Production of γ-terpinene by metabolically engineered Escherichia coli using glycerol as feedstock

Gamma (γ)-terpinene, a monoterpene compound, which is generally used in the pharmaceutical and cosmetics industries, due to its physical and chemical properties, is expected to become one of the more influential compounds used as an alternative biofuel in the future. It is necessary to seek more sustainable technologies such as microbial engineering for γ-terpinene production. In this study, we metabolically engineered Escherichia coli to produce γ-terpinene by introducing a heterologous mevalonate (MVA) pathway combined with the geranyl diphosphate synthase gene and γ-terpinene synthase gene. Subsequently, the culture medium and process conditions were optimised with a titre of 19.42 mg L⁻¹ obtained. Additionally, in-depth analysis at translation level for the engineered strain and intermediate metabolites were detected for further analysis. Finally, the fed-batch fermentation of γ-terpinene was evaluated, where a maximum concentration of 275.41 mg L⁻¹ with a maintainable feedstock of glycerol was achieved.

A sustainable technology for gamma (γ)-terpinene was constructed in an engineered Escherichia coli. In-depth analysis at translation level for the engineered strain and intermediate metabolites were analyzed. The fed-batch fermentation of γ-terpinene was 275.41 mg L⁻¹.     

α-Glucosidase inhibitors from Chinese bayberry (Morella rubra Sieb. et Zucc.) fruit: molecular docking and interaction mechanism of flavonols with different B-ring hydroxylations

Inhibition of α-glucosidase alleviates postprandial high glycemic levels in diabetic or prediabetic population. In Chinese bayberry fruit, myricetin, quercetin and kaempferol are main flavonols, which differ only in their hydroxylation on the B-ring. Kaempferol (4′-OH) showed high IC₅₀ (65.36 ± 0.27 μmol L⁻¹) against α-glucosidase, while quercetin (3′,4′-OH) exhibited stronger inhibition (46.91 ± 0.54 μmol L⁻¹) and myricetin (3′,4′,5′-OH) possessed the strongest inhibitory activity (33.20 ± 0.43 μmol L⁻¹). Molecular docking analysis illustrated that these flavonols could insert to the active cavity of α-glucosidase. Adjacent hydroxyl groups at B-ring of myricetin and quercetin positively contributed to form hydrogen bonds that were important to the stability of flavonol–enzyme complex, while kaempferol had no adjacent hydroxyl groups. Such observation was further validated by molecular dynamics simulations, and in good consistency with in vitro kinetic analysis and fluorescence spectroscopy analysis. Among three flavonols tested, myricetin possessed the strongest inhibition effects on α-glucosidase with the lowest dissociation constant (K_i = 15.56 μmol L⁻¹) of myricetin-α-glucosidase, largest fluorescence quenching constant (K_sv) of (14.26 ± 0.03) × 10⁴ L mol⁻¹ and highest binding constant (K_a) of (1.38 ± 0.03) × 10⁵ L mol⁻¹ at 298 K with the enzyme. Bio-Layer Interferometry (BLI) and circular dichroism (CD) analysis further confirmed that myricetin had high affinity to α-glucosidase and induced conformational changes of enzyme. Therefore, myricetin, quercetin and kaempferol are all excellent dietary α-glucosidase inhibitors and their inhibitory activities are enhanced by increasing number of hydroxyl groups on B-ring.

Inhibition of α-glucosidase alleviates postprandial high glycemic levels in diabetic or prediabetic population.     

Lymphotoxin-α (LTα) Supports Development of Splenic Follicular Structure That Is Required for IgG Responses

