Substituted l-tryptophan-l-phenyllactic acid conjugates produced by an endophytic fungus Aspergillus aculeatus using an OSMAC approach

The endophytic fungus Aspergillus aculeatus isolated from leaves of the papaya plant Carica papaya was fermented on solid rice medium, yielding a new l-tryptophan-l-phenyllactic acid conjugate (1) and thirteen known compounds (11, 14–25). In addition, an OSMAC approach was employed by adding eight different sodium or ammonium salts to the rice medium. Addition of 3.5% NaNO₃ caused a significant change of the metabolite pattern of the fungus as indicated by HPLC analysis. Subsequent isolation yielded several new substituted l-tryptophan-l-phenyllactic acid conjugates (1–10) in addition to three known compounds (11–13), among which compounds 2–10, 12–13 were not detected in the rice control culture. All structures were unambiguously elucidated by one and two dimensional NMR spectroscopy and by mass spectrometry. The absolute configuration of the new compounds was determined by Marfey''s reaction and X-ray single crystal diffraction. Compounds 19–22 showed cytotoxicity against the L5178Y mouse lymphoma cell line with IC₅₀ values of 3.4, 1.4, 7.3 and 23.7 μM, respectively.

Indole-derived alkaloids from Aspergillus aculeatus using an OSMAC approach.     

Induction of cryptic metabolites of the endophytic fungus Trichocladium sp. through OSMAC and co-cultivation

The endophytic fungus Trichocladium sp. isolated from roots of Houttuynia cordata was cultured on solid rice medium, yielding a new amidepsine derivative (1) and a new reduced spiro azaphilone derivative (3) together with eight known compounds (4–11). Co-cultivation of Trichocladium sp. with Bacillus subtilis resulted in induction of a further new compound (2) and a 10-fold increase of 11 compared to the axenic fungal culture. Moreover, when the fungus was cultivated on peas instead of rice, a new sesquiterpene derivative (13) and two known compounds (12 and 14) were obtained. Addition of 2% tryptophan to rice medium led to the isolation of a new bismacrolactone (15). The structures of the new compounds were elucidated by HRESIMS, 1D and 2D NMR as well as by comparison with the literature. A combination of TDDFT-ECD, TDDFT-SOR, DFT-VCD and DFT-NMR calculations were applied to determine the absolute and relative configurations of 13 and 15. Compounds 7, 11 and 15 exhibited strong cytotoxicity against the L5178Y mouse lymphoma cell line with IC₅₀ values of 0.3, 0.5 and 0.2 μM, respectively.

The endophytic fungus Trichocladium sp. isolated from roots of Houttuynia cordata yielded fifteen compounds including five new ones through OSMAC and co-cultivation approaches.      

Expanding the chemical diversity of an endophytic fungus Bulgaria inquinans,an ascomycete associated with mistletoe,through an OSMAC approach

An endophytic fungus Bulgaria inquinans (isolate MSp3-1), isolated from mistletoe (Viscum album), was subjected to fermentation on solid Czapek medium. Chromatographic workup of the crude EtOAc extract yielded five new natural products (1–5). Subsequent application of the “One Strain, MAny Compounds” (OSMAC) strategy on this strain by the addition of a mixture of salts (MgSO₄, NaNO₃ and NaCl) to solid Czapek medium induced the accumulation of nine additional new secondary metabolites (6–13, 16), with most of them (8, 10–12) not detectable in cultures lacking the salt mixture. The structures of the new compounds were established on the basis of the 1D/2D NMR and HRESIMS data. The TDDFT-ECD method was applied to determine the absolute configurations of the new compounds 1, 4 and 6 as well as of the previously reported bulgarialactone B (14), for which the absolute configuration was unknown so far. The modified Mosher''s method was performed to assign the absolute configurations of 12 and 13. TDDFT-ECD analysis also allowed determining the absolute configuration of (+)-epicocconone, which had an enantiomeric absolute configuration in the tricyclic moiety compared to that of bulgarialactone B (14). All the isolated metabolites were evaluated for their cytotoxic activity. Compound 2 was found to possess strong cytotoxic activity against the murine lymphoma cell line L5178Y with an IC₅₀ value of 1.8 μM, while the remaining metabolites were shown to be inactive.

