeit with increased accuracy. As depicted in Fig. 1, two molecules of 1 interact via a crystallographic center of inversion. The pair of short intermolecular O⋯I contacts thus generated can be perceived as red areas on the interaction-sensitive Hirshfeld surface.³¹Open in a separate windowFig. 1Two neighbouring molecules of 1, related by a crystallographic center of inversion. The short intermolecular I⋯O contacts show up in red on the Hirshfeld surface³² enclosing the left molecule. (90% probability ellipsoids, H atoms omitted, symmetry operator 1 − x, 1 − y, 1 − z).The high resolution of our diffraction data ^¶ for 1 allowed an atom-centered multipole refinement^33,34 and thus an improved model for the experimental electron density which takes features of chemical bonding and lone pairs into account. Fig. 2 shows the deformation density, i.e. the difference electron density between this advanced multipole model and the IAM in the same orientation as Fig. 1.^|| The orientation of an oxygen lone pair (blue arrow) pointing towards the σ hole of the heavy halogen in the inversion-related molecule and the region of positive charge at this iodine atom (red arrow) are clearly visible. Single-bonded terminal halides are associated with one σ hole opposite to the only σ bond, thus resulting in a linear arrangement about the halogen atom. Different geometries and potentially more than a single σ hole are to be expected for λ3-iodanes such as our target molecule, and as a tendency, the resulting halogen bonds are expected to be weaker than those subtended by single-bonded iodine atoms.³⁵ In agreement with these theoretical considerations, the closest I⋯O contacts in 1 amount to 2.9822(9) Å. This distance is significantly shorter than the sum of the van-der-Waals radii (I, 1.98 Å; O, 1.52 Å (ref. 36)) but cannot compete with the shortest halogen bonds between iodine and oxygen^37,38 or iodine and nitrogen.^39–41Fig. 1 and ​and22 show that the C_aryl–I⋯O contacts are not linear; they subtend an angle C10–I1⋯O1′ of 141.23(3)° at the iodine atom. On the basis of theoretical calculations, Kirshenboim and Kozuch³⁵ have suggested that the split σ holes should be situated in the plane of the three substituents of the hypervalent atom and that halogen and covalent bonds should be coplanar. Fig. 3 shows that the C_aryl–I⋯O interaction in 1 closely matches this expectation, with the next oxygen neighbour O1′ only 0.47 Å out of the least-squares plane through the heavy halogen I1 and its three covalently bonded partners C1, O1 and C10.Open in a separate windowFig. 2Deformation density for the pair of neighbouring molecules in 1; the dashed blue and red arrows indicate regions of opposite charge. (Contour interval 0.10e Å⁻³; blue lines positive, red lines negative, green lines zero contours, symmetry operator 1 − x, 1 − y, 1 − z).Open in a separate windowFig. 3A molecule of 1, shown [platon] along O1⋯C1, and its halogen-bonded neighbour O1ⁱ. Symmetry operator 1 − x, 1 − y, 1 − z.The Laplacian, the scalar derivative of the gradient vector field of the electron density, emphasizes local charge accumulations and depletions and it allows to assess the character of intra- and inter-molecular interactions. A detailed analysis of all bonds in 1 according to Bader''s Atoms In Molecules theory⁴² is provided in the ESI.^‡ We here only mention that the electron density in the bond critical point (bcp) of the short intermolecular I⋯O contact amounts to 0.102(5)e Å⁻³; we are not aware of charge density studies on λ3-iodanes, but both the electron density and its small positive Laplacian match values experimentally observed for halogen bonds involving O and terminal I in the same distance range.⁴³The crystal structure of 1 necessarily implies additional contacts beyond the short halogen bond shown in Fig. 1 and ​and2.2. The shortest among these secondary interactions is depicted in Fig. 4: it involved a non-classical C–H⋯F contact with a H⋯F distance of 2.55 Å.Open in a separate windowFig. 4C–H⋯F contact in 1; additional information has been compiled in the ESI.^‡ Symmetry operators i = 1 − x, 1 − y, 1 − z; ii = 1 − x, 1 − y, −z.The topological analysis of the experimental charge density reveals that this non-classical C–H⋯F hydrogen bond and all other secondary contacts are only associated with very small electron densities in the bcps. Table S8 in the ESI^‡ provides a summary of this analysis and confirms that the I⋯O halogen bond discussed in Fig. 1 and ​and22 represents by far the most relevant intermolecular interaction.The relevance of this halogen bond extends beyond the crystal structure of 1: Insight into the spatial disposition of electrophilic and nucleophilic regions and hence into the expected reactivity of a molecule may be gained from another electron-density derived property, the electrostatic potential (ESP). The ESP for the pair of interacting molecules in 1 is depicted in Fig. 5.Open in a separate windowFig. 5Electrostatic potential for a pair of molecules in 1 mapped on an electron density isosurface (ρ = 0.5e Å⁻³; program MoleCoolQt^44,45). Fig. 5 underlines the complementary electrostatic interactions between the positively charged iodine and the negatively charged oxygen atoms. One can easily imagine to “replace” the inversion-related partner molecule in crystalline 1 by an incoming nucleophile.The ESP tentatively obtained for a single molecule in the structure of 1 did not differ significantly from that derived for the inversion-related pair (Fig. 5), and even the results from theoretical calculations in the gas phase for an isolated molecule²⁰ are in good qualitative agreement with our ESP derived from the crystal structure. In the absence of very short contacts, polarization by neighbouring molecules only has a minor influence on the ESP. The experimentally observed electron density matches the proven reactivity for the title compound, and we consider it rewarding to extend our charge density studies on related hypervalent reagents.     

Determination of berberine in Rhizoma coptidis using a β-cyclodextrin-sensitized fluorescence method

In this work, a method for the determination of berberine in Rhizoma coptidis using β-cyclodextrin-sensitized fluorescence technology is established. Berberine is the main extract of Rhizoma coptidis, a medicinal material, which causes an envelope reaction with β-cyclodextrin to generate fluorescence sensitization. In the environment of its own aqueous extract, with 0.0065 mol L⁻¹ of β-cyclodextrin, a fluorescence excitation wavelength (λ_ex) of 345 nm and an emission wavelength (λ_em) of 540 nm were selected to avoid interference from other distractors. The fluorescent sensor for the detection of berberine exhibits a low limit of detection (3.59 × 10⁻⁹ mol L⁻¹) and a wide linear range from 2.7 × 10⁻⁷ mol L⁻¹ to 2.7 × 10⁻⁶ mol L⁻¹. Our sensor can be also used to detect berberine in real medicinal materials. The content of berberine in Rhizoma coptidis medicinal material was found to be 7.60% using this method with an average recovery rate of 99.5%. The result obtained by thin-layer chromatography with fluorescence detection was 7.61%, which is consistent with the result from the β-cyclodextrin sensitized fluorescence method. This method is simple and environmentally friendly with high sensitivity and good selectivity and gives reliable results, which is promising for practical application.

In this work, a method for the determination of berberine in Rhizoma coptidis using β-cyclodextrin-sensitized fluorescence technology is established.

Rhizoma coptidis¹ is the dried rhizome of Coptis chinensis (Chuan Lian), Coptis chinensis (Ya Lian) or Yunlian (Chuan Lian), and is an important traditional Chinese medicine commonly used in clinics. Rhizoma coptidis has the functions of clearing heat and dampness, purging fire and detoxifying.² Its active ingredients are isoquinoline alkaloids,³ such as berberine hydrochloride (also known as berberine), coptisine hydrochloric (also known as coptisine), palmatine hydrochloride (also known as palmatine), and jatrorrhizine hydrochloride (also known as jatrorrhizine). Berberine is the main effective component of Rhizoma coptidis. Antibacterial,⁴ antiviral,⁵ and anti-diabetic⁶ effects of berberine have been reported, as well as its use for the treatment of liver cancer .⁷ Thus, the content of berberine is critical to the quality control of Rhizoma coptidis.At present, many methods are used for the determination of berberine in Rhizoma coptidis, such as UV-vis spectroscopy, capillary electrophoresis with field amplified sample stacking, high-performance liquid chromatography, high-performance liquid chromatography-mass spectrometry, near-infrared spectroscopy, proton nuclear magnetic resonance and nuclear magnetic resonance proton spectroscopy.^8–14 However, there are no relevant reports on the systematic study of fluorescence spectroscopy for berberine. Fluorescence analysis is the direct measurement of the secondary light emission of a substance. Fluorometric analysis has the advantages of high sensitivity and good selectivity compared to photometric methods and is simple, rapid and inexpensive compared to chromatographic methods. It is gradually receiving increasing attention for the analysis of proprietary Chinese medicines.^15–18Cyclodextrins (CDs), as shown in Fig. 1, are cyclic oligosaccharides consisting of (α-1,4)-linked α-d-glucopyranose units. The primary hydroxyl group extends at one end and the secondary hydroxyl group extends at the other end. Because the primary hydroxyl group can rotate freely to cover a part of the ring, while the secondary hydroxyl group is relatively rigid, CDs are not cylindrical but slightly tapered. The glycoside oxygen atom with an unbonded lone pair of electrons in the glucose group points to the center of the molecule, which has a high density. As a result of these factors, the center of the molecule is hydrophobic and the surface is hydrophilic. As shown in Fig. 1(b), it can match the size and cavity of the molecule, and the hydrophobic guest molecules form an inclusion complex.¹⁹ Cyclodextrin can form an inclusion complex with dapoxyl sodium sulfonate (DDS). A large fluorescence enhancement of DSS was observed upon formation of the inclusion complex.^20,21 The formation of inclusion complexes by cyclodextrin has the potential for tuning the optical properties of other chromophoric molecules by varying the cavity size of the macrocyclic host molecule.²²Open in a separate windowFig. 1Molecular structure (a) and molecular shape (b) of CDs.Berberine itself has weak fluorescence but produces stronger fluorescence when it forms an inclusion complex with other substances. Berberine can form an inclusion complex with cucurbit⁷ uril and produce a highly sensitive fluorescence response.²³ Berberine can form an inclusion complex with β-CD and a new spectrofluorimetric method for the determination of berberine in the presence of β-CD was developed. The linear range was 1.00–4.00 μg mL⁻¹ with a detection limit of 5.54 ng mL⁻¹.²⁴Both the collision between the fluorescent molecules and the quenchers around the fluorescent molecules can cause the inactivation of the fluorescent molecules and produce non-radiation. When the fluorescent molecule berberine enters the hydrophobic cavity of β-CD to form the inclusion compound, β-CD plays a shielding role, making berberine reduce the non-radiation deactivation process and quenching process, thus improving the fluorescence quantum yield.^25,26 It has been proved that both ends of the berberine molecule are matched with the pores of β-CD molecules, and 1 : 1 and 2 : 1 type inclusion compounds can be formed, as shown in Fig. 2.¹⁹ The experiment found that β-CD had no fluorescence enhancement effect on other isoquinoline alkaloids, so based on the sensitization and selectivity of β-CD for berberine, a new fluorescence analysis method for the determination of berberine content in the Chinese medicine Rhizoma coptidis was proposed.Open in a separate windowFig. 2Molecular structure of berberine (a) and models of the β-CD–BH inclusion mixtures: 1 : 1 inclusion mixture (b), 1 : 1 inclusion mixture (c), 2 : 1 inclusion mixture (d).In this work, a method for the determination of berberine in Rhizoma coptidis using β-cyclodextrin-sensitized fluorescence technology is established. Berberine is the main extract of Rhizoma coptidis medicinal material, which causes an envelope reaction with β-cyclodextrin to generate fluorescence sensitization. The fluorescent sensor for the detection of berberine exhibits a low limit of detection (1.33 ng mL⁻¹) and a wide linear range from 0.1 μg mL⁻¹ to 1.0 μg mL⁻¹. Our sensor can be also used to detect berberine in real medicinal materials. The content of berberine in Rhizoma coptidis medicinal material was found to be 7.60% with this method and the average recovery rate was 99.5%. The result obtained by thin-layer chromatography with fluorescence detection was 7.61%, which is consistent with the result from the β-cyclodextrin-sensitized fluorescence method.     

Protein-activated transformation of silver nanoparticles into blue and red-emitting nanoclusters

Proteins are very effective capping agents to synthesize biocompatible metal nanomaterials in situ. Reduction of metal salts in the presence of a protein generates very different types of nanomaterials (nanoparticles or nanoclusters) at different pH. Can a simple pH jump trigger a transformation between the nanomaterials? This has been realized through the conversion of silver nanoparticles (AgNPs) into highly fluorescent silver nanoclusters (AgNCs) via a pH-induced activation with bovine serum albumin (BSA) capping. The BSA-capped AgNPs, stable at neutral pH, undergo rapid dissolution upon a pH jump to 11.5, followed by the generation of blue-emitting Ag₈NCs under prolonged incubation (∼9 days). The AgNPs can be transformed quickly (within 1 hour) into red-emitting Ag₁₃NCs by adding sodium borohydride during the dissolution period. The BSA-capping exerts both oxidizing and reducing properties in the basic solution; it first oxidizes AgNPs into Ag⁺ and then reduces the Ag⁺ ions into AgNCs.

Protein capping can trigger nanoparticle to nanocluster transformation at elevated pH.