LTα-deficient (LTα^−/−) mice show altered splenic microarchitecture. This includes loss of normal B cell–T cell compartmentalization, of follicular dendritic cell (FDC) clusters, and of ability to form germinal centers (GC). LTα^−/− mice immunized with sheep red blood cells (SRBC) produced high levels of antigen-specific IgM but no IgG in either primary or secondary responses, demonstrating failure of Ig class switching. This inability to switch to IgG could have been due to the altered splenic microarchitecture in these mice. Alternatively, it could have been due directly to a requirement for LTα expression by lymphocytes cooperating in the antibody response. To investigate this, we performed reciprocal spleen cell transfers. When irradiated LTα^−/− mice were reconstituted with wild-type splenocytes and immunized immediately with SRBC, splenic microarchitecture remained disturbed and there was no IgG response. In contrast, when irradiated wild-type animals received splenocytes from LTα^−/− mice, follicle structure and a strong IgG response were retained. These data indicate that LTα-deficient B cells and T cells have no intrinsic defect in ability to generate an IgG response. Rather, the altered microenvironment characteristic of LTα^−/− mice appears to result in impaired ability to switch to a productive IgG response. To investigate whether prolonged expression of LTα could alter the structure and function of spleen follicles, reciprocal bone marrow (BM) transplantation was performed. Six weeks after reconstitution of LTα^−/− mice with wild-type BM, spleen follicle structure was partially restored, with return of FDC clusters and GC. B cell/T cell compartmentalization remained abnormal and white pulp zones were small. This was accompanied by restoration of IgG response to SRBC. Reconstitution of wild-type mice with LTα^−/− BM resulted in loss of FDC clusters and GC, and loss of the IgG response, although compartmentalized B cell and T cell zones were largely retained. Thus, defective IgG production is not absolutely associated with abnormal B cell and T cell compartmentalization. Rather, expression of LTα supports the maturation of spleen follicle structure, including the development and maintenance of FDC clusters, which supports Ig class switching and an effective IgG response.Lymphotoxin-α (LTα)¹ shares structural features with the related cytokine, TNFα. Both LTα and TNFα exist in solution as homotrimeric proteins. In these forms, they share biological activities by virtue of their similar binding to the two defined TNF receptors, TNFR-I, and TNFR-II. Signaling via these two receptors modulates a wide variety of immune and inflammatory responses (1, 2). LTα also exists in a heteromeric form with the type II membrane protein LTβ, which in its most prevalent form on the membrane has the stoichiometry LTα₁LTβ₂. The LTα₁LTβ₂ heteromer has no measurable affinity for TNFR-I or TNFR-II, but does interact with high affinity with the TNFR-related protein (also designated the LTβ receptor, or LTβR) (3–5).LTα^−/− mice are born with defective development of LN and Peyer''s patches (PP) (6). In LTα^−/− mice, spleen structure is also disturbed, with small white pulp follicles that fail to segregate T cell and B cell zones, and fail to generate clusters of FDC or GC (6–8). Proper spleen microarchitecture, including the presence of primary and secondary lymphoid follicles that contain FDC, is thought to be required for all features of a mature T cell–dependent B cell response, including Ig class switching, affinity maturation, and development of antibody secreting cells (9–11). Consequently, it was striking that LTα^−/− mice were able to generate high affinity anti-NP IgG antibody after immunization with high doses of the T cell–dependent antigen 4-hydroxy3-nitrophenyl-ovalbumin (NP-OVA) adsorbed to alum (8). In contrast, there was impaired production of high affinity anti-NP IgG when LTα^−/− mice were immunized with low doses of NP-OVA absorbed to alum. Banks et al. (12) using an independently derived LTα^−/− mouse strain, have shown impaired IgG responses in LTα^−/− mice after subcutaneous immunization with KLH absorbed to alum or immunization with viral antigens. Further studies examining mice deficient in both TNFα and LTα demonstrated variable IgG responses, with deficient IgG responses after intraperitoneal immunization with SRBC but retained responses against vesicular stomatitis virus after an infectious challenge (13). Recently prepared TNFα^−/− mice (14) are reported also to have an impaired IgG anti-SRBC response but a strong response to DNP-KLH adsorbed to alum. Thus, the role of LTα in supporting an effective IgG response to SRBC remains incompletely defined.LTα^−/− mice manifest a complex phenotype that includes an absence of LTα expression and also established abnormalities of lymphoid tissue development and structure (6–8). The present study was undertaken to investigate the requirements for LTα expression in lymphoid cells and for intact lymphoid tissue structure to support production of an IgG responses to the T cell–dependent antigen SRBC. We report that LTα^−/− mice responded with high levels of IgM but very low levels of IgG after immunization with SRBC in the absence of adjuvant. Experiments in which suspensions of mature spleen cells or of  T cell–depleted bone marrow were transferred to wild-type or LTα^−/− mice demonstrated that certain elements of spleen follicle structure are plastic and are determined by the presence of LTα-expressing cells. These experiments also demonstrated that LTα^−/− B cells and T cells in a structurally intact lymphoid tissue environment are competent to perform Ig isotype switching. In contrast, disturbed lymphoid tissue structure caused by absence of LTα and manifested by absence of clusters of FDC is associated with an inability to form an effective IgG response. Thus, LTα produced by bone marrow (BM)–derived cells establishes a permissive environment for an effective IgG response.     