OSMAC approach on endophytic Bulgaria inquinans by addition of a mixture of salts (MgSO₄, NaNO₃ and NaCl) to solid Czapek medium induced the accumulation of new secondary metabolites.     

Versiquinazolines L–Q,new polycyclic alkaloids from the marine-derived fungus Aspergillus versicolor

Further chemical examination of a coral-associated fungus Aspergillus versicolor LZD-14-1 by the PHLC-DAD detection resulted in the isolation of six new polycyclic alkaloids, namely versiquinazolines L–Q (1–6). Their structures were determined by extensive analyses of spectroscopic data, including quantum ECD calculation and X-ray single crystal diffraction for the assignment of absolute configurations. Versiquinazoline L bearing a d-Ala residue and versiquinazoline M containing an l-serine residue are rarely found in the fumiquinazoline-type alkaloids, while versiquinazoline P displayed an unusual scaffold with a spiro-γ-lactone. Versiquinazolines P and Q exhibited significant inhibition against thioredoxin reductase (TrxR) with IC₅₀ values of 13.6 ± 0.6 and 12.2 ± 0.7 μM, which showed higher activity than the positive control curcumin (IC₅₀ = 25 μM). The weak cytotoxicity and potent inhibition toward TrxR suggested that versiquinazolines P and Q are potential for microenvironmental regulation of tumor progression and metastasis.

Further chemical examination of a coral-associated fungus Aspergillus versicolor LZD-14-1 by the PHLC-DAD detection resulted in the isolation of six new polycyclic alkaloids, namely versiquinazolines L–Q (1–6).     

Metabolomic profiling for the identification of novel diagnostic markers and therapeutic targets in prostate cancer: an update

Introduction: An altered metabolic regulation is involved in the development and progression of different cancer types. As well as this, many genes associated with tumors are shown to have an important role in control of the metabolism. The incidence of prostate cancer (PCa) is increased in men with metabolic disorders. In particular, obesity is an established risk factor for PCa. An increased body mass index correlates with aggressive disease, and a higher risk of biochemical recurrence and prostate cancer-specific mortality. Increased lipogenesis is also one of the most significant events in PCa metabolism reprogramming.

Areas covered: In this article, we provide an updated review of the current understanding of the PCa metabolome and evaluate the possibility of unveiling novel therapeutic targets.

Expert opinion: Obesity is an established risk factor for PCa, and an increased BMI correlates with aggressive disease, and a higher risk of biochemical recurrence and prostate cancer-specific mortality. PCa metabolome is characterized by the accumulation of metabolic intermediates and an increased expression of genes in the tricarboxylic acid cycle, the induction of de novo lipogenesis and cholesterogenesis. PCa cells can induce different alterations in their microenvironment by modulating the crosstalk between cancer and stromal cells.     

Metabolomic profiling of matured coconut water during post-harvest storage revealed discrimination and distinct changes in metabolites

The metabolites of coconut water stored at room temperature were analyzed using UPLC-MS/MS and multivariate statistical analysis to identify the differential biomarkers and metabolic pathways during post-harvest storage. Principal component analysis (PCA), partial least squares-discriminate analysis (PLS-DA), and orthogonal projections to latent structures discriminant analysis (OPLS-DA) were employed to analyze the UPLC-MS/MS data set of 34 matured coconut water samples collected after 0, 1, 2, 3, 4, and 5 months of storage (MOS); moreover, the p-value and fold change were chosen to identify the differential biomarkers; furthermore, a KEGG pathway was applied to analyze the metabolic pathways. All samples were discriminated well in the OPLS-DA model and were divided into two clusters: groups A (0 MOS, and so on), B, C, and D were in one cluster, and groups E and F were in another. A total of 18 biomarkers were identified among all groups and 12 biomarkers between groups A and E, from which we concluded that the post-harvest storage life of matured coconut water shall not exceed 3 months and the pathways of the TCA cycle, protein hydrolysis from coconut meat, and interconversion among amino acids were mainly enriched during the post-harvest storage.