Noble metal nanomaterials, especially silver (Ag) and gold (Au), have witnessed exceptional research exploration in the last couple of decades from both fundamental and application perspectives.¹ These nanomaterials mainly exist in two distinct size regimes with unique optical characteristics. Ultra-small nanoclusters (NCs) (size typically <3 nm) contain only a handful of atoms (few to hundred), while relatively large nanoparticles (NPs) may comprise thousands of atoms. NPs may display strong extinction (absorption or scattering) spectra in the UV-vis region but are generally non-fluorescent.² In contrast, metal nanoclusters (MNCs) exhibit bright emission but not so noteworthy absorption spectra.^3,4 The distinct optical characteristics of the two nanomaterials have been exploited in various applications. For example, metal nanoparticles (MNPs) are extensively used in photothermal therapy⁵ and imaging,⁶ while NCs are more suited in fluorescence imagining⁷ and sensing⁸ applications. A facile transformation between the two nanomaterials could enable us to combine the complementary optical properties in a single system. Moreover, the kinetics of transformation can provide insights on various intermediate processes like dissolution, etching and digestive ripening etc.^9–11Silver nanoparticles (AgNPs) and nanoclusters (AgNCs) are of particular interest, as it not only possess the intriguing physicochemical properties of MNPs and MNCs, but also feature unique properties pertaining to silver.^3,12,13 For example, metallic silver has been well known for its capability to prevent infection since the ancient times, while recent studies revealed that ultrasmall AgNCs exhibit even superior antibacterial properties towards a broad spectrum of bacteria.^13,14 Moreover, due to superior plasmonic properties and bright fluorescence, AgNPs and AgNCs are preferred over other metal nanomaterials.^15,16The fluorescence properties of AgNCs mainly be attributed to the quantum confinement effect or surface ligand effect.¹⁷ The strong fluorescence generally arises from the electronic transition between occupied d band and states above the Fermi level (sp bands) or the electronic transition between highest occupied molecular orbital (HOMO) and the lowest unoccupied molecular orbital (LUMO).¹⁸ Several reviews have been devoted for the fundamental understanding of the fluorescence origin of AgNCs.^17,19 Recently, it was demonstrated that aggregation-induced emission (AIE) may also contribute to the luminescence pathway of MNCs.^19,20 The origin of AIE from MNCs could be attributed to the restriction of intramolecular vibration and rotation of ligand on the surface of MNCs after aggregation, which facilitates the radiative energy relaxation via inhibiting of non-radiative relaxations.^21,22Protein capping is quite common for obtaining both NPs^23,24 and NCs.^25–29 Serum proteins, bovine serum albumin (BSA) and human serum albumin (HSA) are the most popular among trials with different proteins.^26–29 BSA is a large protein which provides steric stabilization to the MNCs with its various functional group like –OH, –NH₂, –COOH, –SH.^25,30 The disulfide bond of BSA may have strong interaction with the MNCs where sulfur may be covalently bonded to the MNCs core.^24,31 The nanomaterials are synthesized within the protein template at very different pHs. AgNPs are obtained from the reduction of silver salts at neutral pH (6–8),²⁴ whereas the same process at a higher pH (>11) leads to AgNCs.^30,32 The protein capping itself may reduce Ag⁺; AgNCs are formed without any external reducing agent.^25,33 However, an external reducing agent may change the nature and kinetics of the NCs significantly.³⁰Thus, the influence of pH on the protein structure may govern the selective synthesis of AgNPs or AgNCs. BSA can achieve several conformations – N (native), B (basic), A (aged) and U (unfolded) as the pH of the medium gradually changes from neutral to highly alkaline.^34,35 It may be possible that a specific type of nanomaterial is stable within a particular conformation dictated by the pH of the medium. Hence, by simply changing the pH, we may expect a significant modulation of the morphology of the nanomaterial. Herein, we applied this concept to show an effortless transformation from AgNP to AgNC. Although BSA template is exceptionally popular in the preparation of both AgNPs and AgNCs, however, to the best of our knowledge, no report is available on the conversion from AgNP to AgNC within the protein capping.The BSA-capped AgNPs (BSA-AgNPs) were first synthesized at a neutral pH (pH = 6) using sodium borohydride reduction (see ESI^†). The AgNPs show a sharp surface plasmon resonance (SPR) band at 415 nm (Fig. 1a) and have uniform diameters of 12.5 ± 1.5 nm (Fig. 1b). The AgNPs are quite stable at this pH with no apparent change in the SPR band even after 15 days (Fig. S1^†).Open in a separate windowFig. 1(a) UV-vis spectrum and (b) TEM image of BSA-protected AgNPs synthesized at pH 6. The insets show the appearance of the AgNP solution under regular and UV light (left panel), and size distribution histogram (right panel).However, when the BSA-AgNPs were treated with NaOH to elevate the pH to 11.5, we observed a remarkable decrease in the SPR band at 415 nm and a color change from dark to light brown within 2 h of the pH jump (Fig. 2a). The observations indicate the dissolution of AgNPs, which was further confirmed from the TEM images taken quickly (∼10 min) after the NaOH treatment (Fig. S2^†). Heterogeneous distribution of AgNPs was obtained with sizes varying from 2.6 nm to 17 nm, which is in sharp contrast to the uniform AgNPs before the addition NaOH (cf.Fig. 1b). Upon further incubation (6 h), the light brown color gradually faded to light yellow with a further decrease in the SPR band absorbance (Fig. S3^†).Open in a separate windowFig. 2Time evaluation of the (a) UV-visible and (b) emission spectra (λ_ex = 370 nm) of BSA-capped AgNPs after enhancement of the pH from 6 to 11.5 (by addition of NaOH at t = 0). The inset shows the snapshot of the final blue-emitting AgNC solution under normal and UV lights. (c) TEM of the blue-emitting AgNCs along with the HRTEM image and the analyzed size distribution in the inset. (d) MALDI-mass spectra of the BSA protein and BSA-capped blue-emitting AgNCs.Interestingly, the solution also develops distinct fluorescence with a maximum at ∼460 nm after the addition of NaOH (Fig. 2b). The fluorescence intensity gradually grows up upon incubation, and finally, an intense blue fluorescence was developed within ∼9 days. The final NaOH-treated AgNP solution appears to be light yellow under normal light and blue-fluorescent when viewed under a hand-held UV lamp (Fig. 2b, inset). The blue-emitting AgNCs exhibit a single band excitation spectrum with a maximum at 372 nm (Fig. S4^†).TEM image of the optimized NCs (after 9 days incubation at 37 °C) exclusively reveals uniform AgNCs of ∼2.10 ± 0.28 nm diameter without any trace of large NPs (Fig. 2c). The mass of the BSA-capped AgNCs (67 375 Da) was shifted by 845 Da from that of native BSA (66 530 Da) (Fig. 2d). Thus, the new species should correspond to Ag₈ cluster. The characteristics of the blue-AgNCs were quite similar to the human serum albumin (HSA)-protected blue-AgNCs, directly prepared from silver salt.³³ However, the formation time of those AgNCs was significantly less (∼10 h) than the present method (∼9 days).³³ Thus, the initial dissolution process, although quite fast, may have a crucial role in the kinetics of the protein-protected NCs. When we performed a similar pH jump experiment on a citrate-stabilized AgNP,³⁶ the extinction spectrum of the AgNPs showed much less variation compared to the BSA-AgNPs. Instead of a strong decrease, the SRP band showed a red-shift with an extended tail indicating aggregation rather than dissolution of NPs (Fig. S5^†).Furthermore, a red-emitting cluster was generated when an external reducing agent, sodium borohydride (NaBH₄), was added during the dissolution process. NaBH₄ was added after ∼11 min of the NaOH addition when the SPR band of BSA-AgNP was already decreased by half (Fig. 3a). The SPR band (λ_max = 415 nm) of AgNP continues to diminish similarly before and after the addition of NaBH₄ (Fig. S3^†). Thus, NaBH₄ may not have any significant effect on the dissolution process of AgNP. However, it has a strong impact on the modulation of the fluorescence; a new fluorescence band was developed at ∼650 nm within a much shorter duration (1 h) (Fig. 3b). The solution exhibits a bright-red fluorescence under a UV lamp (Fig. 3b, inset) with a quantum yield of 3.5%.Open in a separate windowFig. 3Early time evolution of (a) UV-visible and (b) emission spectra (λ_ex = 370 nm) of the BSA-protected AgNPs upon subsequent treatments with NaOH (pH 11.5) and NaBH₄ at t = 0 and 11 min, respectively. The decrease of the SPR band at 415 nm and a concomitant increase of the fluorescence band at ∼650 nm indicates dissolution of the AgNPs and formation of the red-emitting cluster. The inset shows the visuals of the AgNCs formed after 1 h under normal light and UV light. (c) TEM of the red-emitting AgNCs along with the HRTEM image and the analyzed size distribution in the inset. (d) MALDI-mass spectra of the BSA protein and BSA-capped red-emitting AgNCs.TEM measurements of the red-emitting species show homogeneous distribution of AgNCs with ∼2.25 ± 0.25 nm diameter (Fig. 3c). MALDI-mass experiment further assigned the red-emitting species as Ag₁₃ cluster (Fig. 3d). The excitation spectrum (λ_em = 650 nm) displays two distinct peaks at 370 nm and 470 nm, which match closely to the reported excitation peaks of the Ag_13–15 clusters within BSA/HSA capping (Fig. S4^†).^30,32,33 Moreover, the fluorescence decay of the red-emitting-AgNCs converted from AgNP almost matches with those prepared directly from AgNO₃; both display very similar average lifetimes (0.95 ns vs. 0.89 ns) (Fig. S6 and Table S1^†).³⁷Another important observation is that the red-emitting AgNCs have only transient stability at 37 °C. With further incubation, the absorbance at ∼470 nm (characteristic excitation peak of the red-emitting cluster) reduces and the absorbance at 370 nm (excitation peak of the blue-emitting cluster) increases simultaneously (Fig. 4a). The red-emission at 650 nm also decreases gradually with a concomitant increase of a blue emission band at 465 nm (Fig. 4b). Thus, both absorption and emission measurements clearly indicate transformation of red- to blue-emitting clusters which takes up to ∼15 days for completion. The solution finally becomes light yellow and exhibits a bright blue fluorescence under UV light similar to the blue-emitting cluster obtained earlier from the AgNP in the absence of NaBH₄. Interestingly, other characteristics of the regenerated blue-emitting AgNCs (converted from Ag₁₃NCs) also match quite nicely with the directly prepared blue-AgNCs (converted from AgNPs in the absence of NaBH₄). The size of this blue cluster was 2.04 ± 0.12 nm, which is similar to the previously obtained direct blue-emitting cluster (2.10 ± 0.28) (Fig. 4c). Furthermore, MALDI-mass measurement reveals that both the blue-emitting clusters may have the same composition, Ag₈ (Fig. 4d). In addition to this, the average lifetime (0.53 ns) of the blue-emitting AgNCs synthesized from AgNP agrees well to the average lifetime (0.40 ns) of the blue-emitting AgNCs converted from the red-emitting AgNCs (Fig. S7 and Table S2^†). However, the quantum yield (23%) of blue-emitting AgNCs, converted from red-emitting AgNCs, was higher than the quantum yield (18%) of the blue-emitting AgNCs, converted from AgNPs. Since, the emission characteristics of the blue and the red-emitting clusters nearly matches with earlier report, we expect that silver may be present in the zero oxidation state as determined in those studies.^30,31Open in a separate windowFig. 4Transformation of red-emitting to blue-emitting cluster: (a) UV-visible and (b) emission spectra (λ_ex = 370 nm) showing transformation of the BSA-protected red-emitting Ag₁₃NCs (obtained at 1 h) to blue-emitting AgNC upon prolonged incubation. Red and blue arrows respectively denote the decrease/increase of the red/blue cluster absorbance and emission intensity with time. The inset (b) shows a magnified wavelength region in 580–720 nm of the emission spectra. (c) TEM image of the blue-emitting silver nanocluster while its inset shows HRTEM image with size histogram of corresponding silver nanocluster. (d) MALDI-mass spectra of BSA and BSA-containing blue-emitting silver nanocluster synthesized from Ag₁₃NCs.Moreover, the atomic composition of the NCs can be also be estimated from the Jellium model using the equation^38,39E_em = E_Fermi/N^0.33where E_Fermi is the Fermi energy of the metal (Ag), E_em is the emission energy of the MNCs and N is the number of atoms constituting a MNC. Using the model equation, the number of silver atoms for the blue-emitting AgNCs can be predicted as 8.45 (∼8) Ag atoms which is a good agreement with our MALDI data (8 Ag atoms). However, the theoretical calculation estimated as N ∼ 24 for the red-emitting AgNCs, which is not in agreement with the MALDI data (13 Ag atoms). This is because of the well-known deviation of the Jellium model for higher number of Ag atoms in AgNCs because of increase in the electronic screening effects and the harmonic distortion in the potential energy well.¹⁹Although the red-emitting cluster is not very stable at the experimental condition (pH 11.5, 37 °C), it may be easily stabilized by lowering the temperature or pH. The fluorescence intensity of the red-emitting and blue-emitting cluster kept at 4 °C, was almost preserved for more than 15 days (Fig. S8^†). On the other hand, lowering the pH to 6, also inhibits the red to blue-cluster transformation (Fig. S9^†). The observations indicate that the red-blue transformation has a moderate activation barrier and the conversion may be governed by the change in the structure of the protein in the alkaline condition. Acidification of the solution can stop the transformation of the protein conformation and inhibits the process.From these observations, we may conclude that the conversion from NPs to NCs occurs in two steps. First, a rapid dissolution of AgNP occurs in the alkaline medium. The kinetics of the dissolution process can be monitored through a time-dependent decrease of the SPR band and the time constant was found to be ∼13 min (Fig. S10^†). Dissolution of AgNPs is an important issue and assumed to be the leading cause of toxicity of AgNPs in biological mediums.⁴⁰ The dissolution is commonly favored at a low pH but drastically inhibited at high pH.⁴¹ The swift dissolution of the BSA-protected AgNP observed here at a high pH (11.5) is unprecedented. Thus, the BSA capping may have an active role in the dissolution process. We comprehend that the oxidation power of protein may be activated in the basic medium.Organothiols (R-SH) are known to promote dissolution of AgNPs; R-SH progressively reacts with Ag atoms to form RS-Ag complex.⁴² Since cysteine is also an organothiol, it is expected to play an essential role in the dissolution of AgNPs. Gondikas et al. showed that excess cysteine could favor the dissolution process of AgNPs, whereas another amino acid, serine (S–H bond is replaced by O–H bond), has no effect.⁴³ Zang and coworkers showed that only the isolated or reduced cysteine in a protein has a dominant role in the dissolution of NPs.⁴⁴ Although BSA contains as many as 35 cysteine residues; 34 of them are involved in S–S bond formation and only a single cysteine is present in free form (S–H). Hence, the dissolution of AgNPs at neutral pH may be negligible.Most proteins rich in sulfur-containing residues (cysteine and methionine) may degrade in alkaline solution. Florence reported that about 5 of 17 S–S bridges in BSA may be cleaved in the presence of 0.2 M NaOH.⁴⁵ Thus, at higher pH, some disulfide bonds may be cleaved and more cysteine residues may participate in the dissolution of BSA-capped AgNPs.In the second step, Ag⁺ ions generated from the dissolution of AgNPs, can be reduced either by the protein capping itself or by an external reducing agent to form NCs (Scheme 1). The tyrosine residues may be responsible for the reduction of the metal ions to NCs.^25,33 At a pH, higher than the pKa (10.46) of tyrosine, the reduction capability of tyrosine is enhanced by deprotonation of the phenolic group.^25,33,46 Moreover, the addition of a strong reducing agent (e.g., NaBH₄) may lead to a faster reduction, which favors quicker nucleation and growth of Ag atoms forming the bigger NCs (Ag₁₃NCs). However, the large Ag₁₃NCs may not be adequately stabilized by the protein conformation at that condition and hence may transform into the more stable blue-emitting Ag₈NCs.Open in a separate windowScheme 1Schematic representation of the transformation of the BSA-capped AgNPs to blue- and red-emitting AgNCs.The conformation change of the protein capping during the conversion was also supported by the circular dichroism (CD) measurements (Fig. S11^†). The formation of AgNPs results in a negligible change in the protein conformation (Table S3^†). However, the formation of red Ag₁₃ cluster results in a substantial modification in the BSA conformation. The α helix content reduces from 57% to 49%, whereas coil randomness increases from 17% to 21% without a major change in the β sheet. Interestingly, blue-emitting Ag₈ cluster perturbed the conformation of the BSA to a much larger extent (Table S3^†). As the cysteine disulfide bond has a direct role on maintaining the folded conformation of BSA, its breaking may change the protein conformation. The addition of NaOH induces breaking of S–S bond, which leads to formation of AgNCs with subsequent change in protein secondary structure.In conclusion, we report an unprecedented fast dissolution of AgNPs through activation of the protein (BSA) capping by elevating the pH of the medium to 11.5. At higher pH, the disulfide bonds may be cleaved, and the free cysteine may activate the dissolution process. The protein capping also plays a crucial role in the formation of fluorescent nanocluster after the completion of the dissolution process. Thus, we explored multiple roles of the BSA capping – (1) a stable capping agent at neutral pH to stabilize the AgNPs (2) activates the dissolution process probably via oxidative dissolution of the AgNPs (3) adsorbing the nascent silver ions within its scaffold and (4) finally reducing them to fluorescent nanocluster.     

Construction of fluorescence active MOFs with symmetrical and conformationally rigid N-2-aryl-triazole ligands

1,4-Bis-triazole-substituted arene (NAT) was designed and synthesized for the construction of metal organic frameworks. Unlike the tri-phenyl analogs, which give a twisted conformation between three benzene rings due to the A-1,3 repulsion, the NAT-ligand gave the energetically favored co-planar conformation with the strong fluorescence emission. With this ligand, two new MOFs, NAT-MOF-Cd (2,3,4-c) and NAT-MOF-Cu (4-c), were successfully obtained with the structure confirmed by X-ray. With the six-coordinated Cd(ii) cluster, an interesting metal–ligand coordination and H-bonding hybridized porous polymeric structures were observed. In contrast, a typical Cu(ii) paddle wheel coordination was obtained with NAT and Cu, giving a new MOF structure with moderate stability in aqueous solution from pH 1–11 for 24 hours, which suggests a promising future for applications in fluorescence sensing and photocatalysis.

1,4-Bis-triazole-substituted arene (NAT) was designed and synthesized for the construction of fluorescent metal organic frameworks.