Design,synthesis, and bioactivity evaluation of antitumor sorafenib analogues

Malignant tumors are a serious threat to human health and are generally treated with chemical therapy. This chemical therapy uses agents that act on signal transduction pathway mechanism of tumor with good selectivity and low toxicity. Sorafenib is a multikinase target inhibitor with good tumor inhibitory activity and a protein kinase inhibitor. In this research, a novel series of sorafenib analogues and derivatives were designed, synthesized, and evaluated as tumor inhibitors. These compounds used sorafenib as the lead compound and achieved modifications using bioisosteres and the alkyl principle. The in vitro the results showed that compounds 3c, 3d, 3h, 3n, 3r, and 3z had good inhibitory effects on human cervical cancer cells (Hela), while compounds 3t and 3v had good inhibitory effects on human lung cancer cells (H1975 and A549). Among these, compound 3d had an inhibitory activity (IC₅₀) of 0.56 ± 0.04 μmol L⁻¹ against Hela cells (human cervical cancer), the compound 3t had an IC₅₀ of 2.34 ± 0.07 μmol L⁻¹ against H1975 cells (human lung cancer), and compound 3v had an IC₅₀ of 1.35 ± 0.03 μmol L⁻¹ against A549 cells (human lung cancer). The in vivo results showed that these compounds had good antitumor effects and low acute toxicity.

Malignant tumors are a serious threat to human health and are generally treated with chemical therapy.     

Gum acacia-based silver nanoparticles as a highly selective and sensitive dual nanosensor for Hg(ii) and fluorescence turn-off sensor for S2− and malachite green detection

A facile and green method was adopted to synthesize highly selective gum acacia-mediated silver nanoparticles as dual sensor (fluorescence turn-on and colorimetric) for Hg(ii) and fluorescence turn-off sensor for S²⁻ and malachite green. The mechanism proposed for a dual response towards Hg(ii) is the redox reaction between Ag(0) and Hg(ii), resulting in the formation of Ag(i) and Hg(0) and electron transfer from gum acacia to Ag(i), which further leads to the formation of an Ag@Hg nanoalloy. The enhanced fluorescence signal was quenched selectively by S²⁻ owing to the formation of Ag₂S and HgS. The reported nanosensor was found to be useful for sensing malachite green via the inner filter effect. The linear ranges were 3 nmol L⁻¹ to 13 μmol L⁻¹ for Hg(ii), 3–170 μmol L⁻¹ for S²⁻ and 7–80 μmol L⁻¹ for malachite green, and the corresponding detection limits were 2.1 nmol L⁻¹ for Hg(ii), 1.3 μmol L⁻¹ for S²⁻ and 1.6 μmol L⁻¹ for malachite green.

Gum acacia-stabilized silver nanoparticles for the detection of Hg(ii), S²⁻ and malachite green.     

A thio-β-cyclodextrin functionalized graphene/gold nanoparticle electrochemical sensor: a study of the size effect of the gold nanoparticles and the determination of tetrabromobisphenol A

In this study, a novel tetrabromobisphenol A (TBBPA) sensor was fabricated based on a CTAB-capped gold nanoparticle (AuNPs)-thio-β-cyclodextrin (SH-β-CD)/graphene oxide modified glassy carbon electrode (GCE). The peak current of TBBPA was dramatically enhanced by the AuNPs with a diameter of 6.2 nm on the modified electrodes compared with the other sized particles (10.1 or 16.1 nm). To further improve the electrochemical performance of the modified electrode, the influence of pH of the buffer solution and the accumulation time on the determination were investigated. The optimum pH and accumulation time were 7.0 and 180 s, respectively. The developed sensor exhibited good reproducibility, and excellent sensitivity and selectivity, showing a low detection limit (1.2 × 10⁻⁹ mol L⁻¹) and a linear range from 1.5 × 10⁻⁸ to 7 × 10⁻⁶ mol L⁻¹. In addition, a possible oxidization mechanism of TBBPA was also discussed. Finally, this sensor was successfully applied to detect TBBPA in water samples, and the results were consistent with those acquired by high-performance liquid chromatography.

In this study, a novel tetrabromobisphenol A (TBBPA) sensor was fabricated based on a CTAB-capped gold nanoparticle (AuNP)-thio-β-cyclodextrin (SH-β-CD)/graphene oxide modified glassy carbon electrode (GCE).     