The metabolites of coconut water stored at room temperature were analyzed using UPLC-MS/MS and multivariate statistical analysis to identify the differential biomarkers and metabolic pathways during post-harvest storage.     

Semi-synthesis,structural modification and biological evaluation of 5-arylbenzofuran neolignans

5-Arylbenzofuran neolignans, a newfound class of natural products, were reported to possess several kinds of pharmacological activities. To solve the lack of natural sources and promote the research of 5-arylbenzofuran neolignans in all fields, an available semi-synthesis methodology of 5-arylbenzofuran neolignans was developed, and a detailed structural modification was conducted. In the meantime, a one-pot process of Waker-type cyclization and Wacker-type oxidation was developed. To explore the potential of 5-arylbenzofuran neolignans as bioactive substances, 5-arylbenzofuran neolignans and their derivatives were evaluated for their cytotoxicity. As a result, a preliminary structure–activity relationship was obtained. Most derivatives revealed low cytotoxic effects suggesting that they were relatively safer than the natural 5-arylbenzofuran neolignan. Several derivatives showed high cytotoxicities which were found to be closely associated with apoptosis-inducing. The selectivity assay for cytotoxicity showed tumor cells were more sensitive to the promising compounds than normal cells.

5-Arylbenzofuran neolignans, a newfound class of natural products, were semi-synthesized, and a series of derivatives were designed, synthesized and evaluated for cytotoxicity.     

Seasonal metabolic profiling of Valencia orange leaf essential oil using GC coupled with chemometrics,nano-formulation,and insecticidal evaluation: in vivo and in silico

Mosquitoes and mosquito-borne infectious diseases are a global challenge, especially with increased resistance to synthetic insecticides. The foregoing study aimed to utilize the essential oil of leaves of Citrus sinensis var. Valencia as a cheap, safe, eco-friendly (green), and effective alternative to chemical insecticides. Essential oil samples were collected from fresh and dried leaves across different seasons. They are subjected to hydrodistillation and then GC analysis to be compared. Seventy-seven compounds were detected in all samples where monoterpene hydrocarbons represented the most abundant class of hydrocarbons in fresh leaves (52.6–74.4%) and dried leaves (58.6–66.9%). Sabinene (8.26–29.2%), delta-3-carene (8.23–16.4%), d-limonene (2.50–11.2%), and β-myrcene (2.40–4.93%) were the major monoterpene hydrocarbons in all seasons. Oxygenated monoterpenes comprising β-linalool, citronellal, terpinen-4-ol, β-citral, and α-citral exhibited also appreciable percentages in fresh (21.2–43.4%) and dried leaves (23.4–33.0%). Hierarchical cluster analysis (HCA) and principal component analysis (PCA) effectively segregated all samples into three discriminate clusters where, β-linalool, terpinen-4-ol, β-elemene enantiomer, sabinene, and β-phellandrene constitute the main discriminatory biomarkers. Essential oil of fresh spring leaves (FS) was chosen for nano-formulation adopting the hot emulsification method. Both FS sample and the prepared nano-hexosomal formula were screened against the 3rd instar larvae Culex pipiens L. (common house mosquito). LC₅₀ and LC₉₅ values of FS and oil loaded nano-formula were (48 and 30 552 mg L⁻¹) and (30 and 1830 mg L⁻¹) respectively. α-Citral followed by citronellal showed the best fitting within the binding sites of acetylcholine esterase enzyme utilizing molecular docking. Thus, it can be concluded that Valencia orange leaf as a nano-formulation could serve as an effective and sustainable insecticidal agent.

Mosquitoes and mosquito-borne infectious diseases are a global challenge, especially with increased resistance to synthetic insecticides.     