With large surface areas, diverse geometries, tunable pore sizes, and accessible functional sites, metal organic frameworks (MOFs) have shown exciting applications in various research areas, including gas molecule storage and separation, chemical catalysis, molecular sensing, luminescence, drug delivery, optical/electronic devices, etc.¹ From the design perspective, the geometry of the ligand is crucial for the overall network construction and its function.² Over the past decade, our group has been focusing on the investigations of 1,2,3-triazole derivatives for applications in chemical synthesis and material preparation.³ These efforts have led to the discovery of interesting new functions associated with certain triazole analogs. One particularly interesting building block is the N-2-aryl-1,2,3-triazole (NAT) moiety recently reported from our lab.⁴ Unlike the ubiquitous N-1 isomers, which were prepared from CuAAC (click chemistry), the NATs have shown excellent co-planar conformation between triazole and arene rings.⁵ As a result, NAT gives strong fluorescence while N-1 isomer shows almost no emission.⁶ Both the unique structure and interesting photo activity makes NAT interesting molecular building blocks in materials development.⁴ Herein, we report two new NAT based MOFs with moderate water (acid/base) stability and fluorescence emissions.Our interest in NAT-MOF synthesis was initiated by the realization of twisted conformation associated with the poly-arene system. It is well-known that spacing linkers are important in MOF design to alter pore size.⁷ Considering the required structure rigidity often required for stable MOF structure, poly-arenes are one very popular linker motif.⁸ However, as shown in Scheme 1A, the typical bi-aryl molecule could not adopt co-planar conformation due to the A-1,3 repulsion between the ortho-protons on adjacent benzene rings.⁹ As a result, two conformations could be adopted: (A) twisted (all three rings twisted) and (B) orthogonal (rings 1 and 3 are parallel and twisted the same angle with ring 2).¹⁰ Overall, it is impossible to have all three aryl rings lining up to form a planar conformation, which could be crucial to open the window for effective π–π stacking. In contrast, with a coplanar conformation, NAT could serve an interesting and ideal new ligand for the extended poly-arene linker MOF construction. Besides the structure novelty, two other important concerns for MOF construction are material stability and practical ligand synthesis for potential applications.¹¹ With low electron density, 1,2,3-triazole is very stable toward oxidative conditions.¹² Moreover, the N-2-substitution successfully avoid potential triazole ring opening through N₂ extrusion.¹³ We then put in our efforts to develop a practical synthesis of the triazole-arene ligands. After evaluating various protecting groups and reaction sequences, a general route was developed as shown in Fig. 1.Open in a separate windowScheme 1Tri-aryl linker in MOF construction: twisted conformation.Open in a separate windowFig. 1Synthesis of NAT ligands: (a) PMBBr (1.1 equiv.), K₂CO₃ (4 equiv.), MeCN, 60 °C, 96%; (b) p-B(OH)₂–C₆H₄–COOMe, 5% Pd(PPh₃)₄, K₂CO₃ (4 equiv.), 1,4-dioxane : H₂O(1 : 1), 100 °C, 97%; (c) TFA, 120 °C, 54%; (d) 1,4-phenylenediboronic acid (0.5 equiv.), Cu(OAC)₂ (1.5 equiv.), pyridine (2 equiv.), THF, 1 atm O₂, 70 °C, 50%; (e) KOH, MeOH : THF (1 : 1), 100 °C, 90%.The synthesis starts from commercial available 4,5-dibromo-triazole 1. Protecting triazole with PMB group followed by the Suzuki coupling gave 4,5-diaryl triazole in excellent overall yields. Deprotection of PMB group gave NH triazole 2, which was applied to copper mediated Ullman coupling to afford the tetra-esters. Saponification followed by recrystallization gave the tetra-acid NAT ligand. Overall this route could afford the desired NAT ligand in gram scale with five linear steps. The resulting NAT tetra-acid is fluorescence active both in solution and in a solid state as expected. Notably, this route could be easily applied to the coordination with other linkers (besides benzene) for the coordination of new functional groups into the central arene, which is currently under investigation in our lab. With tetra-acid NAT available, we explore the possibility of applying them in MOF construction through coordination with two typical cations, Cd²⁺ from group 12 and Cu²⁺ from group 11. Fortunately, both complexes were successfully obtained under typical MOF preparation conditions (see details in ESI^†). The X-ray crystal structures of both MOFs were successfully obtained.As shown in Fig. 2, NAT-MOF-Cd was prepared through solvothermal condensation with NAT ligand and Cd(NO₃)₂ in the mixture of DMF, EtOH, water, and HNO₃ (5 : 1 : 1 : 1) at 85 °C. The needle-like transparent crystal was obtained. X-ray crystal structure reveals the typical 7-coordinated Cd cluster from three carboxylates and one DMF (Fig. 2A). Interestingly, as a tetra-acid ligand in coordination with a six-coordinated Cd cluster, the NAT ligand shows two different coordination patterns. For NAT-1, both carboxylates on each end of the ligand coordinated with Cd²⁺, forming cycle I (9.5 Å × 13.7 Å) through binding with neighboring NAT-1. On the other hand, only one carboxylate on each side of NAT-2 ligand coordinates with Cd²⁺ (to satisfy the overall six-coordination Cd cluster). The NAT-2 binding with Cd-SBU not only links the cycle 1 but also produces metallocycle 2 with a larger size (23.2 Å × 33 Å). As expected, both NAT-1 and NAT-2 show good co-planar conformation with the dihedral angle <30° in both cases, which highlight the unique structural feature of NAT over poly-arenes. While the cycle-1 and cycle-2 form large network extension, it is overall a 2D layer structure (Fig. 2B). However, with the un-coordinated free COOH in NAT-2, a layer-by-layer hydrogen bonded metal organic framework was achieved. As a result, the overall 3D packing framework is composed of layer-by-layer H-bonding and vertical six-coordinated Cd(ii) cluster MOF along the b axis as shown in Fig. 2C. The H-bond between COOH of each layer (d_OH⋯O = 2.623 nm) is confirmed and perfectly lining up the open cavity, forming cylindrical pores in the vertical direction (Fig. 2D).Open in a separate windowFig. 2(A) coordination environments of both ligand and Cd(ii) ions; (B) two coordination patterns of NAT-1 and NAT-2; (C) 3D packing view of NAT-MOF-Cd along b axes with fitted pores; (D) layer-by-layer H-bonded framework of NAT-MOF-Cd.The topology of this new NAT-MOF-Cd is calculated to be 2,3,4-c net with stoichiometry (2-c)(3-c)2(4-c) while the point (Schlafli) symbol is {4·8²}2{4²·8²·10²}{8}. The PXRD of the crystal structure was uniform with simulated data (Fig. S3a^†). FT-IR of NAT-MOF-Cd (Fig. S5a^†) showed the disappearing of 2990 cm⁻¹ adsorption associated with carboxylic acid and the appearance of 1390 cm⁻¹ and 1590 cm⁻¹ signal associated with the symmetric and asymmetric stretching of carboxylates. Thermogravimetric analyses (TGA) revealed the good thermal stability of NAT-MOF-Cd up to 351 °C (no decompositions) and completely collapsed at around 456 °C (Fig. S6a^†).The successful synthesis of NAT-MOF-Cd confirmed our hypothesis that fluorescence active NAT ligand could be used as a novel co-planar linker to coordinate with secondary building units (SBU) for porous MOF construction. The three-dimensional H-bonded MOF structure is interesting molecular architecture. However, it raised the concern whether it will be stable in an aqueous solution where many applications are taking place. Soaking NAT-MOF-Cd in water for only 30 minutes caused a total collapse of MOF structures confirmed by PXRD (Fig. S4^†). To achieve more rigid MOF structures with this NAT, we put our attention to Cu²⁺. It is well known that Cu(ii) cations could coordinate with four carboxylates to form a paddle wheel SBU. The good match of tetra-acid and four-coordinated Cu makes NAT ligand ideal for MOF construction via Cu-SBU. After screening various conditions, we were pleased to identify the combination of NAT with Cu(NO₃)₂·3H₂O (1 : 2) in a mixture of DMF, EtOH, water, and AcOH (5 : 1 : 1 : 1) as the optimal conditions for MOF synthesis. The FT-IR spectra (Fig. S5b^†) of resulting MOF revealed the disappearance of carboxylic acid groups around 2983 cm⁻¹ and the symmetric and asymmetric stretching of carboxylate groups at 1394 cm⁻¹ and 1581 cm⁻¹. The cubic-shape blue crystal was obtained with the structure confirmed by X-ray.As revealed by X-ray, the NAT-MOF-Cu is constructed with a typical Cu(ii) paddle wheel SBU (Fig. 3A). Each paddle wheel is coordinated with carboxylates from four NAT with two DMF bound on the c direction (Fig. 3B). Similar to NAT-MOF-Cd, the ligand in NAT-MOF-Cu shows good co-planar conformation with the dihedral angle <30°. The overall packing structure with pores along the c axis of NAT-MOF-Cu is shown in Fig. 3C. Meanwhile, the topology of NAT-MOF-Cu is calculated to be 4-c net with the point (Schlafli) symbol of {4⁴·6²}. The PXRD pattern was consistent with the simulated curve of the crystal structure (Fig. S3b^†). Thermalgravimetric analyses confirmed the thermal stability of NAT-MOF-Cu that it could behave intact as crystal scaffold until 354 °C before completely decomposed at around 551 °C (Fig. S6b^†).Open in a separate windowFig. 3(A) Paddle wheel SBU of NAT-MOF-Cu; (B) coordination environments of NAT-MOF-Cu; (C) 3D packing view of NAT-MOF-Cu along c axes; (D) CO₂ sorption and pore volumes of NAT-MOF-Cu at 195 K.The porosity of both NAT-MOFs was identified by CO₂ adsorption at 195 K. The framework behaves reversible type-I isotherm adsorption features, in which gas molecules present sharp adsorption at relatively low pressure (P/P₀ < 0.1) and reach to a plateau at 131 cm³ g⁻¹ with NAT-MOF-Cu (Fig. 3D). The Brunauer–Emmett–Teller (BET) and Langmuir surface area were calculated to be 309 m² g⁻¹ and 436 m² g⁻¹ for NAT-MOF-Cu because of large pore volumes from Horvath-Kawazoe calculation mode and regular stacking. Gas adsorptions of CO₂ and N₂ for NAT-MOF-Cu at 273 K were also obtained (Fig. S7b^†). The maximum adsorption of CO₂ was 43 cm³ g⁻¹ and the value of CO₂/N₂ selectivity (Fig. S7d^†) was obtained as 44.2 by ideal solution adsorbed theory (IAST) with a good correlation factor (R² > 0.999). Gas adsorption including BET surface area (37 m² g⁻¹) and CO₂/N₂ selectivity (8.5) at 273 K of NAT-MOF-Cd was also measured. Detailed figures are provided in ESI (Fig. S7 and S8^†).The stability of both NAT-MOFs was evaluated. As demonstrated previously, with the layer-by-layer H-bond, NAT-MOF-Cd is not stable in protic solvents. In contrast, the NAT-MOF-Cu showed excellent stability. First, immersing NAT-MOF-Cu in various organic solvents, including MeOH, EtOH, THF, MeCN, DCM, m-xylene, and 2-propanol, at room temperature for 24 h showed no crystal decomposition based on crystal PXRD (Fig. 4A). Furthermore, stability in aqueous solution under different pH was also evaluated by soaking the MOF in the aqueous solutions. NAT-MOF-Cu showed almost no decomposition in aqueous media with pH from 1 to 11 (Fig. 4B). Further increasing to pH = 13 or reducing to pH = 0 did give complex decomposition over time. However, the ability to survive aqueous solution over a large pH range showed the good stability of this new NAT-MOF-Cu porous materials.Open in a separate windowFig. 4Solvent stability tests of NAT-MOF-Cu in (A) organic solution and (B) aqueous solution.In general, the formation of carboxylate MOF complexes could enhance ligand fluorescence intensity due to the locked conformation that prevents undesired excitation state relaxation. NAT-MOF-Cd showed enhanced fluorescence (Φ = 26%) compared with NAT ligand (Φ = 6.7%) in solid state (Fig. 5). Interestingly, NAT-MOF-Cu gave significant fluorescence quenching with the resulting MOF showed almost no emission (Φ < 0.1%). This result suggested plausible electron or charge transfers in the excitation state with this new MOF material. The exact mechanism is currently under investigation along with the potential application of this new photoactive MOF as sensor or photocatalysts. The results will be reported in due course. Nevertheless, the intrinsic photoactivity along with the co-planar conformation makes NAT a promising new ligand system for future porous material construction.Open in a separate windowFig. 5Solid state fluorescence emission of NAT and NAT-MOFs.In summary, we designed and synthesized 1,4-bis-triazole-substituted arene (NAT) as a new ligand for the construction of metal organic frameworks. Compared with poly-arenes system, NAT adopts co-planar conformation with the dihedral angle <30°. NAT-MOF-Cd (2,3,4-c) was formed by six-coordinated Cd(ii) clusters with interesting H-bonded MOF framework while NAT-MOF-Cu (4-c) was obtained with Cu(ii) paddle wheel SBU. NAT-MOF-Cu showed reasonable stability in wide pH range (1–11) and organic solvent for over 24 h. Photoluminescence properties were observed upon the formation of NAT-MOF, suggesting potential applications of this photoactive porous material through the new ligand design.     

A top-down design for easy gram scale synthesis of melem nano rectangular prisms with improved surface area

An unprecedented top-down design for the preparation of melem by 1 h stirring of melamine-based g-C₃N₄ in 80 °C concentrated sulfuric acid (95–98%) was discovered. The melem product was formed selectively as a monomer on the gram scale without the need for controlled conditions, inert atmosphere, and a special purification technique. The as-prepared air-stable melem showed a distinctive nano rectangular prism morphology that possesses a larger surface area than the melems achieved by traditional bottom-up designs making it a promising candidate for catalysis and adsorption processes.

A novel practical method for the gram scale preparation of melem possessing a nano rectangular prism morphology and improved specific surface area through a top-down depolymerization design was developed.

Triamino-s-heptazine or 2,5,8-triamino-tri-s-triazine known as “melem”, is a mysterious molecule and invaluable intermediate in the density of melamine rings to graphitic carbon nitride (g-C₃N₄) with a rigid heptazine structure with three pendant amino substituents.¹ Melem does not bear two of the strongest emission quenchers, namely C–H and O–H groups;² In this way it has unique optical properties³ and is known as an efficient metal-free luminescent material.⁴ High stability, the possibility for supramolecular self-assembly, tunable band gap, and an already rich physicochemical chemistry are some of the known properties for melem.^1b For this reason, melem has the potential to be used in photocatalysts, MOFs, COFs, electrochemistry sensors, flame retardants, TADF and related OLEDs, and liquid crystals.¹ The use of melem in solar hydrogen evolution⁵ and bioimaging⁶ is also known.Very few reports of its catalytic application are available, nevertheless, in recent years it has attracted much attention because of exploring its unique properties. Metal-free g-C₃N₄/melem hybrid photocatalysts have been used for visible-light-driven hydrogen evolution.⁷ Lei et al. used melem single crystal nanorods as a photocatalyst with modulated charge potentials and dynamics.⁸ Recently, Liu et al. improved the photocatalytic properties of carbon nitride for water splitting by attaching melem to Schiff base bonds.⁹ In another report, a promotion in photocatalytic activity was obtained by construction of melem/g-C₃N₄ vermiculite hybrid photocatalyst for photo-degradation of tetracycline.¹⁰ Lei et al. reported that H₂ evolution activity of melem derived g-C₃N₄ was 18 times higher than g-C₃N₄.¹¹ Melem was also utilized as a precursor for the preparation of rod-like g-C₃N₄/V₂O₅ heterostructure with enhanced sonophotocatalytic degradation for tetracycline antibiotics.¹² CO₂ cycloaddition into cyclic carbonates,¹³ non-sacrificial photocatalytic H₂O₂ production,³ water treatment,¹⁴ simultaneous reductions of Cr(vi) and degradation of 5-sulfosalicylic acid,¹⁵ are some of the catalytic applications of melem at various fields of sciences.The main protocol of preparing melem is the annealing of cyanamide, dicyanamide, or melamine, which requires precise temperature control under an inert atmosphere such as N₂ or argon. Just recently, the synthesis approaches for molecular s-heptazines as well as their applications and properties have been reviewed by Audebert et al.^1b Most of the reported methods do not lead to the preparation of pure monomer melem and are often mixed with its oligomers and polymerized derivatives,¹⁶ meanwhile the possibility of forming triazine oligomers or oligomers between melem and triazine cannot be precluded.^5,16c Complete polymerization of melamine at 500–550 °C leads to g-C₃N₄ and at 400–450 °C leads to melem-like derivatives,¹⁷ mostly a mixture of different products requiring careful attention during isolation and purification.⁵ Recently, Kessler and his colleague investigated the thermolysis of melamine, the formation of melem, and the formation of poly(triazine imide) from melem precursor via ionothermal as well as thermal condensation (conventional synthesis) as the back reaction of the melem condensation.¹⁸The growing demands for employing melem in new applications besides the serious problems in preparing pure samples necessitate the development of a simple and operational scale-up method that does not have any acute and controlled conditions.It is well-known that the polycondensation mode of g-C₃N₄ and consequently the chemical and thermal stability as well as texture properties strongly depend on the nitrogen rich precursors (cyanamide, dicyandiamide, urea, and melamine) as well as annealing temperature.¹⁹The interaction between the molecular precursors and/or intermediate compounds are critical factors.¹⁷ Due to some drawbacks associated with the g-C₃N₄ such as low electronic conductivity, a high rate of photogenerated electron–hole pairs, a low surface area, poor visible-light absorption, low quantum yield, and low solubility in almost all of the traditional solvents,²⁰ it has been subjected to various acid treatments, to promote its properties and photochemical activity.²¹ Various nanosheets with different properties and morphologies have been obtained depending on the precursor used, acid nature and concentration, as well as reaction temperature and time.²² However, the oxidation products such as cyameluric or cyanuric acids (Scheme 1) under high reaction temperatures and times have been reported.^21cOpen in a separate windowScheme 1The selective production of the monomer melem from melamine-based g-C₃N₄ presented in this work. Other molecules are possible decomposition and/or oxidation products of g-C₃N₄.Inspired by the previous reports to prepare the acidified g-C₃N₄, we started with melamine to synthesize the g-C₃N₄ by calcining at 550 °C under air,²³ followed by the treatment with H₂SO₄. Nevertheless, we discovered that stirring the melamine-based g-C₃N₄ at concentrated H₂SO₄ (95–98%) at 80 °C for a limited time (1 h), afforded selectively monomer melem in high yield (Scheme 1). Following the intercalation, chemical exfoliation, and protonation of nitrogen atoms of the g-C₃N₄ sheets at concentrated H₂SO₄,^22,24 the bridging C–NH–C groups between s-heptazine units breaks which releases the triamino-s-heptazine (melem) molecules as monomer (Scheme S1^†). Under these conditions the formation of oligomers was precluded because of the effective breaking of the bridging amino groups, however, the limited reaction time and moderate temperature prevented the tri-s-triazine ring-opening as well as the formation of the oxidation products such as cyameluric (or cyanuric) acids.^21c Thus, we developed a facile and easy gram-scale synthesis of melem from acidic depolymerization of melamine-based g-C₃N₄ with no need for controlled conditions, and inert atmosphere. The air-stable white powder was insoluble in most common solvents (H₂O, C₂H₅OH, CH₃OH, DMF, CH₃CN, acetone, etc.) and only dissolved in DMSO with a very limited solubility exactly like that reported for the isolated pure monomer melem.^5,25 A new and distinctive rectangular prism morphology with an improved surface area was detected for the as-prepared melem,^5,8,26 which makes our study even more unique and novel.^5,8,27 It is well known that both morphology and specific surface area play important roles in affecting the photocatalytic activity of semiconductors.^22,28 Thus, our study not only provides a novel practical method for the preparation of nanostructured monomer melem, but also paves a new pathway for increasing its surface area. The chemical structure and purity of the as-prepared melem were verified by the combination of different techniques including FT-IR, ¹H and ¹³C NMR, mass spectra, elemental analysis, XRD, XPS, DRS, and photoluminescence spectroscopy.FT-IR spectrum of g-C₃N₄ and the as-prepared melem are depicted in Fig. 1. While the peaks at 803 and 796 cm⁻¹ exhibited the vibrations of tri-s-triazine moieties in g-C₃N₄ and melem respectively, two intense bands at 1622 and 1471 cm⁻¹, consistent with those of monomer melem. The lack of obvious C–NH–C vibrations at around 1230 cm⁻¹ featured the absence or negligible amount of dimelem or further melem-oligomers in the product.^29a,5,8 In the region of NH-stretching frequencies, a spectrum characteristic of amides is observed: three diffuse absorption bands (3415, 3360, and 3106 cm⁻¹) indicate the presence of strong intermolecular hydrogen bonds and strong interaction between the amino-groups and the ring.^29a Inspection of the characteristic bands of melem presented in Fig. 1, no evidence for the formation of cyameluric acid or other oxidation products (Scheme 1) was detected.²⁹Open in a separate windowFig. 1FT-IR of the as-synthesized g-C₃N₄ and Melem.In the ¹³C NMR spectrum (Fig. S1^†), two signals at 165.8 and 156 ppm are assigned to carbon atoms adjacent to the amino groups and CN3 groups in heptazine rings, respectively.^1a,5 The ¹H NMR (Fig. S1^†) showed a sharp signal at 7.4 ppm assigned to six protons of the terminal amino groups of melem along with two weak broad signals at ∼8 ppm which can be attributed to the partial protonation of some nitrogens. The lack of the signal at 149.37 ppm in ¹³C NMR^29b and a high-field signal at ¹H NMR (10.9 ppm or higher)³⁰ strongly confirmed that our method precludes the formation of cyameluric acid accompanied by the desired melem.The mass spectrometry depicted in Fig. S2^† shows that the bulk material contains almost entirely monomer melem evidenced by the main peak at m/z 218 pertinent to a single unit of melem and a very little peak at m/z 419 corresponding to dimelem and nothing of higher mass.³¹ Also, no trace of the oxidation products was observed in mass spectra (m/z 129 and 221 for cyanuric and cyameluric acids, respectively).The C/N atomic ratio is one of the most significant clues to prove the successful formation of melem. The ratio of 0.605 found for the produced melem is very close to the theoretical value in the monomer melem (C/N = 0.6).^5,8A substantial evidence for the exclusive formation of monomer melem was achieved by the XRD pattern. Fig. 2 shows XRD patterns of melem and its polymeric graphitic carbon nitride used in this work. A great match with literature was observed.⁵ Two characteristic peaks of g-C₃N₄ at 2θ = 13.28° (100) and 27.47° (002) related to the in-plane structural packing motif, and interlayer-stacking of aromatic systems respectively, significantly changed after treatment with 80 °C concentrated sulfuric acid for 1 h and showed strong evidence for the formation of the monomer melem.^5,8 The former peak (100) became pronounced and shifted to a lower angle of 12.52°, while the latter one (002) was shortened in the melem and shifted to the higher angle of 2θ = 27.6° caused by decreased stacking distance between the melem inter-layers. More important is the emergence of a new intense peak at 6.16°, which is the unique characteristic of monomer melem,⁵ while, other weak peaks located at about 19, 23, 25, 29 and, 31 are almost looked at in the XRD patterns of both monomer and oligomers.^5,8 No trace of cyameluric acid as the possible oxidation product was detected in the XRD pattern of the resulting product.^21c,30Open in a separate windowFig. 2XRD patterns of g-C₃N₄ and the as-synthesized melem.Next, XPS was used to identify the chemical environments of the product as shown in Fig. 3 and S3.^† Only C, N, and trace amounts of O and S caused by the negligible remaining sulfuric acid and water can be detected (Fig. S3^†). The C1s signals (Fig. 3 left) can be fitted into five components with binding energies of 284.5 eV, 285.18 eV, 287.78 eV, 288.58 eV, and 293.58 eV. The C signal at 284.5 eV is exclusively assigned to carbon atoms (C–C bonding) in a pure carbon environment, such as graphitic or amorphous carbons.^26,32 The signals at 285.16 and 287.78–288.3 eV attributed to graphitic carbon sp² C–C, and the sp² trigonal C–N bonding (s-triazine ring), respectively, characteristic of melem structure.^8,33 The advent of a high-energy satellite at 293.7 eV corresponds to the Π-electron delocalization in the heptazine system of melem.²⁷ The N 1s signals (Fig. 3 right) were deconvolved into five peaks. The signals with binding energies of 398.4, 400, and 401–404 eV are associated with the sp²-hybridized nitrogen (C N–C), tertiary nitrogen (N–(C)₃), and protonated amino groups (C–N–H) in melem, respectively.^33a,34 The emergence of (N–(C)₃) undoubtedly indicated the preservation of tri-s-triazine units (C₆N₇, basic part of melem molecule) during treatment with 80 °C concentrated acid. Thus, no significant changes in the carbon nitride heterocycles such as the oxidation transformation of terminal C–NH–C to C–OH–C and/or tri-s-triazine ring-opening reactions occurred.^21c,36 The advent of a satellite at high binding energy of 406 eV corresponds to the partial protonation of some nitrogens (N–H⁺).³⁵Open in a separate windowFig. 3XPS spectra of the as-prepared melem, left: C 1s and right: N 1s.The morphology of the as-synthesized product was determined by FESEM (Fig. 4A). The FESEM images clearly show microsized rectangular prisms with thickness ranging from ∼50 to 350 nm, which was completely different with carbon nitride with the main nanosheets morphology.³⁷ To the best of our knowledge, this is the first report for such a morphology for melem,⁸ that aroused our curiosity to assess its surface properties. The porosity of the samples was determined by N₂ physisorption experiments. The N₂ adsorption/desorption isotherms and pore size distributions of the as-prepared melem are given in Fig. 4B. The sample exhibited typical type IV isotherms with H3 hysteresis loop according to the IUPAC classification,²⁷ suggesting mesoporous structures with slit-shaped pores resulting from the aggregation of plate-like particles.³⁸ The BET specific surface area of the as-synthesized melem was found to be 19.54 m² g⁻¹ which is about 3–4 folds larger than those reported for bulk melems as 5.63 m² g⁻¹,²⁷ and 7.02 m² g⁻¹,¹² as well as melem nanorods as 4.87 m² g⁻¹,⁸ obtained from the condensation of melamine. These results clearly show the superiority of our easy-to-make melem over the other samples obtained by the traditional bottom-up design under quite controlled conditions.^5,8,27 The mesoporous nature of the as-synthesized melem was further supported by the pore-size distribution analysis depicted as an inset of isotherm (in Fig. 4B) indicating an average diameter of pore size at 2.1 nm.Open in a separate windowFig. 4(A) FESEM image and (B) BET N₂ adsorption/desorption isotherms of the as-synthesized melem.The TG analysis of the as-synthesized melem exhibited three mass loss steps (Fig. S4^†). At the first step, the sample lost about 10% of its weight at less than 200 °C caused by removing water and ethanol molecules absorbed during the elution process. The second one was begun at around 240 °C and continued to 500 °C with the evolution of ammonia and small amounts of HCN, attributed to the condensation polymerization of the monomer.The third thermal decomposition was accelerated above 500 °C (with releasing HCN and C₂N₂),⁶ rendering strong evidence for the absence of triazine derivatives (or lower) in the as-prepared product and once again ruled out the tri-s-triazine ring-opening reactions during the synthesis of melem in this work.^21c,39 The high thermal stability of the produced melem,⁶ is comparable with the parent g-C₃N₄, making it more appropriate for comparative studies and applied goals that add further benefits to our sample.Lastly, the optical properties of the sample were evaluated using UV-Vis diffuse reflectance spectroscopy (DRS). As shown in Fig. S5,^† the absorption maximum wavelength of the resulting melem locates at 310 nm coincides with that reported in the literature.^5,40 The band edge of melem shifted to the lower wavelength (380 nm) compared to the polymer g-C₃N₄ (460 nm), caused by decreasing in Π-electron delocalization in the heptazine system of melem which stretches the band gap from 2.7 eV (polymer) to 3.45 eV (monomer melem) in excellent agreement with reported theoretical value for monomer melem (3.497 eV).^5,27 Further support for this claim was obtained by fluorescence spectra. Fig. 5 shows the comparative fluorescence spectra of the as-synthesized melem, under 355 nm light excitation. As shown in Fig. 5, it is found that the fluorescence emission of polymer g-C₃N₄ peaked at 476 nm,^40a shifted to 412 nm in the melem coincide with Ricci report (415 nm).⁴¹ In addition, the photoluminescence intensity of the resulting melem increased significantly compared to polymer g-C₃N₄ in broad agreement with literature indicating that the condensation of melem to g-C₃N₄ causes the weaker photoluminescence.^40aOpen in a separate windowFig. 5Photoluminescence spectra of g-C₃N₄ and the as-synthesized melem under 355 nm light excitation.Finally, our formulation is very simple and robust with respect to processing conditions to overcome the potential scale-up problems to make it operational and amenable to scalability readily. As an example, a 5 fold semi-scaled-up procedure using 1.0 g g-C₃N₄ led to the isolation of the related pure monomer melem in 95% yield within 1 h.In summary, we developed a novel operational protocol for easy gram scale preparation of air-stable monomer melem through a top-down synthesis design with no need for any control conditions and further purification. Our analyses ruled out the presence of the starting polymer as well as the formation of oligomers and oxidation products in the final product highlighting the selectivity of the method toward the monomer of melem. The distinctive nano rectangular prism morphology with desired surface area and thermal stability, as well as the appropriate photoluminescence property qualifies our synthesized melem for applied goals and makes it a promising alternative for catalysis and adsorption processes which is under investigation in our lab.     