Continuous hydrogen production from glucose/xylose by an anaerobic sequential batch reactor to maximize the energy recovery efficiency

Fermentation of both glucose and xylose is essential to realize efficient bioconversion of renewable and abundant lignocellulosic biomass to hydrogen. In this study, a mixture of glucose and xylose at different ratios was used as a substrate for biological hydrogen production by an anaerobic sequential batch reactor (ASBR). An average glucose and xylose consumption of 80% and 50% with a high hydrogen production rate of 7.1 ± 0.9 mmol L⁻¹ h⁻¹ was obtained, respectively. Hydraulic retention time (HRT) played a critical role in hydrogen production at high glucose to xylose ratios. A maximum hydrogen production rate of 8.9 mmol L⁻¹ h⁻¹ was achieved at an optimized HRT of 12 h with a high glucose and xylose consumption of 92.2% and 82.2%, respectively. Upon further energy conversion analysis, continuous hydrogen production by ASBR provided the maximum energy conversion efficiency of 21.5%. These results indicate that ASBR can effectively accelerate the hydrogen production rate, improve substrate consumption regardless of the glucose to xylose ratio, and thus provides a new direction for efficient hydrogen production from lignocellulosic feedstock.

Fermentation of both glucose and xylose is essential to realize efficient bioconversion of renewable and abundant lignocellulosic biomass to hydrogen.     

Thermally controlled biotransformation of glycyrrhizic acid via an asymmetric temperature-responsive polyurethane membrane

Separating a target product from a relatively complex bioreaction system is often difficult. In this work, a “smart” bioreaction system was developed by using the special characteristic of temperature-responsive polyurethane (TRPU). By combining solvent evaporation with a wet phase inversion technique, an asymmetric membrane consisting of an integral and dense skin layer supported by a porous sublayer was prepared from a thermally responsive polyurethane that experiences a sudden free volume increase upon heating through a phase transition temperature of 56 °C. Subsequently, the asymmetric TRPU membrane served as the carrier of an immobilized enzyme, wherein β-glucuronidase was multipoint-conjugated by using biotin and streptavidin on the porous sublayer. Then, the material-asymmetric TRPU membrane served jointly as the antennae as well as the actuator, which reversibly responds to temperature to switch (on–off) the access of the reactant glycyrrhizic acid (GL). Under the optimal temperature (40 °C) and pH (7.0) conditions, the immobilized β-glucuronidase contributed to almost 33% yield of glycyrrhetinic acid 3-O-mono-β-d-glucuronide (GAMG) of the isolated counterpart for the same concentration of substrate (250 mg L⁻¹) reaction for 24 h, while costing 1% of that of the isolated β-glucuronidase. Kinetic results showed that V_max and K_m values were 8.89 × 10³ mg L⁻¹ and 2.30 × 10³ mg L⁻¹ h⁻¹, respectively. The specific functional polymer-immobilized β-glucuronidase design serves as a bioreactor of GL into GAMG, as well as a separator deliberately irritated and controlled by temperature. This “smart” support material presents a potential facilitator for the separation of complex biotransformation reactions.

A “smart” bioreaction system was developed by using the special characteristic of a temperature-responsive polyurethane (TRPU). This “smart” support material presents a potential benefit of separation for complex biotransformation reactions.     

Fabrication of an (α-Mn2O3:Co)-decorated CNT highly sensitive screen printed electrode for the optimization and electrochemical determination of cyclobenzaprine hydrochloride using response surface methodology

A new chemically optimized screen-printed electrode modified with a cobalt-doped α-Mn₂O₃ nanostructure on carbon nanotube paste (α-Mn₂O₃:Co@CNTs) has been constructed for the recognition of cyclobenzaprine hydrochloride. The prepared paste is based on the incorporation of oxide ion conductors, such as the α-Mn₂O₃ nanostructure with cobalt and ion pairs (tetraphenyl borate coupled with the drug), as electroactive species in the screen-printed electrode to increase the sensor surface area and decrease electrical resistance. The central composite design is a useful methodology for the estimation and modeling of the exact optimum parameters specifically designed for this process. This is a good way to graphically clarify the relationship between various experimental variables and the slope response. The proposed sensor, α-Mn₂O₃:Co@CNTs, possesses very good sensitivity and the ability to recognize the drug over the concentration range of 1 × 10⁻⁶ to 1 × 10⁻² mol L⁻¹ at 25 ± °C with a detection limit of 2.84 × 10⁻⁷ mol L⁻¹. It exhibits a reproducible potential and stable linear response for six months at a Nernstian slope of 58.96 ± 0.76 mV per decade. The proposed electrode approach has been successfully applied in the direct determination of the drug in its pure and dosage forms.