In vitro evaluation of double and triple combinations of antifungal drugs against Aspergillus fumigatus and Aspergillus terreus

Microdilution broth checkerboard techniques based on the National Committee for Clinical Laboratory Standards methodology were used to study double and triple antifungal combinations against clinical isolates of Aspergillus fumigatus and A. terreus. The influences of the end-point definition (partial or complete inhibition) and the mode of reading (visually or spectrophotometrically) were determined. Interactions between antifungal drugs were also evaluated by agar diffusion tests. Combinations of caspofungin with either amphotericin B or voriconazole were additive for all the isolates, and antagonism was not observed. The interaction between caspofungin and flucytosine was synergistic for 62% of the isolates. In contrast, the interaction between voriconazole and flucytosine was never synergistic and antagonism was noted for 93% of the isolates. The triple combination of caspofungin with flucytosine and amphotericin B was synergistic for all the isolates tested. The triple combination of caspofungin with flucytosine and voriconazole was also mostly synergistic; but complex interactions were obtained for some isolates, with synergy or antagonism depending on the concentrations of caspofungin and voriconazole. Analysis of the influence of the reading technique on the results showed that spectrophotometric reading was a good alternative to the recommended visual reading. The results of these in vitro tests suggest that the activity of flucytosine as part of a double combination with caspofungin and as part of a triple combination with caspofungin and amphotericin B against Aspergillus spp. warrants further investigations. Animal studies are needed to evaluate the in vivo efficacies of these combinations.     

An evaluation of approaches to assessing the quality of nursing care using (predetermined) quality assurance tools

This paper examines the implementation of four of the most common approaches to nursing quality assurance in England, namely Monitor, Qualpacs, nursing audits and a patient satisfaction questionnaire entitled 'What the Patient Thinks'. The primary aim of the study was to look more closely at the context, processes and outcomes of selecting and implementing specific quality assurance tools. Data were analysed at three distinct levels. The findings are presented around the structure of this three-level framework and indicate that the process of implementing a quality assurance tool is more important than the tool itself. It is suggested that a bottom-up approach to implementation, which locates ownership and control of the quality assurance tool with practitioners, is seen to result in more favourable staff responses and positive programme outcomes. The implementation of Qualpacs is used to illustrate some of the tensions that can occur between the inherent principles of the tool and the method of implementation. In studying the factors that might influence the method of implementing a quality assurance tool, a number of organizational and managerial factors are identified.     

Biological and pharmacological approaches to the screening and evaluation of natural products

Many traditional herbal remedies contain active principles, the effects of which can be demonstrated pharmacologically; the biological activity of the whole plant extract is usually related to the constituents that have been identified. Thus, active extracts detected by screening methods are often subjected to more elaborate bioassays to determine their specific pharmacological activity. This paper aims to elucidate the biological, pharmacological and clinical approaches used in the screening of bioactive natural substances. In addition, a scientific process with the goal of finding substances with useful pharmacological activity in plant extracts is rigorously investigated.     

Molecular evaluation of the plasma membrane proton pump from Aspergillus fumigatus

The gene encoding the plasma membrane proton pump (H+ -ATPase) of Aspergillus fumigatus, PMA1, was characterized from A. fumigatus strain NIH 5233 and clinical isolate H11-20. An open reading frame of 3,109 nucleotides with two introns near the N terminus predicts a protein consisting of 989 amino acids with a molecular mass of approximately 108 kDa. The predicted A. fumigatus enzyme is 89 and 51% identical to H+ - ATPases of Aspergillus nidulans and Saccharomyces cerevisiae, respectively. The A. fumigatus PMA1 is a typical member of the P-type ATPase family that contains 10 predicted transmembrane segments and conserved sequence motifs TGES, CSDKTGT, MLTGD, and GDGVN within the catalytic region. The enzyme represents 2% of the total plasma membrane protein, and it is characteristically inhibited by orthovanadate, with a 50% inhibitory concentration of approximately 1.8 microM. H+ -ATPases from Aspergillus spp. contain a highly acidic insertion region of 60 amino acids between transmembrane segments 2 and 3, which was confirmed for the membrane-assembled enzyme with a peptide-derived antibody. An increasing A. fumigatus PMA1 copy number confers enhanced growth in low-pH medium, consistent with its role as a proton pump. These results provide support for the development of the A. fumigatus H+ -ATPase as a potential drug discovery target.     