Rhodamine-based cyclic hydrazide derivatives as fluorescent probes for selective and rapid detection of formaldehyde

We describe fluorescent probes to detect formaldehyde (FA) in aqueous solutions and cells. The probes rapidly respond to FA in aqueous solutions and have great selectivity toward FA over other biologically relevant analytes. The results of cell studies reveal that probe 1 can be utilized to monitor endogenous and exogenous FA in live cells.

We developed a fluorescent probe that is useful to monitor endogenous and exogenous formaldehyde in live cells.

Reactive carbonyl species (RCS) produced through metabolic processes are highly reactive and, thus, their overproduction causes damage to a variety of organisms.¹ Formaldehyde (FA), the simplest RCS, is a human toxin and carcinogen as a result of its ability to crosslink DNA and proteins.² FA is generated in cells by several metabolic events, including methanol oxidation, histone demethylation and N-methylamine deamination.³ During normal metabolic processes, the concentration of FA is maintained at physiological levels in the range from 100 μM in blood to 200–400 μM in brain,⁴ where it is involved in spatial memory formation and cognition.⁵ However, upregulation of FA-producing enzymes or exposure to exogenous FA (e.g., industrial pollutants, cigarette smoke, and natural products) can lead to abnormal elevation of FA levels up to as much as 800 μM.^4,5 Elevated levels of FA cause memory impairments, cancers, diabetes and neurodegenerative disorders.⁶ Owing to the physiological and pathological significance of FA, selective and sensitive tools to monitor this RCS in cells are in critical demand.Fluorescence imaging is a powerful method to detect intracellular analytes (e.g., ions, reactive species and biomolecules) owing to its advantageous features such as operational simplicity, sensitivity and non-invasive properties.⁷ Several fluorescent probes for detection of FA in cells, which are based on specific chemical reactions including 2-aza-Cope rearrangement, formimine reaction and aminal formation, have been devised thus far.⁸ However, most of these probes have drawbacks such as low selectivities over other aldehydes and/or slow fluorescence responses to FA.To develop FA-responsive fluorescent probes that do not suffer from the limitations described above, we designed rhodamine-based cyclic hydrazide derivatives 1 and 2 (Scheme 1), which should be weakly fluorescent owing to the absence of an appropriate fluorophore. We reasoned that the tethered amine groups in 1 and 2 would react with FA to form iminium ions A, which would then undergo intramolecular addition of the hydrazide NH to generate cyclic aminals B. We also envisaged that rapid opening of spirocyclic moiety in B would take place to generate highly fluorescent xanthenes C.⁹Open in a separate windowScheme 1The proposed mechanism of fluorescence sensing of formaldehyde by rhodamine cyclic hydrazide-based probes 1 and 2.On the basis of this strategy, the new FA-reactive fluorescent probes 1 and 2 were synthesized using reactions of rhodamine B acid chloride with the corresponding amine-appended hydrazines (Schemes S1–S3^†). All new compounds were characterized using standard spectroscopic methods. To assess the design strategy displayed in Scheme 1, we measured time-dependent changes in the intensities of fluorescence arising from the probes following treatment with FA at a physiologically relevant concentration (10 equiv., 100 μM) in PBS buffer (1% DMSO, pH 7.4).⁴ As shown in the spectra and plots (Fig. 1a and S1^†), the secondary amine tethered probe 1 underwent an immediate fluorescence response to FA and the emission intensity reached a maximum within 5 min. In the case of probe 2 containing a primary amine appendage, the fluorescence intensity promoted by treatment with FA reached a plateau after 2 min but a lesser extent than that from 1 (Fig. S2^†). The pseudo-first-order rate constants for the fluorescence-monitored reactions of 1 and 2 with FA were determined to be k = 1.8 × 10⁻² M⁻¹ s⁻¹ and 4.6 × 10⁻² M⁻¹ s⁻¹, respectively (Fig. S3^†).¹⁰ The FA-concentration dependencies of reactions of probes 1 and 2 in aqueous buffer were determined by measuring fluorescence intensities, 10 min after treatment of the probes with 10 equiv. FA. The results showed that 1 and 2 exhibit a respective 15- and 6.5-fold enhancement in fluorescence intensity after addition of 50 equiv. FA (Fig. 1b and S4^†).Open in a separate windowFig. 1(a) Time-dependent change of the fluorescence spectra of 1 (10 μM) promoted by addition of 10 equiv. FA in PBS buffer (1% DMSO, pH 7.4) at 37 °C (λ_ex = 520 nm). Inset is a plot corresponding to the time-dependent increase in fluorescence intensity of 1 following addition of FA (λ_ex/λ_em = 520/583 nm). (b) FA concentration-dependent changes of the fluorescence spectra of 1 (10 μM), 10 min after each addition of FA. Inset is a plot corresponding to the FA concentration-dependent increase in fluorescence intensity of 1 (10 μM). (c) Color and fluorescence (FI) images of probe 1 in the absence and presence of FA.The selectivity of fluorescence responses of the probes toward FA was then assessed by individually treating 1 and 2 (10 μM) with various biologically relevant analytes, including reactive carbonyl species (FA, acetaldehyde, benzaldehyde, 4-hydroxybenzaldehyde, ethyl pyruvate, ethyl glyoxalate, propionaldehyde, glucose), reactive oxygen species (H₂O₂, HOCl, NO˙, ¹O₂, O₂˙⁻, ˙OH) and cations (Cu²⁺, Fe²⁺, Fe³⁺, K⁺, Zn²⁺). The results showed that both probes respond to FA but not to the other analytes (Fig. 2 and S5^†). Taken together, the above findings indicate that probes 1 and 2 respond rapidly and selectively to FA in aqueous buffer.Open in a separate windowFig. 2Change of fluorescence intensity of 1 (10 μM) at 583 nm (λ_ex = 520 nm) promoted by addition of each of biologically relevant analytes (10 equiv.) in PBS buffer (1% DMSO, pH 7.4) at 37 °C. Numbering of the analytes in the graph is as follows: 1, acetaldehyde; 2, benzaldehyde; 3, 4-hydroxybenzaldehyde; 4, ethyl pyruvate; 5, ethyl glyoxalate; 6, propionaldehyde; 7, d-glucose (1 mM); 8, H₂O₂; 9, HOCl; 10, NO˙; 11, ¹O₂; 12, O₂˙⁻; 13, ˙OH; 14, Cu²⁺; 15, Fe²⁺; 16, Fe³⁺; 17, K⁺; 18, Zn²⁺; 19, FA.To shed light on the mechanistic basis for the responses of probes to FA, the product generated by reaction of 1 and FA was isolated (see ESI^† for the detailed procedure) and characterized by using spectroscopic methods. Analysis of the ¹H and ¹³C NMR spectra of the isolated product in CDCl₃ suggests that a 1,2,4-triazinane ring system exists, as judged from chemical shifts that correspond to bridging methylene protons (CH₂) and carbon (3.24 ppm (s, 2H) and 72.5 ppm, respectively) (Scheme 2). Also, the spectral analysis suggests that the isolated product contains a spirocyclic ring system because of the presence of a signal at 61.3 ppm in the ¹³C NMR spectrum, which is expected for a quaternary carbon in a spiro ring. Furthermore, analysis of UV-Vis absorption and fluorescence spectra revealed that the product in CH₂Cl₂ displays very weak absorbance at 560 nm as well as very weak fluorescence at 583 nm (Fig. S6^†). These observations led us to conclude that the product in CH₂Cl₂ has the spirocyclic structure represented by 3 (Scheme 2).Open in a separate windowScheme 2Solvent (nonpolar organic solvent versus aqueous buffer) dependence of the equilibrium between ring-closed (3) and open (4) forms of the product generated by reaction of 1 with FA.In contrast, the isolated product in aqueous buffer (1% DMSO, pH 7.4) had a strong absorbance at 560 nm and intense fluorescence at 583 nm (Fig. S7^†), spectral properties that are quite similar to those of the substance generated by treatment of 1 with FA in aqueous buffer. Collectively, the results suggest that while the product of the reaction of 1 with FA exists in the ring-closed form 3 in nonpolar organic solvents, in aqueous buffer it exists in the ring-opened xanthene containing form 4 (Scheme 2). As a consequence, it is reasonable to conclude that the reaction responsible for fluorescence generation when 1 is treated with FA in aqueous buffer involves formation of 4. In addition, extinction coefficients (ε₅₁₄), quantum yields and fluorescence outputs (quantum yield × ε₅₁₄) of 1, 2 and 3 were determined (Table S1^†). Furthermore, the limits of detection of 1 and 2 for FA were calculated to be 1.24 μM and 0.59 μM, respectively (Fig. S8^†).Next, the utility of 1 to image FA in live cells was evaluated. Because 1 displayed a larger increase in fluorescence intensity upon treatment with FA than does 2, 1 was utilized in the cell studies described below. To determine the optimal conditions for cell imaging, HeLa cells (human cervical cancer cells) were incubated with 10 μM 1 for various times and with various concentrations of 1 for 1 h. Analysis of confocal fluorescence microscopy images showed that cells exposed to 10 μM 1 for 0.5–1 h display the intense fluorescence signal (Fig. S9^†). In addition, based on the results of an MTT assay as well as the observation of an intact nucleus morphology, 1 had negligible cell death activity under these treatment conditions (Fig. S10^†).We also probed the FA concentration-dependence of the fluorescence response of 1. For this purpose, HeLa cells were first treated with 10 μM 1 and then incubated with concentrations of FA (0–1.0 mM) that are in a physiologically relevant concentration range (ca. 400 μM in normal cells and up to 700–800 μM in cancer tissues).^4,11 The results showed that fluorescence signals arising from 1 in cells increase gradually as the FA concentration increases (Fig. 3), indicating the ability of 1 to serve as a probe for FA in live cells.Open in a separate windowFig. 3Detection of FA in cells using probe 1. HeLa cells were incubated with 1 (10 μM) for 1 h followed by treatment with several concentrations of FA for 1 h. Cell images were obtained using confocal fluorescence microscopy (scale bar = 10 μm). The nucleus was stained with Hoechst 33342. The graph shows normalized fluorescence intensity (FI) in the treated cells (mean ± s.d., n = 3).To evaluate the detection of endogenous FA in cells, several cell lines, including HeLa, MRC-5 (human fibroblast cell line derived from normal lung tissue), HaCaT (human keratinocytes), HEK293T (human embryonic kidney cells) and MCF-7 cells (human breast cancer cells), were incubated with 1 for 1 h. Analysis of cell images revealed that whereas the treated HEK293T cells display the low fluorescence intensity,⁸ⁱ the other four cell lines exhibit similarly strong fluorescence (Fig. 4 and S11^†). The findings indicate that while HEK293T cells produce a low level of FA, the other four cells generate high levels of FA.Open in a separate windowFig. 4Detection of endogenous FA in cells using probe 1. MRC-5, HaCaT, HEK293T, HeLa and MCF-7 cells were incubated individually with 1 (10 μM) for 1 h. Cell images were obtained using confocal fluorescence microscopy (scale bar = 10 μm). The nucleus was stained with Hoechst 33342. The graph shows fluorescence intensities in the treated cells (mean ± s.d., n = 3).It is known that FA is generated in cells by the actions of several demethylases and oxidase enzymes.³ The enzyme lysine-specific demethylase 1 (LSD1) catalyzes the removal of one or two methyl groups from modified lysines to produce free lysine and FA.^3,12 Also, it is known that GSK-LSD1 serves as a potent inhibitor of LSD1.¹³ As a result, production of FA by LSD1 in cells was evaluated by incubating MCF-7 cells with 1 in the absence and presence of GSK-LSD1. The results revealed that the intensity of the fluorescence arising from probe 1 in MCF-7 cells is slightly attenuated when GSK-LSD1 is present (Fig. 5 and S12^†).^8b The findings suggest that LSD1-promoted generation of FA in cells does not occur at high levels in comparison to the amounts formed by several other metabolic events. Taken together, the findings demonstrate that the rhodamine-based probe 1 is capable of detecting endogenous and exogenous FA in live cells.Open in a separate windowFig. 5The effect of an inhibitor on LSD1-promoted generation of FA in cells. MCF-7 cells were incubated with 1 μM GSK-LSD1 for 20 h followed by incubation with 1 (10 μM) for 1 h. Cell images were obtained using confocal fluorescence microscopy (scale bar = 10 μm). The nucleus was stained with Hoechst 33342. The graph shows normalized fluorescence intensities in the treated cells (mean ± s.d., n = 3).In conclusion, we have developed novel rhodamine-based cyclic hydrazide derivatives as fluorescent probes for the detection of FA in both aqueous media and live cells. Upon addition of FA to the probes in aqueous buffer, a fluorescence enhancement occurs within a few minutes. In addition, the probes respond to FA but not to other biologically relevant species, indicating that they have a high selectivity toward FA. Furthermore, the results of cell studies demonstrate that probe 1 can be employed to image exogenous and endogenous FA in live cells. As a result, this probe should be useful in efforts aimed at gaining a more detailed understanding of FA-associated biological processes.     