A new chemically optimized screen-printed electrode modified with a cobalt-doped α-Mn₂O₃ nanostructure on carbon nanotube paste (α-Mn₂O₃:Co@CNTs) has been constructed for the recognition of cyclobenzaprine hydrochloride.     

Highly sensitive determination of lead(ii) and cadmium(ii) by a large surface area mesoporous alumina modified carbon paste electrode

Nanosized mesoporous γ-alumina (M-γ-Al₂O₃) was first prepared and then modified into a carbon paste to fabricate a novel modified carbon paste electrode. The prepared alumina has pores with an amorphous wall and large surface area. The electrochemical behavior of the modified carbon paste electrode was investigated using cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) methods. The modified carbon paste electrode was employed to determine Pb²⁺ and Cd²⁺ simultaneously by a differential pulse voltammetry (DPV) method. Amperometric determination was carried out in 0.1 mol L⁻¹ NaAc–HAc buffer solution (pH 6.0) after enriching for 360 s at −1.0 V. The oxidation peak currents of Pb²⁺ and Cd²⁺ were proportional to their concentration in the range of 0.001–10 μmol L⁻¹ and 0.01–10 μmol L⁻¹, respectively. The detection limits of Pb²⁺ and Cd²⁺ were 0.20 nmol L⁻¹ and 2.0 nmol L⁻¹ (S/N = 3), respectively. The modified carbon paste electrode shows good stability, repeatability and sensitivity. The proposed method was applied to the determination of Pb²⁺ and Cd²⁺ in water samples with satisfactory results.

Nanosized mesoporous γ-alumina (M-γ-Al₂O₃) was first prepared and then modified into a carbon paste to fabricate a novel modified carbon paste electrode.     

A simple Fe3+/bisulfite system for rapid degradation of sulfamethoxazole

Sulfate radical (SO₄˙⁻) based oxidation technologies have been widely used in the remediation of antibiotic-containing wastewater. Activated persulfates are efficient reagents for achieving SO₄˙⁻, but the storage and transportation of concentrated persulfates present associated safety issues. In this study, bisulfite (BS) was used as an alternative precursor for replacing persulfates, and a simple advanced oxidation system (Fe³⁺/BS) for generating SO₄˙⁻ and hydroxyl radical (HO˙) was formulated and evaluated for removing sulfamethoxazole (SMX) from contaminated water. The initial pH, dosages of Fe³⁺ and BS, as well as the water matrix were investigated to improve the SMX degradation. The results indicated that 1 μmol L⁻¹ SMX was completely removed within 5 min at optimum initial pH of 4.0, Fe³⁺ dosage of 10 μmol L⁻¹, BS dosage of 100 μmol L⁻¹ and temperature of 25 °C. The presence of HCO₃⁻ and natural organic matter (NOM) showed obviously negative effects on SMX degradation, while Cu²⁺ could slightly promote the degradation of SMX if its concentration was in an appropriate range (∼1 μmol L⁻¹). Scavenger quenching experiments confirmed the presence of SO₄˙⁻ and HO˙, which resulted in efficient SMX degradation in the Fe³⁺/BS system. During the radical chain reactions, Fe²⁺ and Fe³⁺ could be converted into each other to form self-circulation in this system. The degradation pathway of SMX by Fe³⁺/BS was proposed including hydroxylation and bond cleavage.

Sulfate radical (SO₄˙⁻) based oxidation technologies have been widely used in the remediation of antibiotic-containing wastewater.     

Quantification and isotherm modelling of competitive phosphate and silicate adsorption onto micro-sized granular ferric hydroxide