荧光染色在鼻窦真菌快速诊断中的应用评价

目的评价荧光染色在快速诊断真菌性鼻-鼻窦炎中的应用效果。方法对96例2017年7月至2018年4月首都医科大学附属北京同仁医院临床诊断为真菌性鼻-鼻窦炎的患者术中留取少量真菌团块组织,拉片制成薄层涂片,滴适量荧光染料,加盖玻片,室温放置5 min,在荧光显微镜下从低倍至高倍观察真菌结构。常规进行乳酸酚棉蓝染色及1.79 mol/L KOH湿片处理,以真菌培养结果作为金标准。真菌培养阴性、镜检阳性的样本提取核酸通过ITS序列扩增,进行真菌成分的分子检测。结果荧光染色、乳酸酚棉蓝染色、1.79 mol/L KOH湿片标本一次镜检阳性率分别为95.83%(92/96),93.75%(90/96)及91.67%(88/96),差异无统计学意义;涂片阴性标本再次涂片染色镜检,除1.79 mol/L KOH漏检2例外,荧光及乳酸酚棉蓝染色均可捕捉到真菌结构。培养阳性的65例标本,荧光着色理想,可见清晰的真菌结构,尤其对曲霉菌属与暗色真菌属菌丝区分良好,直接荧光染色拟诊与培养结果一致。有3例经乳酸酚棉蓝染色及1.79 mol/L KOH湿片观察到特征性顶囊结构及分生孢子判断为烟曲霉及链格孢属,结果与培养一致,而荧光染色此类结构着色不清。31例镜检阳性、培养阴性的样本分子检测结果显示,黄曲霉9例、烟曲霉3例、杂色曲霉1例、产黄青霉1例、链格孢霉属2例、尖端赛多孢1例、近平滑假丝酵母菌1例,13例样本为阴性结果。结论荧光染色法可用于鼻窦真菌的快速拟诊,但在观察真菌特征性产孢及产色素结构上,与常规方法相比并无显著优势。     

Hydrogen-bond super-amphiphile based drug delivery system: design,synthesis, and biological evaluation

Drug delivery systems (DDSs) show great application prospects in tumor therapy. So far, physical encapsulation and covalent grafting were the two most common strategies for the construction of DDSs. However, physical encapsulation-based DDSs usually suffered from low drug loading capacity and poor stability, and covalent grafting-based DDSs might reduce the activity of original drug, which greatly limited their clinical application. Therefore, it is of great research value to design a new DDS with high drug loading capacity, robust stability, and original drug activity. Herein, we report a super-amphiphile based drug delivery system (HBS-DDS) through self-assembly induced by hydrogen bonds between amino-substituted N-heterocycles of the 1,3,5-triazines and hydrophilic carmofur (HCFU). The resulting HBS-DDS had a high drug loading capacity (38.1%) and robust stability. In addition, the drug delivery system exhibited pH-triggered size change and release of drugs because of the pH responsiveness of hydrogen bonds. In particular, the anticancer ability test showed that the HBS-DDS could be efficiently ingested by tumor cells, and its half-maximal inhibitory concentration (IC₅₀ = 3.53 μg mL⁻¹) for HeLa cells was close to that of free HCFU (IC₅₀ = 5.54 μg mL⁻¹). The hydrogen bond-based DDS represents a potential drug delivery system in tumor therapy.

Drug delivery systems (DDSs) show great application prospects in tumor therapy.     

Design,synthesis and biological evaluation of N-arylsulfonyl carbazoles as novel anticancer agents

In this work, a set of structurally diverse synthetic carbazoles was screened for their anticancer activities. According to structure–activity relationship studies, carbazoles with an N-substituted sulfonyl group exhibited better anticancer activity. Moreover, compound 8h was discovered to show the most potent anticancer effects on Capan-2 cells by inducing apoptosis and cell cycle arrest in G2/M phase. Finally, the in vivo study demonstrated that 8h prevented the tumor growth in PANC-1 and Capan-2 xenograft models without apparent toxicity.

In this work, a set of structurally diverse synthetic carbazoles was screened for their anticancer activities.