Colour-tunable ultralong organic phosphorescence upon temperature stimulus

A new class of single-component molecular crystal with colour-tunable ultralong organic phosphorescence (UOP) was designed and synthesized through alkyl chain engineering. Forming a more rigid environment at 77 K, the colour-tunable UOP from yellow-white to blue-green is achieved through dual-emission of crystal and amorphous states.

A new class of single-component molecular crystal with colour-tunable ultralong organic phosphorescence (UOP) was designed and synthesized through alkyl chain engineering.

Ultralong phosphorescence, a kind of phosphorescence that can be observed by the naked eye after removing the excitation source, has received great attention in the fields of sensing,¹ displays,² imaging,³ anti-counterfeiting⁴ and so on during the past years. Unfortunately, it is limited to inorganic materials because of the weak spin–orbit coupling (SOC) and strong non-radiative transition of pure organic materials.⁵ Compared with inorganic materials, however, organic materials have some excellent merits, such as inexpensive cost, relative safety to the environment, and soft preparation conditions.⁶ The realization of UOP becomes very significant. As mentioned above, UOP can be achieved by promoting intersystem crossing (ISC) through enhancing SOC and suppressing non-radiative transitions. In view of these factors, various strategies such as polymerization,⁷ H-aggregation,⁸ crystallization,⁹ host–guest doping,¹⁰ and freezing conditions¹¹ have been explored to achieve UOP with the unremitting efforts of scientific researchers.Intelligent-response organic luminescent materials can change their luminescent properties such as colour, lifetime, intensity, etc. after being stimulated by the external factor, such as mechanical forces, temperature, pH, light, solvent, etc., which have caught great attraction. The sensitivity of luminescent molecule to excitation wavelength, temperature or oxygen can be applied in sensors, optical recording and so on.^12–14However, the luminescence of these materials with intelligent-response are mostly limited to fluorescence or room temperature phosphorescence (RTP) with relatively short lifetimes.^15–18 Few samples display persistent luminescent feature.¹⁹ Most organic compounds can only tune their afterglow properties by changing molecular side groups or multi-component doping.^20–23 Therefore, the development of single component UOP materials with intelligent response remains a challenge. Inspired by the alkyl-chain engineering²⁴ and freezing conditions, herein we speculate that temperature response UOP might be induced by controlling the activity of the alkyl chain to regulate non-radiative transition rate. By means of reducing the temperature to restrict the molecular motions at amorphous state, colour-tunable UOP with temperature-response can be realized by dual-emission of molecules at both amorphous and crystalline states.In our previous study, MCzT crystals showed yellow UOP at RT.²⁵ The crystals showed the same UOP at 77 K with that at RT (Fig. 1a). Here, 9-(2-((4,6-dimethoxy-1,3,5-triazin-2-yl)oxy)ethyl)-9H-carbazole (MTOD) was designed and via alkyl chain attaching carbazole with a triazine core. The target molecule was characterized by ¹H NMR and ¹³C NMR (Fig. S1–S3^†). Its melting point reaches 130 °C. (Fig. S4^†) MTOD achieved a lifetime of up to 860 ms under ambient conditions. Surprisingly, the UOP changed from yellow-white to blue-green after the removal of the UV-lamp for several seconds at 77 K (Fig. 1b), demonstrating colour-tunable property of UOP with a low temperature stimulus.Open in a separate windowFig. 1Molecular structures and UOP photographs of MCzT (a) and MTOD (b).In order to explore the reasons for the colour-tunable UOP of MTOD at low temperature. The photophysical properties of MTOD in the crystal state were first investigated under ambient conditions. As shown in Fig. 2a, the photoluminescence (PL) spectrum of MTOD shows two main emission peaks at 375 and 413 nm and a shoulder at 435 nm. From the lifetime decay profiles (Fig. S5a^†), it was confirmed that they were all assigned to the fluorescence. Notably, the crystals of MTOD presented yellow afterglow after turning off the UV lamp. From the delayed phosphorescence spectrum, the main emission of MTOD was located at 414, 556 and 600 nm with lifetimes of 681.91, 860.56 and 860.59 ms, respectively (Fig. 2b and S5b^†). Among these, the emission around 414 nm is assigned to triplet–triplet annihilation (TTA) fluorescence originating larger π–π overlaps of carbazole groups from the crystal data (Fig. S6^†).²⁶ Remaining two emission peaks are attributed to UOP emission. From the phosphorescence excitation-emission spectra of MTOD, UOP can be efficiently excited from 260 to 380 nm, with the optimal excitation at 360 nm (Fig. S7^†).Open in a separate windowFig. 2Photophysical properties of MTOD at room temperature and at 77 K. (a) Steady-state photoluminescence (PL, blue dashed line) and phosphorescence (red solid line) spectra at RT and 77 K. Inset: photographs taken after removing excitation. (b) Time-resolved phosphorescence decay of the emission bands at 556 and 600 nm at room temperature, respectively. (c) Time-resolved phosphorescence decay of the emission bands at 478, 564 and 612 nm at 77 K, respectively. (d) PL (blue dashed line) and phosphorescence (red solid line) spectra at molten state.Subsequently, we measured the PL and phosphorescence spectra of MTOD at 77 K (Fig. 2a). The steady-state PL spectrum showed four peaks at 361, 376, 410 and 436 nm, with little change compared with its corresponding spectrum at RT. However, a new phosphorescence peak appears at about 480 nm with an intense emission than others, which displays an ultralong lifetime of 2.5 s. Obviously, the new emission peak plays an indispensable role in the colour-changed UOP at 77 K. The yellowish-white UOP observed by naked eyes at 77 K is generated by the combination of three phosphorescence peaks. However, as time goes by, only the blue-green afterglow at about 480 nm due to the shorter lifetime of long-wavelength phosphorescence can be observed.To find out the origin of this new peak at 480 nm, the PL and phosphorescence spectra of MTOD at molten state were measured as amorphous emission. As shown in Fig. 2d, the PL peak is located at 380 nm and the phosphorescence spectrum shows a broad band with a main peak at 484 nm. It is suggested that the new phosphorescence emission peak at 77 K may be attributed to amorphous state. Taken together, we deduce that the colour-tunable property of the crystal is possibly due to the presence of an amorphous state at low temperature.The X-ray single crystal diffraction of MTOD crystal was taken to explore the mechanism of UOP at room temperature. Abundant intermolecular and intramolecular interactions (Fig. 3a, S6, S8 and S9a^†) in the crystals strongly restrict the torsional molecular configuration. The dihedral angle between the triazine and carbazole groups is about 75° (Fig. S10^†). In the crystal of MTOD, the single molecule is limited by multiple intermolecular interactions, including C–H⋯N (2.681 Å), π-H⋯π (2.791, 2.876 Å), π-H⋯N (2.703, 2.711 Å) (Table S3^†). The rich intermolecular interactions are beneficial to limit molecular motions to suppress non-radiative transitions of excited molecules, leading to UOP. However, the amorphous molecules around the crystals with weak restriction displayed negligible phosphorescence at room temperature due to the strong motions of alkyl chains.Open in a separate windowFig. 3Intermolecular interactions (a) at room temperature and (b) at 100 K.Comparatively, the single crystal of MTOD molecule at 100 K was measured in order to explore the colour-changing mechanism of UOP at low temperature (Fig. 3b and S9b^†). By comparison, MTOD crystals exhibit more intermolecular interactions at 100 K and the distance become shorter. Molecular conformation of MTOD changed slightly, the dihedral angle between triazine and carbazole changed from 74.95° to 75.32° (Fig. S10^†). These increased interactions can constrain the molecules more effectively and the stronger restriction of alkyl chain and carbazole will further suppress non-radiative transitions, resulting in the much longer phosphorescence lifetime of over 1.0 s at low temperature. Compared with the molecules in the crystalline state, freezing condition can provide a more rigid environment to minimize the movement of the alkyl chain, greatly reducing the non-radiative transition rate at amorphous state, resulting in the lifetime of short wavelength phosphorescence at 478 nm up to 2.5 s.According to the above results, the photophysical process of colour-tunable phosphorescence can be described by Jablonski diagram as shown in Fig. 4. Upon photoexcitation, both electrons in amorphous and crystalline molecules transforms to lowest singlet states (S₁). Then, the electrons in S₁ would further transform to the lowest triplet (T₁) through ISC. At room temperature, amorphous molecules show strong molecular motions to facilitate the non-radiative transitions. However, crystalline molecules due to closely arrangement can exhibit phosphorescence through radiative decay. At 77 K, both excitons in amorphous and crystal states are dominated by radiative transitions, leading to colour-tunable UOP.Open in a separate windowFig. 4(a) Jablonski diagram of the relevant photophysical processes illustrating amorphous and crystalline UOP process at room temperature (top) and 77 K (bottom). (b) Phosphorescence spectra of MTOD at different temperature and (c) corresponding coordinates in CIE.In view of the interesting luminescent phenomenon, we have investigated a series of phosphorescent spectra of MTOD crystals at different temperatures ranged from 183 to 273 K. As shown in Fig. 4b, as the temperature increases, the phosphorescence intensity of the amorphous molecules gradually decreases. The colour variations of the MTOD crystals in response to the environmental temperatures are shown in the Commission International de l’Eclairage (CIE) coordinate diagram (Fig. 4c). As the temperature was gradually changed from 183 to 273 K, the UOP changed from green to yellow with good linearity of the CIE coordinates. This demonstrated that MTOD crystals may have potential in low temperature sensing.In conclusion, we synthesized a colour-tunable single-component UOP compound through alkyl chain engineering. Combined the spectral and single crystal analyses, it is indicated that colour-tunable UOP comes from dual-emission of molecules at amorphous and crystalline states. Low temperature provides better rigid effect on UOP of amorphous molecules than crystals, resulting in the UOP colour changed from yellow-white to blue-green. More interestingly, red-shifted UOP of MTOD crystals with the increase of temperature can be achieved, demonstrating its potential for temperature sensing. This study will provide a platform for the design of single-component UOP molecules with tunable colour emission and broadens its application field.     

Mesoporous-silica-coated palladium-nanocubes as recyclable nanocatalyst in C–C-coupling reaction – a green approach

We report the straightforward design of a recyclable palladium-core–silica-shell nanocatalyst showing an excellent balance between sufficient stability and permeability. The overall process – design, catalysis and purification – is characterized by its sustainability and simplicity accompanied by a great recycling potential and ultra high yields in C–C-coupling reactions.

A green approach: in a single-step coating process a mesoporous silica shell was tailored onto palladium-nanocubes. Along with a PEG-matrix this core–shell-nanocatalyst could be recovered after C–C-coupling reactions and reused without any significant decrease in product yield.

In contrast to bulk materials, metal-nanocyrstals (NC) possess unique physical and chemical properties. Both, nano-scale and geometry can dictate their optical, electronic and catalytic behavior.^1–3 Consequently, nanomaterials have been implemented already in various fields like medicine,⁴ sensing,⁵ nano-electronics⁶ and organic synthesis.⁷ Smart strategies for a sustainable usage of limited resources such as precious metals and hydrocarbons are inevitable due to the consistently growing population and economy.^8–10 Generally, catalysis gives rise to novel and energy-saving synthetic routes. However, homogeneous catalysis is not widely used in industrial processes, due to the need of mostly toxic ligands and the costly purification along with a restricted reusability potential.^11–13 Heterogeneous catalysis based on the utilization of metal nanocrystals overcomes most of these limitations. Caused by its high surface-to-volume ratio, catalytic activities are drastically increased in comparison to bulk materials. However, the most common disadvantage in NC-based catalysis lies in the occurrence of aggregates during the reaction, which leads to a decrease of the catalytic active surface. Several studies were already presented in literature to delay that phenomenon using micelle-like- or core–shell-nanostructures as potential nanocatalytic systems.^14–20 However, these surfactants or shells are either potentially harmful for the environment or very step-inefficient to produce. Other approaches used the deposition of small NCs in a mesoporous support which offers a large active surface area.^21–24 In this context, it is crucial to find an adequate balance between permeability for small organic molecules and the overall stability of the nanocatalyst. Overcoming these obstacles can contribute to a sustainable supply of drugs and other organic substances.In the present work palladium-nanocubes (Pd-NCubes) were fabricated in aqueous solution using cetyltrimethyl-ammoniumbromide (CTAB) as surfactant. The procedure is adapted from a previously published study.^25,26 The respective transmission electron microscopy (TEM) image and the selected area electron diffraction (SAED) pattern of the as-obtained Pd-NCubes are depicted in Fig. S1.^† The SAED confirms the single-crystallinity of Pd-NCubes bound by {100}-facets. TE micrographs of Pd-NCubes revealed an average edge-length of (18 ± 2) nm (for histogram see Fig. S2^†). The formation of polyhedra and nanorods was found to be less than 1%.To the best of our knowledge, there is no procedure reported that showed the direct fabrication of a mesoporous silica (mSi) shell tailored on Pd-NCs. However, Matsuura et al. demonstrated a single-step coating approach of CTAB-capped gold-nanorods and CdSe/ZnS quantum dots obtaining a mesoporous silica shell.²⁷ The pores were determined to be 4 nm in width with 2 nm thick walls. Since the Pd-NCubes are covered by CTAB, already no surfactant exchange is necessary. Consequently, this procedure could be directly transferred to the as-obtained Pd-NCubes (∼10¹⁵ particles per L) of this study using tetraethyl orthosilicate (TEOS) as silica precursor in an alkaline solution. Here, CTAB serves as organic template for the formation of the mesoporous silica shell.TE micrographs of individual Pd-mSi-nanohybrids are depicted in Fig. 1 showing a spherical silica coating with a thickness of (17 ± 2) nm (for histogram see Fig. S3^†). The porosity is essential to ensure that vacant coordination sites on the palladium-core are present and accessible for catalysis. In contrast to other multistep approaches, pores are formed in situ with no additional etching step necessary.^28,29 This avoids the usage of harmful etching agents such as fluorides or ammonia.³⁰ The silica shell then served as platform for further surface modification using two different PEG-silanes (M_n = 5000 g mol⁻¹ and M_n = 20 000 g mol⁻¹). TE microscopy did not reveal any changes in the structure of the PEG functionalized Pd-mSi-nanohybrids opposed to the unfunctionalized nanocatalyst, since the contrast of polymer is too low (see Fig. S4^†). However, dynamic light scattering (DLS) measurements in diluted aqueous solutions proved an increased hydrodynamic radius with increasing molecular weight of the PEG-chain grafted onto the silica shell (see Fig. 2). These results provide evidence that only individual nanostructures are formed while no larger aggregates are present.Open in a separate windowFig. 1Exemplary TE micrographs of Pd-mSi-nanohybrids.Open in a separate windowFig. 2DLS results along the different stages of the hierarchal fabrication process of the nanocatalyst.The successful functionalization of the Pd-mSi-nanohybrid with PEG provides the dispersibility for the overall nanocatalyst in a PEG matrix. Due to its lack of toxicity and its simple recovery, caused by its melting point at ∼50 °C, PEG is considered as a “green” reaction medium.³¹ It has already been shown that PEG can act as suitable solvent for both homogeneous and heterogeneous catalysis.^32–34 Consequently, further experiments were performed using only the Pd-mSi-nanohybrid functionalized with PEG-silane having an average molecular weight of M_n = 5000 g mol⁻¹ (PEG-5k). For catalytic reactions, Pd-mSi-PEG-5k was dispersed in a PEG matrix (PEG-2000, M_n = 2000 g mol⁻¹) and charged into a Teflon centrifuge tube. Using only one tube for the reaction and the product separation avoids an additional transfer step and prevents any loss of the nanocatalyst between reaction cycles (see recycling Scheme 1). Here, the C–C-coupling between ethyl acrylate and p-iodoanisole served as model Heck-reaction to prove the catalytic activity of the designed catalyst (4.4 mol% overall Pd conc. equal to ca. 0.3 mol% surface-available Pd; conc. is determined by ICP-MS measurements, see Table S1^†). Sodium phosphate was used as base providing the largest product yield when compared to other bases, such as Na₂CO₃, K₂CO₃ and K₃PO₄. Once the catalysis was performed and the PEG-2000 was cooled down, diethyl ether was added to extract the product and separated from the reaction medium via centrifugation.Open in a separate windowScheme 1Recycling process of the Pd-mSi-PEG-5k-nanocatalyst and PEG-2000 as solvent after the Heck-reaction between ethyl acrylate and p-iodoanisole to form ethyl p-methoxycinnamate.To exclude any suspended compounds from the desired product, the mixture was passed through a PTFE-filter. Et₂O was removed in vacuo without the need of column chromatography. Comparing the ¹H-NMR spectra of the as-obtained product with the educts indicate a yield of 98% with only small amounts of PEG-2000 present (identified by the signal at ∼3.6 ppm, < 1 weight-%). The results show that both educts and the base Na₃PO₄ are able to diffuse through the mesoporous silica shell to the palladium core (see Fig. 3). A detailed ¹H-NMR signal assignment of the product is given in Fig. S5.^†Open in a separate windowFig. 3 ¹H-NMR spectra of the educts ethyl acrylate (top) and p-iodoanisole (center) and the product ethyl p-methoxycinnamate (bottom).After recovering the reaction mixture containing Pd-mSi-PEG-5k and PEG-2000, seven further Heck-reaction-cycles were conducted under the same conditions. Results obtained from ¹H-NMR and gravimetry indicate no significant decrease in catalytic activity (see ¹H-NMR spectra in Fig. 4). Along the eight Heck-reactions, product yields were determined between 94% and 99%. Only small traces of p-iodoanisole (identified by the signal at ∼6.75 ppm) were still present while ethyl acrylate could be fully removed in vacuo. The yields obtained after each cycle are displayed in Fig. 5. ICP-MS measurements of the catalysis product were performed to determine the palladium leaching out of the catalytic system. The results are displayed in Table S1^† indicating that leaching is strongly suppressed since the overall Pd-content in the product ranges from 0.3–5.7 ng, only. This corresponds to 0.002–0.044 ppm palladium with respect to the product mass. The data are in good agreement with the high product yields along the eight Heck-reactions. To trace the evolution of the nanocatalyst along the cycles, TE micrographs were taken after the 1^st and the 8^th Heck-cycle (see Fig. 6). It can be seen that the cubical structure of the palladium vanishes during the first reaction (left TEM image). Inside the silica shell spherical palladium particles were formed. An explanation for this rearrangement lies in the suggested Heck-mechanism.³⁵ Here, a Pd²⁺-species forms after the oxidative addition of the p-iodoanisole which can desorb from the Pd-NCube. Once the reductive elimination of the product occurs the Pd⁰-species is generated again that can re-deposit on the palladium-core.²¹ Since the spherical geometry possesses the lowest free surface energy, globules were eventually formed.³ The mesoporous silica shell is not affected significantly by the catalysis and the rearrangement of the palladium. The porosity appears to stay intact, enabling the penetration of further small organic molecules. Control experiments were performed to validate whether these observations are based on either a thermally or a chemically induced rearrangement process of the palladium core. Therefore, only the nanocatalyst was dispersed in PEG-2000 and heated at 110 °C for 24 h without any conducted catalysis reaction. TEM results showed no changes in the structure of neither the palladium core nor the silica shell (see Fig. S6^†). After the 8^th reaction, no core–shell-nanostructures could be detected anymore via TEM. Only small randomly shaped Pd-nanoparticles (≤20 nm) were found indicating a slow leaching of the palladium out of the silica shell. However, no larger aggregates were formed (see right TE micrograph in Fig. 6) which explains the continuously high catalytic activity.Open in a separate windowFig. 4 ¹H-NMR spectra of the product ethyl p-methoxycinnamate after the 1^st Heck-reaction (bottom) up to the 8^th reaction (top).Open in a separate windowFig. 5Conversions of ethyl p-methoxycinnamate obtained via NMR and gravimetry after each respective Heck-reaction (1–8).Open in a separate windowFig. 6TE micrographs of the Pd-mSi-PEG-5k-nanocatalyst taken after the 1^st (left) and the 8^th Heck-reaction (right).     