Adsorption onto ferric hydroxide is a known method to reach very low residual phosphate concentrations. Silicate is omnipresent in surface and industrial waters and reduces the adsorption capacity of ferric hydroxides. The present article focusses on the influences of silicate concentration and contact time on the adsorption of phosphate to a micro-sized iron hydroxide adsorbent (μGFH) and fits adsorption data to multi-component adsorption isotherms. In Berlin drinking water (DOC of approx. 4 mg L⁻¹) at pH 7.0, loadings of 24 mg g⁻¹ P (with 3 mg L⁻¹ initial PO₄³⁻–P) and 17 mg L⁻¹ Si (with 9 mg L⁻¹ initial Si) were reached. In deionized water, phosphate shows a high percentage of reversible bonds to μGFH while silicate adsorption is not reversible probably due to polymerization. Depending on the initial silicate concentration, phosphate loadings are reduced by 27, 33 and 47% (for equilibrium concentrations of 1.5 mg L⁻¹) for 9, 14 and 22 mg L⁻¹ Si respectively. Out of eight tested multi-component adsorption models, the Extended Freundlich Model Isotherm (EFMI) describes the simultaneous adsorption of phosphate and silicate best. Thus, providing the means to predict and control phosphate removal. Longer contact times of the adsorbent with silicate prior to addition of phosphate reduce phosphate adsorption significantly. Compared to 7 days of contact with silicate (c₀ = 10 mg L⁻¹) prior to phosphate (c₀ = 3 mg L⁻¹) addition, 28 and 56 days reduce the μGFH capacity for phosphate by 21 and 43%, respectively.

Adsorption of phosphate onto ferric hydroxide in complex waters is influenced by effects of competition, displacement and surface blockage.     

IL-15 trans-presentation promotes human NK cell development and differentiation in vivo

The in vivo requirements for human natural killer (NK) cell development and differentiation into cytotoxic effectors expressing inhibitory receptors for self–major histocompatability complex class I (MHC-I; killer Ig-like receptors [KIRs]) remain undefined. Here, we dissect the role of interleukin (IL)-15 in human NK cell development using Rag2^−/−γc^−/− mice transplanted with human hematopoietic stem cells. Human NK cell reconstitution was intrinsically low in this model because of the poor reactivity to mouse IL-15. Although exogenous human IL-15 (hIL-15) alone made little improvement, IL-15 coupled to IL-15 receptor α (IL-15Rα) significantly augmented human NK cells. IL-15–IL-15Rα complexes induced extensive NK cell proliferation and differentiation, resulting in accumulation of CD16⁺KIR⁺ NK cells, which was not uniquely dependent on enhanced survival or preferential responsiveness of this subset to IL-15. Human NK cell differentiation in vivo required hIL-15 and progressed in a linear fashion from CD56^hiCD16⁻KIR⁻ to CD56^loCD16⁺KIR⁻, and finally to CD56^loCD16⁺KIR⁺. These data provide the first evidence that IL-15 trans-presentation regulates human NK cell homeostasis. Use of hIL-15 receptor agonists generates a robust humanized immune system model to study human NK cells in vivo. IL-15 receptor agonists may provide therapeutic tools to improve NK cell reconstitution after bone marrow transplants, enhance graft versus leukemia effects, and increase the pool of IL-15–responsive cells during immunotherapy strategies.NK cells participate in host protection by eliminating cells with altered expression of self–MHC class I (MHC-I), which can result from viral infection or transformation (1). Although we are beginning to appreciate a role for viral and stress-induced ligands in NK cell activation, the best described regulatory mechanism of NK cell activity is the expression of inhibitory receptors for self–MHC-I ligands by mature NK cells with high cytotoxic potential. In man, killer Ig-like receptors (KIRs) recognizing the classical MHC-I molecules HLA-A, -B, or -C are expressed on the predominate peripheral NK cell (CD56^loCD16⁺) subset, which possesses abundant intracellular perforin and granzymes and displays spontaneous cytotoxicity (1). In contrast, CD56^hiCD16⁻ NK cells rarely express KIRs, and because they are more prevalent in blood early after bone marrow transplant (2, 3), give rise to CD56^loCD16⁺ NK cells when transferred into NOD/SCID mice (4), and have longer telomeres than CD56^loCD16⁺ NK cells (5), it is likely that CD56^hiCD16⁻ NK cells are (or contain within this population) precursors of CD56^loCD16⁺ NK cells; however, the tools to definitively prove this hypothesis are lacking.Although inhibitory KIRs control reactivity of mature NK cells, their expression also influences the functional maturation of developing NK cells, as NK cells expressing at least one KIR-recognizing self–MHC-I have a lower threshold of activation and appear more functional than NK cells expressing no KIRs or those only expressing KIRs recognizing non-self–MHC-I ligands (6). In addition, patients lacking the transporter associated with antigen processing have dramatically reduced MHC-I surface expression and, consequently, hyporesponsive NK cells to MHC-I–deficient cells (7). KIR⁺ NK cells are present in these patients, indicating that normal MHC-I expression itself is not required for KIR expression (7). This phenomenon termed “licensing” or “disarming” has been better characterized in mice, and suggests a role for KIR–self–MHC-I interactions during human NK cell development. Given the importance of KIR expression in regulating NK cell function, knowledge of elements influencing KIR acquisition would improve our understanding and clinical approaches to diseases where KIR and HLA haplotypes influence susceptibility, progression, or outcome of autoimmune/inflammatory disease, cancer, hematopoietic grafts, and infections (8).Because KIRs are expressed on mature CD56^loCD16⁺ NK cells, factors that influence NK cell homeostasis could potentially influence KIR acquisition in vivo. An elegant series of mouse studies using gene targeting and bone marrow chimeras have revealed that NK cell development requires IL-15Rα–expressing cells to chaperone IL-15 to the surface, where it is bioactive and significantly more potent in inducing activation and proliferation of IL-15Rβγ–expressing cells via trans-presentation (9, 10). When not bound to IL-15Rα, IL-15 appears to have a minimal effect on NK cell homeostasis in vivo (11, 12). In humans, mutations in IL-15Rα have not been reported; however, NK cells are dramatically reduced in patients carrying mutations in the common γ chain (γc) cytokine receptor (used in IL-15, -7, -4, -9, -2, and -21 signal transduction), Jak3, or the shared IL-2/-15Rβ, whereas they are present in IL-7Rα–deficient patients, suggesting that IL-15 may regulate human NK cell development (13, 14).In vivo studies of NK cells have been largely restricted to mice, and although this line of experimentation is valuable, some of this knowledge is not transferable to human NK cell biology. A clear example of this is NK cell development, where the kinetics, frequency, and phenotype are clearly different between species (1). An intermediate between mouse and human in vivo studies exists in the form of human immune system (HIS) mice. A recently developed HIS mouse model is the engraftment of newborn BALB/c Rag2^−/−γc^−/− mice with human hematopoietic stem cells (HSCs) from fetal liver or cord blood (15, 16). BALB/c Rag2^−/−γc^−/− HIS mice represent a practical HIS model with high human chimerism, most lymphocyte lineages generated, and adaptive immune responses occasionally evoked (15, 16).In this study, using HIS BALB/c Rag2^−/−γc^−/− mice engrafted with fetal liver HSCs, we study the in vivo role of IL-15 trans-presentation in human NK cell homeostasis.     