Luminometric dosimetry of X-ray radiation by a zwitterionic uranium coordination polymer

A novel X-ray dosimeter based on a uranium coordination polymer U-Cbdcp was obtained by the judicious synergy between the luminescent uranyl centres and zwitterionic tritopic ligands. Notably, U-Cbdcp exhibits luminescence quenching upon increasing X-ray dose, which in combination with its excellent radiolytic stability, makes it suitable for X-ray dosimetry.

A novel X-ray dosimeter based on a uranium coordination polymer has been developed by the judicious synergy between the luminescent uranyl centres and zwitterionic tritopic ligands.

X-ray radiation has been extensively used in medical diagnosis and treatment, security screening, quality control inspection, scientific instrumentation, etc.^1–3 Overexposure to X-ray radiation cause damage to human cells, which could result in skin burn, tissue damage, and increased incidence of cancer.^4,5 Moreover, X-ray dosimetry is required in many industrial fields, including food irradiation, sterilization, and material modification.⁶ Thus, different types of radiation dosimeters, including ionization chamber, scintillator, semiconductor, thermoluminescence dosimeter, chemical dosimeter, and so on, have been commercialized to quantify the incident X-ray dose.⁶ The former three types of dosimeters are more frequently applied to record the dose-rate of incident radiation.^7–10 Thermoluminescence dosimeters and chemical dosimeters are suitable for dosimetry of accumulated dose, but they suffer from critical drawbacks such as cumbersome reading processing, instrument-demand, or cost-ineffectiveness.^11,12 Therefore, further development of new types of X-ray dosimeters remains essential.Coordination polymers, which are assembled from metal ions and organic ligands, have been met with great interest in diverse fields including catalysis, sensing, sorption, separation, and luminescence.^13–18 Their tunability in terms of chemical composition, structure, and more importantly photophysical property, makes them promising for radiation detection. Indeed, pioneering works by Allendorf and co-workers have demonstrated that scintillating metal–organic frameworks (MOFs) assembled from metal cations and radioluminescent organic ligands can function as a new type of radiation detection materials.¹³ Furthermore, coordination polymers or cluster species showing radiochromism, radio-photoluminescence, fluorochromism, and photoluminescence quenching upon accumulated doses of ionizing radiation have been documented, making them as promising candidates of radiation dosimeters.^19–26 Notably, the abundance of luminescent centers or radio-responsive moieties in some of these materials renders higher saturation point in response to radiation dose. This attribute allows for wider operation ranges or higher upper limits of detection compared with those of traditional metal-ion-doped inorganic dosimeters, e.g. Ag-doped phosphate glass and Mg²⁺-doped LiF (LiF:Mg).²⁷We have recently undertaken a study focused on developing actinide-based coordination polymers or cluster materials for their promising applications in ionizing radiation detection.^19,21,28 The large coordination numbers and diverse coordination geometries of actinide cations engender a myriad of topologies of these materials.^29–31 Moreover, the intrinsically intense green emission from the uranyl cation can be utilized as a radio-luminescent center.^23,26 In addition, the slight radioactivity of ²³⁸U (t_1/2 = 4.47 billion years) can be neglected in the course of high dose radiation detection.^32,33 Herein, a novel zwitterionic uranium coordination polymer is reported, showing rather unique fluorescence quenching response to X-ray radiation. This radio-responsive feature, in combination with its high radiolytic stability, points to the potential implementation of uranium-bearing materials for radiation dosimetry.Solvothermal reaction between UO₂(NO₃)₂·6H₂O, zwitterionic N-(4-carboxybenzyl)-(3,5-dicarboxyl)pyridinium bromide (H₃CbdcpBr), and CH₃COOH in DMF/H₂O mixed solution at 100 °C afforded yellow crystals of UO₂(OH)(H₂Cbdcp)(HCbdcp)·4H₂O (U-Cbdcp) with a yield of 63% based on U.Single crystal X-ray diffraction (SCXRD) analysis revealed that U-Cbdcp crystalizes in the monoclinic P2₁/n space group (Table S1^†). The asymmetric unit of U-Cbdcp network consists of one crystallographically independent UO₂²⁺ cation, two Cbdcp ligands, and one hydroxide group (Fig. S1^†). The coordination geometry of uranyl cation can be best described as a typical pentagonal bipyramid, of which four O atoms on the pentagonal plane are donated from three Cbdcp ligand and the rest one is from a hydroxide group (Fig. 1a).^34–38 One of the organic linkers coordinates with one uranyl cation in a μ₁–η₁ bridging mode, while the other one interconnects with two uranyl cations in a μ₂–η₁:η₂ manner (Fig. 1b). Therefore, these two crystallographically unique ligands can be assigned as H₂Cbdcp⁻ and HCbdcp²⁻ with one and two carboxylate group being deprotonated, respectively. The torsion angles between the carboxybenzyl and (3,5-dicarboxyl)pyridinium moieties are measured to be 110.909° and 112.447° for H₂Cbdcp⁻ and HCbdcp²⁻, respectively, as defined by ∠N–C–C. The assembly of uranyl cations, H₂Cbdcp⁻, and HCbdcp²⁻ ligands results in the formation of a one-dimensional infinite chain extending along the c axis (Fig. 1c). The afforded chains are further extended into a 3D supramolecular network via π–π interactions and hydrogen bonds between the ligands (Fig. 1d). The phase purity of bulky U-Cbdcp sample was confirmed by powder X-ray diffraction (PXRD), showing that the measured pattern matches well with the simulated one (Fig. S2^†).Open in a separate windowFig. 1(a) The coordination environment of UO₂²⁺ cation. (b) The coordination modes of two crystallographically independent ligands. (c) The 1D chain of U-Cbdcp extending along the c axis. (d) Representation showing the network of U-Cbdcp. In figure (a)–(c), U atoms are in green, O atoms are in red, N atoms are in blue, and C atoms are in grey.The solid-state luminescence spectrum (λ_ex = 365 nm) was collected on a tablet of U-Cbdcp, that was fabricated from finely ground powder. As expected, U-Cbdcp exhibits five characteristic bands of uranyl cation centring at 488, 508, 531, 556, and 583 nm (Fig. 2a). This intense green emission can be attributed to the HOMO-LUMO transition occurring in the uranyl bonds upon UV excitation.^38,39 Strikingly, the uranyl-based luminescence is strongly quenched after X-ray radiation (4.7 kGy) as shown by the photographs of U-Cbdcp tablet (Fig. 2a inset). Concomitantly, the intensities of characteristic UO₂²⁺ emission bands, which were measured from the tablet exposed to specific interval of X-ray dose, gradually diminished upon continuous X-ray irradiation (Cu-K_α, 120 Gy min⁻¹). More specifically, approximately 44% luminescence intensity was retained after being exposed to 260 Gy X-ray radiation (Fig. 2a). Further increasing the dose to 4.7 kGy resulted in nearly 90% emission quenching. Interestingly, I₀/I as a function of radiation dose can be well fitted with a linear correlation with R² of 0.9988, where I₀ and I are the luminescence intensities monitored at 508 nm before and after irradiation, respectively. This excellent linearity allows for quantifying X-ray dose in a wide dynamic range spanning from 10 to 4700 Gy via a luminescence “turn-off” manner. To obtain limit of detection (LOD), the calibration curve was established by plotting the quenching rate (I₀ − I)/I₀ as a function of dose at the low dose range (0–30 Gy) (Fig. S3^†). The limit of detection (LOD) is calculated to be 0.093 Gy based on the method reported by Zang and coworkers.⁴⁰ Markedly, this LOD is comparable to 0.047 Gy of the most sensitive photochromic sensor Htpbz@Th-SINAP-2.²¹Open in a separate windowFig. 2(a) X-ray dose-dependent fluorescence spectra and optical micrographs (inset) of a U-Cbdcp tablet. (b) The plot showing the linear correlation between I₀/I and X-ray dose.To decipher the quenching mechanism, the structures of U-Cbdcp before and after X-ray irradiation (5 kGy) were thoroughly characterized by combined techniques including PXRD and SCXRD. The PXRD patterns of U-Cbdcp remained approximately unchanged upon irradiation, ruling out our initial speculation of radiation induced damage to the bulky sample (Fig. S4^†). This supposition is additionally supported by the nearly identical FTIR spectrum of irradiated U-Cbdcp with the nonirradiated one (Fig. S5^†). Furthermore, SCXRD analysis before and after X-ray radiation was conducted on the same single crystal of U-Cbdcp and revealed that the overall network derived from these two datasets retain unchanged as well (Table S1^†). In detail, the local structure as represented by the bond length and bond angle of U-Cbdcp changes slightly, which can be attributed to the standard deviations of these parameters obtained from SCXRD (Table S2^†). This observation further excludes the quenching mechanism via decomposition of U-Cbdcp crystal.There is precedence in literature that the luminescence quenching can be associated with the generation of radicals via radio-induced bond break or electron transfer.^23,26,41,42 Therefore, electron paramagnetic resonance (EPR) spectrum of irradiated U-Cbdcp was collected and indeed shows an intense EPR signal with a g-tensor of 2.0197, corresponding to the value (g = 2.0023) of a free electron (Fig. 3).⁴³ The freshly synthesized sample, however, is EPR silent for comparison. To identify the location of radical species in the coordination polymer, EPR spectra of H₃CbdcpBr ligand before and after irradiation were recorded as well. As shown in Fig. S6,^† the irradiated H₃CbdcpBr exhibits a relatively weak resonance with a g factor of 2.0198, which is comparable with that of U-Cbdcp. In the light of aforementioned results, we may conclude that continuous X-ray radiation generates ligand-based radical species, which functions as a quencher via a nonradiative energy transfer pathway.^44–46Open in a separate windowFig. 3EPR spectra of U-Cbdcp before and after 5 kGy X-ray radiation.Encouraged by the structural integrity of U-Cbdcp upon 5 kGy X-ray irradiation, we further investigated its radiolytic stability by irradiating the sample with high dose β-ray and γ-ray radiations. The radiations were provided by a custom-built electron cyclotron (1.2 MeV) and a ⁶⁰Co irradiation source (2.22 × 10¹⁵ Bq) with dose rates of 150 and 11.8 kGy per h, respectively. PXRD study indicated that no obvious changes in long-range order or loss of crystallinity of U-Cbdcp were observed after radiations, implying excellent radiation resistance of U-Cbdcp (Fig. S7^†).In summary, a new 1D uranium coordination polymer built from uranyl cations and zwitterionic Cbdcp ligands were obtained solvothermally. One of the most intriguing properties of U-Cbdcp is the occurrence of luminescence quenching upon X-ray radiation. This unique radio-induced luminometric response can be utilized as a strategy for X-ray dosimetry. Notably, the quenching response can be well fitted with a linear correlation and the detection limit was calculated to be 0.093 Gy. This finding, in conjunction with the excellent radiation resistance of U-Cbdcp, point to potential applications of uranium bearing materials for radiation detection.     

Micro-supercapacitors based on oriented coordination polymer thin films for AC line-filtering

Reported herein is a facile solution-processed substrate-independent approach for preparation of oriented coordination polymer (Co-BTA) thin-film electrodes for on-chip micro-supercapacitors (MSCs). The Co-BTA-MSCs exhibited excellent AC line-filtering performance with an extremely short resistance–capacitance constant, making it capable of replacing aluminum electrolytic capacitors for AC line-filtering applications.

Micro-supercapacitors exhibiting excellent AC line-filtering with oriented coordination polymer thin-film electrodes are fabricated based on a substrate-independent electrode fabrication strategy.