Effect of ion form of the ion-exchange resin on ε-poly-l-lysine purification from microbial fermentation broth

ε-Poly-l-lysine (ε-PL) is an added-value natural product with widespread application in the fields of food, pharmaceuticals and biopolymer materials. However, the high production cost reduces its application. To improve the efficiency of ε-PL purification for decreasing the cost of downstream processes, the ion form of the ion-exchange resin, which is widely used for ε-PL purification, was investigated systematically in this study. Among eleven cation-exchange resins tested, the Amberlite IRC-50 resin offered the best adsorption capability and the highest desorption ratio. The adsorption kinetics of IRC-50 resin with H⁺, Na⁺ and NH₄⁺ ion forms followed a pseudo-second-order model. The dynamic adsorption and desorption parameters of ε-PL were optimized with a column packed with IRC-50 resin with Na⁺ and NH₄⁺. It is suggested that NH₄⁺ is the optimal ion form of IRC-50 resin for ε-PL extraction. Under optimal conditions, the IRC-50 resin with NH₄⁺ achieved the highest ε-PL adsorption capability, purity and recovery ratio of 307.96 mg g⁻¹, 76.52% and 96.2%, respectively. After further purification, a final ε-PL purity of 97.10% was achieved with a total recovery ratio of 66.01%. This is the first report on improving the ε-PL purification efficiency through optimizing the ion form of the ion-exchange resin. Moreover, it would offer guidance for other natural product recovery processes by ion-exchange chromatography.

This is the first report on improving the ε-PL purification efficiency through optimizing the ion form of the ion-exchange resin.     