Micro-supercapacitors (MSCs), as important Si-compatible on-chip electrochemical energy storage devices, have attracted rapidly growing attention due to their rapid energy-harvesting features and burst-mode power delivery.^1,2 In the past few years, a variety of materials including carbon nanotubes,³ graphene,⁴ graphene oxide and mesoporous conducting polymers,^5,6 have already been explored to fabricate the electrodes of MSCs for improving their electrochemical performance. Unfortunately, fabrication procedures of most of these active materials suffer from high cost, harsh and complicated processing conditions, as well as easy cracking and delamination of active films,^1,4 extremely limiting their commercial applications. Moreover, their performances are unsatisfactory for alternating current (AC) line-filtering, which is a key parameter to implement high-frequency operation in most line-powered devices.^7–9For AC line-filtering, capacitors need to respond harmonically at 120 Hz to attenuate the leftover AC ripples on direct current voltage busses.¹⁰ Notably, the development of more compact and miniaturized capacitors to replace traditional aluminum electrolytic capacitors (AECs) for AC line-filtering has become one of the major tasks for future electronics.¹¹ However, typical supercapacitors are incapable for AC line-filtering at this frequency due to their limited ion diffusion and charge transfer efficiency, corresponding to the unsuitable architectures and low conductivity of electrode materials.^10–12 Therefore, the design and fabrication of highly conductive electrodes with optimized architectures for facial electron/ion transportation is crucial for improving the performance of MSCs in AC line-filtering.^12,13 It is worth mentioning that great advancements have been achieved by utilizing vertically oriented graphene sheets as well as 3-dimensional graphene/carbon nanotube carpets prepared by chemical vapor deposition (CVD),^7,8 yielding efficient filtering of 120 Hz AC with short resistance–capacitance (RC) time constants of less than 0.2 ms, which is competitive with those of porous carbon-based supercapacitors (RC time constant = 1 s) as well as AECs (RC time constant = 8.3 ms).⁸ However, the CVD method necessitated in the fabrication of graphene/carbon nanotube electrodes suffers from high cost and complicate procedures.Coordination polymers with an unrivalled degree of structural and property tunability which could be realized by facial procedures, are promising candidates for energy storage.^14,15 Recently, a remarkable achievement which demonstrated a facile and low-cost solution-processed method towards on-chip MSCs based on an azulene-bridged coordination polymer framework (PiCBA) on a Si wafer-supported Au surface was reported.¹⁴ Nevertheless, the reported preparation of coordination polymer film exhibited strong dependence on the surface chemistry (functionality) of the substrate and further improvement of their electrochemical stability was needed. Therefore, the development of substrate-independent fabrication strategies of large-scale and uniform coordination polymer films is in great need not only for fundamental studies, but also for technological applications especially in electronics.Herein, we demonstrate a facial solution-based substrate-independent approach to fabricate oriented coordination polymer (Co-BTA) thin-film electrodes. Remarkably, rigid and flexible Co-BTA-based MSCs with excellent electrochemical stability and AC line-filtering performance were realized, indicating great application potential in micro-supercapacitors.As demonstrated in Fig. 1a–c, a large scale and continuous Co-BTA coordination polymer film composed of one dimensional (1D) molecules ([Co(1,2,4,5-bta)]_n) was prepared at the air–liquid interface through a coordination reaction between 1,2,4,5-benzenetetramine tetrahydrochloride (BTA) and cobalt acetate tetrahydrate (Co(CH₃COO)₂·4H₂O). Notably, the preparation of Co-BTA film is basing on mild conditions and independent of any substrates. The resulting film can be transferred onto any supports including rigid silicon (Si) wafer, glass, as well as flexible PET substrate, indicating great substrate-independence and making it practically applicable for various applications. Besides of a brown film formed at the air–liquid interface, a powder product is also obtained at the bottom of the reaction bottle.Open in a separate windowFig. 1(a) Synthesis of Co-BTA through the coordination reaction between BTA and cobalt ions. (b) Illustration of the gas–liquid interface growth of Co-BTA film. (c) Photographs of the reaction system before and after the coordination reaction.To study the morphology of the resulting Co-BTA film, the brown film was transferred onto a SiO₂/Si wafer by immersing the wafer down to the reaction mixture and subsequently lifting the film up. The scanning electron microscopy (SEM) image reveals a highly uniform and large-scale distribution of the obtained film without cracks or wrinkles (Fig. 2a), which is superior to other reported coordination polymer films obtained via a similar method.¹⁶ An average thickness of approximately 60 nm of the Co-BTA film is observed from the cross-sectional SEM image as shown in Fig. 2b. Interestingly, thickness of the obtained coordination polymer film could be well controlled and Co-BTA films with thicknesses up to several hundred nanometers could be well prepared by adjusting the ratio of raw materials (Fig. 2c and d).Open in a separate windowFig. 2(a) Planar SEM image of Co-BTA film. Cross-sectional SEM images of Co-BTA films with a thickness of (b) 60 nm, (c) 160 nm and (d) 260 nm.To investigate the structure information of the resulting Co-BTA and further explore the coordination reaction, characterizations including powder X-ray diffraction measurements (PXRD), X-ray photoelectron spectroscopy (XPS) and Fourier transform infrared spectroscopy (FTIR) were carried out. The PXRD pattern of Co-BTA powder shown in Fig. S1a^† is in great agreement with that simulated from the crystal structure of Ni(dhbq)·nH₂O (Fig. S1b^†), suggesting that Co-BTA and Ni(dhbq)·nH₂O is isostructural and forms 1D structures with straight infinite chain extends.¹⁷ More interestingly, PXRD measurements employing two different scattering geometries (Fig. S2^†) on the Co-BTA thin-film demonstrate two quite different diffraction patterns. As observed in Fig. 3a, the diffraction pattern observed for the out-of-plane scattering geometry exhibits three characteristic peaks of the Co-BTA film at ∼12°, 24° and 36°, which are corresponding to (001), (002) and (003), respectively. In contrast, the in-plane PXRD profile employing grazing-incidence XRD (GIXRD) technique at an incident angle (α) of 0.2° demonstrates a main peak at ∼18°, which is assigned to the (110) diffraction peak. Different diffraction peaks observed through these two XRD scattering geometries indicate an orientation nature of the as-prepared Co-BTA film,¹⁸ which exhibits better crystallinity compared with the powder Co-BTA product. In addition, the N 1s core level spectrum for Co-BTA film exhibit one typical peak at 399.1 eV, which is corresponding to the amido coordinated with Co^II, indicating the strong coordination between Co^II and BTA (Fig. 3b). The weak peak at ∼401 eV is assigned to N–O due to the oxidation of ligand BTA in ambient environment before reaction. The atomic ratio of N : Co is calculated to be 3.53 : 1 for Co-BTA film and 3.71 : 1 for Co-BTA powder respectively (Fig. S3 and Table S1^†), which is close to the theoretical stoichiometric ratio (4 : 1) for Co-BTA structure, suggesting a high degree of coordination in the resulting product through one Co cation and two benzenetetramine groups. Moreover, the disappearance of two characteristic N–H stretching modes from –NH₂ after the coordination reaction whereas the phenyl-related vibration still exists, further confirms the existence of –NH– in the product through the loss of one H per –NH₂ (Fig. S4^†).¹⁹Open in a separate windowFig. 3(a) PXRD profiles of out-of-plane XRD, in-plane XRD and simulated PXRD pattern of Ni(dhbq)·nH₂O,¹⁷ respectively. *SiO₂/Si substrate. (b) N 1s core level spectra of the Co-BTA film.On the basis of facial fabrication, substrate independence, highly orientation nature, low band gap (1.68 eV, calculated from Fig. S5^†) and excellent stability in acid environment (Fig. S6^†), the resulting Co-BTA film is considered as a promising candidate for MSCs application. Fig. 4a schematically depicts the stepwise fabrication of a planar Co-BTA film based MSC on a SiO₂/Si wafer and its electrochemical performance is first examined by cyclic voltammetry (CV) with scan rates ranging from 50 mV s⁻¹ to 1000 V s⁻¹ (Fig. 4b and c). At a low scan rate of 50 mV s⁻¹, the 60 nm-thick Co-BTA film based MSC exhibited a pronounced pseudocapacitive effect, implying the occurance of faradaic reaction.²⁰ With the increase of scan rate, a gradual transition of the CV curves from the pseudocapacitive to the typical electrical double-layer capacitive behavior with a nearly rectangular CV shape was observed. Remarkably, the device exhibited a maximum volumetric capacitance of 23.1 F cm⁻³ at 50 mV s⁻¹, which is comparable with those of reported carbon- or graphene-based MSCs (Table S2^†), e.g., onion-like carbon,²¹ vertically oriented graphene,⁸ and carbon nanotubes/graphene.⁷ Even though a trend that C_V decreased gradually with increasing scan rate was observed, the Co-BTA-based electrode still delivered a C_V of 2.7 F cm⁻³ even at a high scan rate of 1000 V s⁻¹, suggesting an excellent capacitive performance of this Co-BTA-based MSC device.⁷Open in a separate windowFig. 4(a) Schematic illustration of the fabrication of MSC device with the Co-BTA film electrode. (b) CV curves of Co-BTA-based MSCs in the H₂SO₄–PVA gel electrolyte at different scan rates. (c) C_V evolution of the MSCs at different scan rates.Electrochemical impedance spectroscopy (EIS) measurements were performed to evaluate the charge transport properties within the Co-BTA-based MSCs. The Nyquist plot shown in Fig. S7^† indicated the kinetic features of electron transfer/ion diffusion at the electrode, from which an almost straight line especially in the low frequency region is observed. Notably, the plot shows a closed 90° slope without a charge transport semicircle at high frequency which is corresponding to an almost ideal capacitive ion diffusion mechanism, due to the excellent charge transfer property of the oriented Co-BTA electrode film. Moreover, this microdevice exhibited a low equivalent series resistance of 13.48 Ω (Fig. S7^† (inset)), further suggesting the ultrafast ion diffusion characteristic in such a Co-BTA-based-MSC.²² It''s suggested that the unique kinetic feature of fast ion diffusion and charge transfer benefits from the intrinsic characteristics of the oriented polymer film composed of 1D molecules, which can not only facilitate rapid ionic diffusion but also facilitate the interfacial charge transfer and faradaic redox reaction between the electrode material and electrolyte.What''s more, the dependence of the phase angle on frequency shown in Fig. 5a delivered a high characteristic frequency f₀ of 6812 Hz at the phase angle of −45° (the resistance and reactance of the device have equal magnitudes),¹⁰ which is much higher than that of an active carbon supercapacitor (5 Hz),²³ sulfur-doped graphene MSCs (3836 Hz),²² or an azulene-bridged coordination polymer framework based MSCs (PiCBA-MSCs) (3620 Hz) and so on,¹⁴ as summarized in Table S2.^† Moreover, a max phase angle of −80° at a frequency of 18 Hz is observed, indicating the performance of this Co-BTA based MSCs is 89% of that of an ideal capacitor. Importantly, a large impedance phase angle of −78.6° was obtained at a frequency of 120 Hz, which is the largest reported value for coordination polymer based MSCs (Table S2^†), suggesting an excellent AC line-filtering performance of our microdevice.⁷ To further conform the ultrahigh fast ion diffusion in Co-BTA-based-MSCs, the relaxation time constant τ₀ (τ₀ = 1/f₀, the minimum time needed to discharge all the energy from the device with an efficiency of greater than 50% of its max. value) was calculated to be only 0.15 ms (6812 Hz), which is orders of magnitude higher than that of conventional electrical double-layer capacitors (1 s),⁸ activated or onion-like carbon MSCs (<200 ms, <10 ms),^21,23 and much shorter than those of MSCs based on carbon nanotubes/reduced graphene oxide (4.8 ms) as well as azulene-bridged PiCBA coordination polymer framework film (0.27 ms).^14,24 Moreover, a short RC time constant (τ_RC) of 0.32 ms was obtained (Fig. 5b) through a series-RC circuit model, making it capable of replacing AECs for AC line-filtering application. To the best of our knowledge, this is the first report of coordination polymer-based MSCs exhibiting such a small relaxation time constant and RC time constant.Open in a separate windowFig. 5(a) Impedance phase angle on the frequency for the Co-BTA-based microdevices. (b) Plot of capacitance (C_V′ = volumetric real capacitance and C_V′′ = imaginary capacitance) versus the frequency of Co-BTA-based microdevices. (c) Cycling stability of Co-BTA film with 10 000 cycles at the scan rate of 50 V s⁻¹. Inset displays the CV curves at the first, five thousandth and ten thousandth cycle, respectively. (d) Ragone plots for Co-BTA film, compared with commercially applied Li-thin-film batteries,²¹ electrolytic capacitors,² CNT-graphene carpets,²⁴ PiCBA coordination polymer and MXene-reduced graphene oxide.^14,25Impressively, this oriented electrode structure exhibits excellent long-term electrochemical stability with ∼96.3% capacitance retention even after 10 000 cycles of charging/discharging at a scan rate of 50 V s⁻¹ (Fig. 5c), which has also been confirmed by comparing the CV curves before and after testing for 10 000 cycles (inset of Fig. 5c). It''s worth pointing out that the as-made Co-BTA-based MSCs exhibit the best electrochemical stability among reported MSCs with coordination polymer electrodes.¹⁴ On the basis of the above discussion, it is reasonable to conclude that the ultrahigh fast ion diffusion/charge transfer in Co-BTA-based-MSCs attributed to the oriented architecture of Co-BTA thin-film electrodes, the excellent AC line-filtering performance, as well as remarkable electrochemical stability contributes to the excellent performances of Co-BTA-based-MSCs. Moreover, the power density and energy density of the as-made device is calculated and compared with that of MSCs based on other electrode materials to evaluate the energy storage performance of the Co-BTA based MSCs. The Ragone plot in Fig. 5d reveals a high power density of 1056 W cm⁻³ for our device, which is at least five orders of magnitude higher than that of commercial thin-film lithium batteries. What''s more, our device exhibits an energy density of up to 1.6 mW h cm⁻³ at 50 mV s⁻¹, which is at least one order of magnitude higher than that obtained for MSCs based on CNTs-graphene carpet and high-power electrolytic capacitors.^2,24To further demonstrate the substrate independence of this fabrication strategy, flexible Co-BTA-based-MSC device was fabricated and investigated basing on a flexible polyethylene terephthalate (PET) substrate instead of rigid Si substrate in the same way (Fig. S8–S10^†). The as-fabricated device exhibited a maximum volumetric capacitance of 22.0 F cm⁻³ at 50 mV s⁻¹, a short relaxation time constant τ₀ of 0.15 ms and a RC time constant (τ_RC) of 0.42 ms, which are close to the properties of devices with a Si substrate, confirming the substrate independence of this fabrication scheme. As a proof-of-concept application, bending tests were carried out and the bended device (radius = 1 cm) exhibited a small relaxation time constant τ₀ of 0.21 ms and RC time constant (τ_RC) of 0.42 ms, suggesting that the Co-BTA-based MSC with PET substrate in a bended state still delivers a good ion diffusion and AC line-filtering performance.In conclusion, we have demonstrated a facile method that can be used to construct large scale and highly oriented uniform Co-BTA coordination polymer thin films using a very convenient and fast process. With this method, Co-BTA-based MSCs are fabricated without any dependence of the substrate. The as-fabricated MSCs on Si substrate exhibit high specific capacitance, energy density as well as excellent electrochemical stability. Particularly, the fabricated Co-BTA based MSCs deliver excellent AC line-filtering performance with an extremely short RC time of 0.32 ms, attributed to the facilitated ion diffusion beneficial from the oriented architecture of Co-BTA thin film. The high-performance electrochemical properties of Co-BTA-MSCs makes Co-BTA films promising materials to provide more compact AC filtering units for future electronic devices.     

Green synthesis of crystalline bismuth nanoparticles using lemon juice

Lemon juice effectively served as a reducing and capping agent for an easy, cost-effective, and green synthesis of crystalline bismuth nanoparticles (BiNPs) in basic aqueous media. Spherical BiNPs with a rhombohedral crystalline structure are capped by phytochemicals and stably dispersible in aqueous media. The BiNPs effectively catalyze the reduction of 4-nitrophenol to 4-aminophenol by NaBH₄.

Lemon juice effectively served as a reducing and capping agent for an easy, cost-effective, and green synthesis of crystalline bismuth nanoparticles (BiNPs) in basic aqueous media.

Nanostructures of bismuth, the heaviest element among the ‘safe ones’ earning the status of a ‘green element’,¹ are particularly interesting due to their large magnetoresistance and excellent thermoelectric properties.^2–4 Bismuth nanoparticles (BiNPs) are the most extensively used nanostructures of bismuth. For example, BiNPs are utilized as contrast agents for computed tomography, photoacoustic imaging and infrared thermal imaging,^5,6 catalysts for the reduction of nitroaromatic compounds,^7,8 and removal of NO from air.⁹ Furthermore, BiNPs can act as intermediates for the synthesis of other nanostructures of bismuth, such as thermoelectric Bi₂Te₃ (ref. 10) and seeds for the solution–liquid–solid growth of nanowires.^11,12 Techniques for the synthesis of pure BiNPs could be categorized into (i) thermal decomposition,¹² (ii) mechanochemical processing,¹³ (iii) photochemical reduction,¹⁴ and (iv) solution-phase chemical reduction methods.^6,15–19 The thermal decomposition method requires harsh preparation conditions, expensive organometallic precursors, high temperature, and long reaction time, while producing high-quality monodispersed BiNPs. The mechanochemical processing technique is advantageous in terms of using inexpensive and nontoxic bulk bismuth pellets as precursors. However, the energy consumption and costly instrumentations are the limitations. The photochemical reactions typically require long time for sufficient conversion to bismuth nanoparticles, while they can also produce highly monodispersed BiNPs. The solution-phase chemical reduction methods are most popular due to the facile procedures and accessible reagents. However, stabilizers and toxic reducing agents often used are the limitations. To resolve the above-mentioned limitations, simple and green alternative methods are highly demanded.The use of abundant plant sources, which has been applied for synthesis of various metal nanoparticles such as Ag and Au, is a promising solution.^20,21 Plant sources contain a wide variety of biomolecules potentially serving as reducing and capping agents. Edible plant sources are obviously safest. To the best of our knowledge, there is no report on the synthesis of crystalline BiNPs using plant sources, while P. Poltronieri et al. reported synthesis of amorphous BiNPs using hydroalcoholic extract of Moringa oleifera.²² We focused on lemon, a very common fruit containing abundant antioxidants such as polyphenols, limonoids, citric acid, ascorbic acid, and vitamins potentially reduce ions with high oxidation states. Some of the phytochemicals, namely carbohydrates and proteins bearing ionic moieties, can be capping agents. Accordingly, lemon juice was successfully applied for the formation and in situ stabilization of silver and gold nanoparticles in aqueous media.^23,24 We presumed that a similar mechanism also works for BiNPs.In this communication, we introduce a greener strategy for the synthesis of crystalline BiNPs using lemon juice as a reducing and capping agent. For example, the synthesis was carried out using Bi(NO₃)₃·5H₂O (0.25 mmol) and freshly prepared lemon juice (25 mL) at 80 °C for 2 h under an aerobic basic condition. The X-ray diffraction (XRD) pattern (Fig. 1) of the obtained product was indexed to the pure rhombohedral phase of elemental bismuth (JCPDS no. 44-1246), indicating that the obtained product is BiNPs without detectable oxide phases. The average crystallite size was calculated to be 20 nm applying the Scherrer''s equation on the peaks of the (012), (104) and (110) plane. This result indicates that the phytochemicals present in lemon juice have ability to reduce bismuth salts to form BiNPs. The plausible phytochemicals for reduction are ascorbic acid, citric acid, and sugars. The reduction of Bi³⁺ with glucose⁸ and ascorbic acid¹⁷ was reported, and we accordingly performed the control experiment using possible reducing agents contained in lemon juice, namely ascorbic acid, glucose, and starch without any stabilizing agents. All of them could form elemental bismuth in highly aggregated forms as confirmed by the XRD patterns (ESI, Fig. S9^†) and SEM images (ESI, Fig. S10^†).Open in a separate windowFig. 1XRD pattern of obtained BiNPs synthesized using lemon juice with that of authentic Bi (JCPDS no. 44-1246).Electron microscopy was employed to confirm the size and the morphology of synthesized BiNPs. The scanning electron micrography (SEM) image (Fig. 2a) shows spherical objects having the size in the range of 50 to 100 nm agglomerated and covered with amorphous substances presumably originating from lemon juice. This agglomeration occurred during drying as confirmed by the stable water dispersibility of BiNPs with an average hydrodynamic diameter (D_h) of 255 nm investigated by dynamic light scattering (DLS) (ESI, Fig. S1a^†). The high colloidal dispersibility of the BiNPs and the coating layer observed in the SEM image suggest that phytochemicals present in lemon juice act also as capping agents. The larger D_h measured by DLS than the size observed by SEM can be attributed to the surrounding hydration layer and swelled phytochemicals attached to the surface of the BiNPs. We then performed transmission electron microscopy (TEM) analysis to get the actual size of the Bi cores. The TEM images of the BiNPs (Fig. 2b and c) indicated the presence of heavy elements in the matrix of light elements. The particles are spherical, and the diameter ranges from 8 to 30 nm. The high-resolution TEM image (Fig. 2d) shows the lattice fringes of 0.298 nm, 0.253 nm, and 0.224 nm of the typical crystallite agreeing well with the distance of the (012), (104) and (110) plane, respectively, of the rhombohedral Bi(0).Open in a separate windowFig. 2(a) SEM and (b–d) TEM images of BiNPs synthesized using lemon juice.The EDX spectrum (ESI, Fig. S2^†) reveals that obtained BiNPs comprise bismuth (Bi), carbon (C), and oxygen (O). The strongest signal of bismuth testifies the successful synthesis of BiNPs, whereas the C and O signals demonstrate the presence of the capping layer on the BiNPs.The FT-IR spectrum of the synthesized BiNPs was analyzed to presume the possible components on the BiNPs surface with the comparison with the solid content of lemon juice (Fig. 3). The FT-IR spectrum of the BiNPs shows the following major absorption bands. The broad absorption band at 3050–3500 cm⁻¹ is assignable to the O–H stretching of alcohol moieties. The absorption band around 2840–2940 cm⁻¹ is assignable to the C–H stretching of alkane, and the absence of the sharp absorption around 2950–3050 cm⁻¹ suggests the absence of trace contents of unsaturated C–H groups. The absence of the adsorptions originating from COO–H around 2400–3100 cm⁻¹ and C O around 1700–1730 cm⁻¹ indicates that citric acid^25,26 and other carboxylic acids contained sufficiently in lemon juice are not the major components on the BiNPs, while the presence of the strong absorption around 1600 cm⁻¹ is assignable to carboxylate moieties. The absorption bands around 930–1130 cm⁻¹ assignable to C–C and C–O stretching and around 1200–1420 cm⁻¹ assignable to O–C–H, C–C–H and C–O–H bending suggest the presence of sugars on BiNPs.^25,26 The content of the organic moieties was estimated approximately 14% by thermogravimetric analysis from weight loss that occurred at the temperature range from 130 to 450 °C (ESI, Fig. S3^†), at which the negligible C and O signals were observed in the EDX spectrum of the residue after TGA (ESI, Fig. S2^†).Open in a separate windowFig. 3FT-IR spectra of solid content of lemon juice and obtained BiNPs synthesized using lemon juice.The ¹H and ¹³C NMR and FTIR spectroscopic analysis (ESI, Fig. S4–S8^†) of the ethanol and chloroform extracts of the BiNPs suggests that the major capping phytochemicals are polysaccharides and fatty acid derivatives. Other minor possible components may include amino acids, terpenes and phenolic compounds contained in lemon juice.The successful formation of BiNPs made us enthusiastic to investigate their catalytic performance in the reduction of nitroaromatic compounds, problematic pollutants arising from explosives, analgesic, and antipyretic drugs and dyes.²⁷ Herein, the catalytic performance of our BiNPs was investigated by selecting the reduction of 4-nitrophenol (4-NP) to 4-aminophenol (4-AP) by NaBH₄ at room temperature as a model reaction.^7,8,28 The progress of the reduction was monitored by UV-vis absorption spectroscopy. The colour of the 4-NP aqueous solution changed from light yellow to deep yellow upon the addition of NaBH₄ to produce 4-nitrophenolate ion and an intense absorption peak at 403 nm appeared instead of the original absorption peak of 4-NP at 316 nm in the UV-vis spectrum.⁷ In the absence of BiNPs, the colour of the solution and the intensity of the peak were retained even after 12 h. In the presence of BiNPs, the deep yellow mixture became almost colourless within 180 min (Fig. 4a). The intensity of the absorption peak at 403 nm decreased overtime, and simultaneously a new absorption peak appeared and grew at 299 nm, indicating the formation of 4-AP (Fig. 4a).^8,27,29 The reaction rate almost follows pseudo-first-order kinetics agreeing with the Langmuir–Hinshelwood mechanism; in which two reactants react after adsorption on the solid surface, and then the product desorbs. The rate constant (k) was determined from the plot of ln(A/A₀) vs. time (Fig. 4b) according to the previous reports.^7,8,28,29 The k value of 0.0134 min⁻¹ (activity factor = 0.1 s⁻¹ g⁻¹) is lower than previously reported PVP-coated bismuth nanodots (6.033 s⁻¹ g⁻¹)⁸ and starch coated BiNPs (0.02751 s⁻¹).⁷ The lower catalytic rate of our BiNPs can be attributed to the electrostatic repulsion of 4-nitrophenolate ions with negatively charged BiNPs (zeta potential value = −31.7 mV) in the reaction mixture. The initial rate is slower than the rate after 50 min. Possible factors are (i) the inductive period for the surface activation of the nanoparticles at the initial stage as reported for bismuth nanodots by Liang et al.⁸ and (ii) partial and gradual release of phytochemicals under the basic conditions. After the catalytic reaction, the D_h value measured by DLS (195 nm) became smaller than the original one maintaining the good water dispersibility, suggesting unravelling of primary particles fused by the capping layer (ESI, Fig. S1b^†). In addition, the weight loss in the TGA curve became lower (ESI, Fig. S3^†), and the SEM image implies the partial loss of the coating (ESI, Fig. S11^†). This decrease of the organic moieties is attributable to the partial removal of coating substances such as fatty acids during the catalytic reaction under the basic and reductive conditions, supported by the disappearance of the absorption band around 1600 cm⁻¹ assignable to carboxylate moieties in the FTIR spectrum of BiNPs after the catalytic reaction (ESI, Fig. S12^†). Carboxylic acids and their esters are reported to be reduced to alcohol by NaBH₄ in the presence of electrophiles.^30–33 The initial presence of the amphiphilic and anionic layer plausibly delays the catalytic reaction.Open in a separate windowFig. 4(a) Optical images and absorption spectra of catalytic reduction of 4-NP ([4-NP] = 15 ppm) by NaBH₄ (4.28 × 10⁻⁴ M) in presence of BiNPs (142 mg L⁻¹); (b) pseudo-first order kinetic plot of catalytic reduction.In conclusion, we have demonstrated a green, cost-effective, and successful approach for synthesis of crystalline BiNPs using lemon juice as a reducing as well as capping agent. The obtained BiNPs effectively catalysed the reduction of 4-NP to 4-AP by NaBH₄. The importance of this method lies in the simple synthetic procedure, uses of safe and low-cost lemon, and good dispersibility over conventional chemical approaches.     