An assessment of an ion exchange resin system for the removal and recovery of Ni,Hg, and Cr from wet flue gas desulphurization wastewater—a pilot study

The paper presents the results of a pilot-scale study investigating the efficiency of an ion exchange resin system in the removal of Ni, Hg, and Cr from flue gas desulphurisation wastewater, in the presence of competitive metals such as Ca, Mg, Al, Fe, and Mn. The core part of the ion exchange installation consisted of two columns that were filled with ion exchange resin with iminodiacetic functional groups (Purolite S930) and one column filled with resin with isothiouronium functional groups (Purolite S920). The results showed that Ni, Hg, and Cr were almost completely removed from the wastewater with nearly 100% efficiency. Purolite S930 almost totally removed Ni, reducing its content from 89.3 ± 35 μg dm⁻³ to below 0.1 μg dm⁻³, while Purolite S920 reduced the remaining Cr content from 2.2 ± 0.6 μg dm⁻³ and most of the Hg content, from 23.5 ± 6.6 μg dm⁻³ to below 0.1 μg dm⁻³. The competitive metals Ca, Mg, Mn, and Al showed low affinity to the studied ion exchange resins. The study also assessed speciation of ion forms and sorption mechanisms. Breakthrough curve analysis was also carried out, which revealed that the selectivity sequence of iminodiacetic resin was Ni > Cr > Hg > Fe > Al > Mn > Ca, Mg. Elution studies were performed on S930 resins that allowed the separation of two streams: one containing mostly Ni and Fe which can be subjected to Ni recovery and the other containing mostly Cr and Hg which can be separated.

Pilot-scale study of an ion exchange resin system for the selective removal/recovery of Ni, Hg, and Cr from desulphurisation wastewater.     

Removal of manganous dithionate (MnS2O6) with MnO2 from the desulfurization manganese slurry

Manganese desulfurization has been increasingly explored, but the generated manganous dithionate (MD) by-product affects the valuable use of the desulfurized slurry. In this study, α-MnO₂, β-MnO₂, γ-MnO₂, and δ-MnO₂ were prepared for MD removal in desulfurization manganese slurry. Results showed that δ-MnO₂ had the best activity among the four because of its porosity and favorable surface properties. The operation conditions showed that 12.00 g L⁻¹ MD can be removed by more than 80.00% under the conditions of 1.4 mol L⁻¹ sulfuric acid, 100 g L⁻¹ δ-MnO₂ dosage, and reaction at 90 °C for 3 h. The MD removal with MnO₂ followed the decomposition–oxidation pass and direct oxidation–reduction reaction and consequently induced structure destruction and crystalline transfer. MD removal with natural MnO₂ ore was also examined, and natural MnO₂ ore in the δ type was found to have prominent activity. Thus, this type of natural MnO₂ may serve as a good alternative to pure MnO₂ for decreasing the cost of MD removal from desulfurization manganese slurry.

The manganous dithionate by-product of the desulfurized slurry could be oxidized with MnO₂ without any impurity. σ-MnO₂ showed the best activity due to its high surface area and expose much more surface-active oxygen.     

Ratiometric fluorescence sensing of d-allulose using an inclusion complex of γ-cyclodextrin with a benzoxaborole-based probe

Because d-allulose has been attracting attention as a zero-calorie sugar, the selective sensing of d-allulose is desired to investigate its health benefits. We report herein a novel fluorescence chemosensor that is based on an inclusion complex of γ-cyclodextrin (γ-CyD) with a benzoxaborole-based probe. Two inclusion complexes, 1/γCyD and 2/γCyD, were prepared by mixing γ-CyD with their corresponding probes in a water-rich solvent, where γ-CyD encapsulates two molecules of the probes inside its cavity to form a pyrene dimer. Both 1/γCyD and 2/γCyD exhibit monomeric and dimeric fluorescence from the pyrene moieties. By the reaction of 1/γCyD with saccharides, the intensities of monomeric and dimeric fluorescence remained unchanged and decreased, respectively. We have demonstrated that 1/γCyD has much higher affinity for d-allulose than for the other saccharides (d-fructose, d-glucose, and d-galactose). The conditional equilibrium constants for the reaction systems were determined to be 498 ± 35 M⁻¹ for d-fructose, 48.4 ± 25.3 M⁻¹ for d-glucose, 15.0 ± 3.3 M⁻¹ for d-galactose, and (8.05 ± 0.59) × 10³ M⁻¹ for d-allulose. These features of 1/γCyD enable ratiometric fluorescence sensing with high sensitivity and selectivity for d-allulose. The limits of detection and quantification of 1/γCyD for d-allulose at pH 8.0 were determined to be 6.9 and 21 μM, respectively. Induced circular dichroism spectral study has shown that the reaction of 1/γCyD with d-allulose causes the monomerisation of the dimer of probe 1 that is encapsulated by γ-CyD, which leads to the diminishment of the dimeric fluorescence.

We proposed an inclusion complex of γ-cyclodextrin with a benzoxaborole-based fluorescent probe as a highly sensitive and selective chemosensor for d-allulose.