Proton transfer network with luminescence display controls OFF/ON catalysis that generates a high-speed slider-on-deck

A three-component network for OFF/ON catalysis was built from a protonated nanoswitch and a luminophore. Its activation by addition of silver(i) triggered the proton-catalyzed formation of a biped and the assembly of a fast slider-on-deck (k₂₉₈ = 540 kHz).

Upon addition/removal of silver(i) ions and due to efficient inter-component communication, a supramolecular multicomponent network acts as an OFF/ON proton relay with luminescence display enabling switchable catalysis.

Proton relays in living systems¹ are often coupled to bio-machinery with enzymatic catalysis.² Frequently, such proton transfer systems comprise intricate networks with information exchange^3,4 amongst multiple components. Using manmade molecules,⁵ however, it is still arduous to create networks that approach the complexity⁶ and functions of biosystems.Although activation via proton transfer plays an eminent role in molecular logic,⁷ switching,^8–10 and machine operation,^11–15 proton transfer networks in combination with the regulation of catalysis remain scarce.^16,17 Beyond that, we present the in situ catalytic synthesis of a highly dynamic supramolecular device by a new proton transfer network requiring interference-free information handling,¹⁸ self-sorting^19–21 and communication²² among multiple components.^23–25Recently, our group has developed various chemically actuated networks of communicating components and used them for the ON/OFF regulation of transition metal catalysis.^26–28 Herein, we describe a proton relay system that reversibly toggles between two networked states and is useful for regulation of acid catalysis. It catalyzes an amine deprotection which was utilized to fabricate in situ a high-speed slider-on-deck,^29,30 for the first time based on aniline → zinc-porphyrin³¹ (N_an → ZnPor) interactions.The communicating network consists of nanoswitch 1 (ref. 32) and luminophore 2 (ref. 33) as receptors (Fig. 1). In the self-sorted networked state (NetState-I), the added proton is captured in the cavity of nanoswitch 1 resulting in [H(1)]⁺, while luminophore 2 remains free. NetState-I is catalytically inactive as the encapsulated proton is unable to catalyze the deprotection of the trityl protected slider biped 4. Upon addition of AgBF₄ to NetState-I, the silver(i) coordinates to switch 1 and the proton is released onto luminophore 2 thus leading to formation of [Ag(1)]⁺ and [H(2)]⁺ in NetState-II (see Scheme 1), which may be followed by characteristic ratiometric emission changes. If NetState-II is formed in presence of 3 and 4, the catalytic trityl deprotection of 4 produces the free biped 5 that eventually binds to deck 3 furnishing the slider-on-deck [(3)(5)].Open in a separate windowFig. 1Chemical structures and cartoon representations of the ligands used in the study.Open in a separate windowScheme 1Schematic representation of the reversible proton relay network. Green circle represents catalytic site.The concept of the catalyzed biped formation through a proton transfer network relies on the following considerations: (a) the initial networked state should be catalytically inactive as expected from a self-sorting of the proton inside the nanoswitch leaving the luminophore free. (b) Addition of silver(i) ions should release the proton from the protonated switch thus forming the protonated luminophore which we anticipated to act as catalyst. (c) A di-protected biped should undergo acid catalyzed deprotection by the protonated luminophore and eventually form a slider-on-deck by coordination to a deck. (d) Finally, the product of the catalysis should not intervene in any state of the operation with the main system.As proton receptor we chose the known phenanthroline–terpyridine nanoswitch 1 (ref. 32) in order to tightly capture the proton/silver(i) as HETTAP (HETeroleptic Terpyridine And Phenanthroline) complexes.³⁴ Luminophore 2 was based on the 2,4,6-triarylpyridine scaffold which is known to fluoresce strongly upon protonation due to intramolecular charge transfer (ICT).³⁵To investigate the self-sorting within the network, we mixed nanoswitch 1, luminophore 2 and TFA in 1 : 1 : 1 ratio in CD₂Cl₂ which furnished NetState-I = [H(1)]⁺ + 2. ¹H NMR shifts of the 4/7-H proton signals to 8.43 and 8.69 ppm and of 9-H and e-H proton peaks to 6.60 and 8.48 ppm confirmed formation of [H(1)]⁺. Moreover, proton signals of c′-H and d′-H at 7.07 and 7.92 ppm corroborated presence of the free luminophore 2 (Fig. 2). Addition of one equiv. of AgBF₄ to NetState-I instantaneously generated NetState-II = [Ag(1)]⁺ + [H(2)]⁺. In the ¹H NMR of NetState-II, the 9-H and e-H proton signals shifted to 6.55 and 8.23 ppm, respectively, which substantiated the formation of [Ag(1)]⁺ whereas the shift of proton peaks c′-H and d′-H to 7.12 and 8.00 ppm, respectively, established the protonated luminophore [H(2)]⁺ (Fig. 2). Upon excitation at 300 nm, NetState-I exhibited emissions at λ = 346, 372 and 393 nm which represent an overlap of the emission peaks of the free luminophore 2 at λ = 346 and 372 nm and of [H(1)]⁺ at λ = 376 and 393 nm (Fig. 3c). In NetState-II, the fluorescence spectrum showed a single emission at λ = 461 nm, which is attributed to [H(2)]⁺ (Fig. 3). In contrast, [Ag(1)]⁺ in NetState-II is nonfluorescent as advocated by the full quenching of the emission of [H(1)]⁺ at λ = 376 and 393 nm upon titration with silver(i) ions (ESI, Fig. S18^†). When one equiv. of tetra-n-butylammonium iodide (TBAI) was added to NetState-II, complete restoration of NetState-I was achieved as confirmed from ¹H NMR data. TBAI acted as a scavenger for silver(i) by removing AgI through precipitation thus reversing the translocation sequence. Multiple switching of NetState-I ⇆ NetState-II was readily followed by ¹H NMR (Fig. 2) and luminescence changes. Upon successive addition of AgBF₄ to NetState-I, the emission changed to λ = 461 nm (sky blue emission of [H(2)]⁺) in NetState-II (Fig. 3a and b) in a clean ratiometric manner that allowed monitoring of the amount of liberated protons. Alternating switching between the NetStates was followed by emission spectroscopy over three cycles with a small decline in emission intensity (Fig. 3c and d), which may be due to the progressive accumulation of AgI.Open in a separate windowFig. 2Partial ¹H NMR spectra (500 MHz, CD₂Cl₂, 298 K) showing reliable switching between NetState-I and II over 3 cycles. (a) After mixing of TFA, 1, and 2 in 1 : 1 : 1 ratio; (b) after adding 1.0 equiv. of AgBF₄ furnishing [Ag(1)]⁺ and [H(2)]⁺; (c) after addition of 1.0 equiv. of TBAI; (d) after adding 1.0 equiv. of AgBF₄; (e) after addition of 1.0 equiv. of TBAI, (f) after adding 1.0 equiv. of AgBF₄; (g) after addition of 1.0 equiv. of TBAI. Blue asterisk marked proton signals represent the tetra-butylammonium ion.Open in a separate windowFig. 3(a) Ratiometric emission titration of 2 (2.1 × 10⁻⁵ M) with TFA (5.3 × 10⁻³ M) in CH₂Cl₂. (b) Change of emission upon protonation of 2. (c) Reversibility of the network NetState-I ⇆ II over three cycles upon addition of silver(i) and TBAI (see text, blue arrow assigns direction of switching) monitored by luminescence changes at λ = 461 nm. (d) Multiple cycles monitored by the change of the fluorescence intensity.For probing the catalytic efficiency of each state towards a simple amine deprotection reaction we chose the trityl-protected aniline 6 (Fig. 4) performing two NetState-I ⇆ NetState-II cycles (Fig. 4b). We first prepared NetState-I by mixing 1, 2 and TFA (1 : 1 : 1) in CD₂Cl₂ and then added 6 and TMB (1,3,5-trimethoxybenzene as an internal standard) in a 10 : 10 ratio. After heating for 2 h at 40 °C, the ¹H NMR spectrum revealed no product 7. Clearly, the proton in [H(1)]⁺ is catalytically inactive (ESI, Fig. S10b^†). Addition of one equiv. of AgBF₄ and probing the catalysis again for 2 h at 40 °C revealed formation of product 7 in (30 ± 2)% yield (ESI, Fig. S10c^†). When the catalytic experiment was probed with only [H(2)]⁺ under identical reaction conditions, it provided the same yield (ESI, Fig. S16^†). This suggested that the reversible proton relay network (Scheme 1) functioned reliably also in presence of 6 with complex [H(2)]⁺ as the catalytically active species. Addition of one equiv. of TBAI to the above NetState-II mixture and probing for catalysis, revealed no further product formation. Apparently, addition of TBAI translocated back the proton to nanoswitch 1 thus restoring NetState-I, which resulted in the OFF state for catalysis. Adding one equiv. of AgBF₄ to the mixture and probing again the catalytic activity resulted in formation of further (29 ± 2)% of product 7, which was basically identical to the yield produced in the first cycle of NetState-II (ESI, Fig. S10e^†). Adding TBAI and the consumed amount of substrate showed no further deprotection. In sum, the robust catalytic performance of the proton relay network as reflected in two successive catalytic cycles resulted in no significant decline of the catalytic activity (Fig. 4b).Open in a separate windowFig. 4(a) Representation of the OFF/ON regulation of the trityl deprotection reaction in NetState-I and II. (b) Reversible switching between the network states furnished reproducible amounts of product 7 in NetState-II (two independent runs). Consumed amounts of substrates were added (green asterisk).The OFF–ON switching of the catalytic trityl deprotection of aniline 6 by the proton relay network inspired us to utilize the system to catalytically fabricate a slider-on-deck based on the N_an (= aniline) → ZnPor interaction. Deprotection of the protected biped 4 should afford the bis-aniline biped 5 and enable its attachment to the tris-zinc porphyrin deck 3 thus forming the slider-on-deck [(3)(5)].Addition of one equiv. of biped 5 (for synthesis and characterization, see ESI^†) to deck 3 (ref. 25) in CD₂Cl₂ quantitatively generated the slider-on-deck [(3)(5)], which was unambiguously characterized by ¹H NMR, ¹H DOSY NMR and elemental analysis (ESI, Fig. S5 and S6^†). In the ¹H NMR-spectrum, several diagnostic shifts of biped and deck proton signals attested the binding of biped 5 to the deck. Formation of the slider-on-deck was further confirmed from a single set in the ¹H DOSY in CD₂Cl₂ (ESI, Fig. S9^†).The single set of protons (p-H, r-H, s-H, t-H, u-H, v-H) in the ¹H NMR spectrum for all three-zinc porphyrin (ZnPor) stations of deck 3 unmistakably suggested rapid intrasupramolecular exchange^29a of the aniline biped across all three ZnPor sites requiring fast N_an → ZnPor bond cleavage. At low temperature (−65 °C) the ZnPor p-H proton signals separated into two sets (1 : 2) (ESI, Fig. S7 and S8^†). Kinetic analysis over a wide temperature range provided the exchange frequency (k) with k₂₉₈ = 540 kHz at 298 K (Fig. 5a) and the activation data as ΔH^‡ = 41.5 ± 0.7 kJ mol⁻¹, ΔS^‡ = 2.8 ± 0.7 J mol⁻¹ K⁻¹ and ΔG^‡₂₉₈ = 40.7 kJ mol⁻¹ (Fig. 5).Open in a separate windowFig. 5(a) Experimental and theoretical splitting of p-H proton signal of nanoslider [(3)(5)] in VT ¹H-NMR (600 MHz) furnishing rate data in CD₂Cl₂. (b) Eyring plot providing kinetic parameters.After determining the kinetic parameters of the slider-on-deck [(3)(5)], we investigated its in situ catalyzed formation by the proton relay network. Therefore, we mixed nanoswitch 1, luminophore 2, TFA, trityl-protected biped 4, deck 3 and TMB in 1 : 1 : 1 : 5 : 5 : 5 ratio in CD₂Cl₂ and heated the reaction mixture for 12 h at 40 °C. The ¹H NMR spectrum suggested no formation of any slider-on-deck in NetState-I (ESI, Fig. S13^†). Addition of one equiv. of AgBF₄ to the same mixture (converting NetState-I to NetState-II) and heating it at 40 °C for 12 h revealed full formation of the slider-on-deck as indicated by the disappearance of the 2′-H, 3′-H and 4′-H proton signals of biped 4 at 6.60, 6.31 and 6.51 ppm (ESI, Fig. S11^†), respectively, and the quantitative emergence of 4′-H proton signal of biped 5 bound to deck 3 at 5.84 ppm (ESI, Fig. S12^†). The upfield shift of the deck''s meso t-H protons from 10.36 to 10.29 ppm allowed monitoring of the formation of [(3)(5)], e.g. by the gradually increased chemical shift difference of proton signal t-H (Δδ_t-H) (Fig. 6a; ESI, Fig. S11^†). In sum, the protonated [H(2)]⁺ in NetState-II catalyzed the deprotection of the trityl-protected biped 4 with the effect that the resultant bis-aniline biped 5 would quantitatively bind to deck 3 affording the slider-on-deck [(3)(5)]. Furthermore, the OFF/ON switching of catalysis was probed for shortened time periods of 3 h. As illustrated in Fig. 6b, the process could be readily followed using the proton signal t-H (Δδ_t-H) (ESI, Fig. S14^†) and the growth of proton signal 4′-H at 5.84 ppm of bound biped 5 (ESI, Fig. S15^†). One clearly sees recurring OFF/ON regulation in NetState-I/II.Open in a separate windowFig. 6Plot of the shift difference of proton signal t-H (Δδ_t-H) vs. time for (a) the formation of [(3)(5)] from 3 and 4. Red curve: formation of [(3)(5)] over 12 h catalyzed by NetState-II. Black curve: OFF state of catalysis in NetState-I. (b) OFF/ON catalytic cycles by switching between NetState-I and NetState-II. Addition of Ag⁺, see blue asterisk; addition of TBAI: red asterisk.In conclusion, we have designed a supramolecular multicomponent network³⁶ that acts as an OFF/ON proton relay with a luminescence display³⁷ and is useful for switchable catalysis.³⁸ The network is toggled by chemical input and intercomponent communication^15,16 and as a result is a conceptual complement to photoacids driving networks.^7,8The release and capture of protons is demonstrated by the ON/OFF trityl deprotection of anilines. To demonstrate functioning in a complex setting, the network was utilized to catalyze formation of a high-speed slider-on-deck assembly based on N_an → ZnPor interactions (sliding frequency of k₂₉₈ = 540 kHz). The robust operation of the proton relay furnishing dynamic machinery³⁹ through acid catalysis followed by self-assembly is a valuable step mimicking sophisticated biological proton relays.²