Correlation between bacterial diversity and flavor substances in Longgang soy sauce.

Longgang soy sauce is one of the traditional fermented condiments in China, but its bacterial community succession and its unique flavor development during the fermentation process are not well-investigated. This study evaluated the bacterial diversity, flavor changes, and their correlation during the fermentation of Longgang soy sauce. The results showed that Weissella was the dominant bacterial genus in the fermentation stage of sauce fermented grains. In the first 31 days of the moromi fermentation stage, a variety of bacterial genera such as Weissella, Halomonas, Bacteroides, Pseudomonas, and Tetragenococcus were the dominant bacteria. Our results showed that these bacteria have a significantly positive correlation with phenylethyl alcohol, ethyl acetate, and 3-methyl-1-butanol. As the fermentation progressed, a flora structure with Halomonas as the main bacterial genus was formed. This genus exhibited a significantly positive and positive correlation with 1-octanol, ethyl palmitate, heptanol, and 2-nonanol, which are the unique flavor components of Longgang soy sauce.

Relationship between flavor compounds and changes of microbial community in the solid fermented vinegar.

The relationship between volatile compounds of vinegar and microorganisms is not clear, especially pyrazine, a trace component. In order to reveal their potential relationship, high throughput sequencing, solid-phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) and Spearman's correlation analysis were used. Results showed that Acetobacter and Lactobacillus with opposite abundance trends were the predominant bacteria, and the total abundance of them exceeds 98%, while the predominant fungal genera were Aspergillus and Malassezia, their highest abundances are 75.4% and 81.5%, respectively. In the whole process of microbial community succession, 6 pyrazines were detected including trimethylpyrazine and tetramethylpyrazine, etc., and Spearman's correlation analysis showed that they were positively correlated with the presence of Vibrionimonas, Paraburkholderia, Paucibacter, Komagataeibacter, Acinetobacter, and Slinibacter. In general, this study further revealed more species related to pyrazines, it will be helpful to understand the formation of pyrazines and promote the improvement of vinegar quality.

Isolation of lipopeptide antibiotics from Bacillus siamensis: a potential biocontrol agent for Fusarium graminearum.

Fusarium head blight (FHB), a destructive fungal disease that can cause damage to various crops and reduce yield and quality, is primarily caused by several species of the soil-borne fungal genus Fusarium, which produce mycotoxins that contaminate grains and may cause various severe chronic diseases in humans and livestock. In recent years, Bacillus spp. have been reported to be good producers of antifungal antibiotics against FHB. This study aimed to explore the potential role of a newly identified Bacillus strain, designated as CU-XJ-9, against FHB. This strain, which was isolated from traditional Chinese fermented food, was identified as Bacillus siamensis and confirmed to produce lipopeptide biosurfactants, which according to the analysis by quadrupole time-of-flight tandem mass spectrometry (Q-TOF-MS/MS) may belong to the iturin lipopeptide family. At 100 µg/mL, the isolated antifungal compounds completely inhibited conidial germination. Observation of the effects of the isolated antifungal compounds on the mycelia of F. graminearum by scanning electron microscopy revealed obvious nodes in the middle of the mycelia and destroyed mycelial structures, and these changes became more pronounced with increasing dose. Overall, this study provides important information regarding the ability of Bacillus siamensis to produce lipopeptide biosurfactants, which showed significant antagonistic activity against F. graminearum.

[Clinical effect of acupuncture based on syndrome differentiation in the treatment of chronic insomnia and its influence on cognitive function].

OBJECTIVE: To compare the effect of acupuncture based on syndrome differentiation and estazolam in the treatment of chronic insomnia and its influence on cognitive function. METHODS: A total of 90 patients with chronic insomnia were randomly divided into an acupuncture group and a medication group, 45 cases in each group. The acupuncture group was treated with acupuncture at Sishencong (EX-HN 1) and bilateral Shenmen (HT 7), Sanyinjiao (SP 6) combined with compatibility of acupoints based on syndrome differentiation, once a day for 6 d and then rest for 1 d, for a total of 4 weeks. The medication group was treated with oral estazolam tablets before bedtime, 1 tablet each time, for a total of 4 weeks. Before and after treatment, the scores of Pittsburgh sleep quality index (PSQI), mini-mental state examination (MMSE), Montreal cognitive assessment (MoCA) and auditory verbal memory test (AVMT) of the two groups were compared, and the effects were evaluated. RESULTS: After treatment, the PSQI sub-item scores and total scores of the two groups were lower than those before treatment ( P<0.05 ), and above scores in the acupuncture group were lower than those in the medication group ( P<0.05 ); the scores of MMSE, MoCA and AVMT in the two groups were higher than those before treatment ( P<0.05 ), and the scores in the acupuncture group were higher than those in the medication group ( P<0.05 ). The total effective rate of the acupuncture group was 80.0% (36/45), which was higher than 53.3% (24/45) in the medication group (P<0.05). CONCLUSION: Syndrome differentiation acupuncture can improve the sleep quality and cognitive function of patients with chronic insomnia, and the curative effect is better than that of estazolam.

Safety evaluation of Staphylococcus and its application in soy sauce brewing.

The strain Staphylococcus PT-1 was isolated from soy sauce mash and whole genome sequencing revealed it didn't contain drug resistance genes and virulence genes. Salt tolerance test showed that PT-1 could withstand 20% NaCl. It was inoculated into a pork broth medium. Through volatile component detection, the content of pyrazine in the fermentation broth was 0.83% pyrazine and 1.36% 2, 5-dimethylpyrazine. When the strain was applied to the brewing process of soy sauce, the contents of total nitrogen and amino acid nitrogen in the resulting product were higher than those in the control group; the contents of ammonium salt and NaCl were lower than those in the control group; the total acid content was similar. Solid phase microextraction-gas chromatography-mass spectrometry was used to detect the volatile components of two finished soy sauce products. Compared to soy sauce fermented by typical means, that added with PT-1 contained unique components, such as 2,5-dimethylpyrazine (1.8027%), 2-methylpyrazine (0.0158%), 3-ethyl-2,5-dimethylpyrazine (0.5809%), and 4-ethyl guaiacol (8.7477%). Sensory evaluation showed that the total score of PT-1-fermented soy sauce was higher than that of the control group and was characterized by full and rich flavor. These results provide new insights for improving the quality of soy sauce by using novel strains for fermentation.

Protein sensing and cell discrimination using a sensor array based on nanomaterial-assisted chemiluminescence.

Cross-reactive sensor arrays, known as "chemical noses", offer an alternative to time-consuming analytical methods. Here, we report a sensor array based on nanomaterial-assisted chemiluminescence (CL) for protein sensing and cell discrimination. We have found that the CL efficiencies are improved to varied degrees for a given protein or cell line on catalytic nanomaterials. Distinct CL response patterns as "fingerprints" can be obtained on the array and then identified through linear discriminant analysis (LDA). The sensing of 12 kinds of proteins at three concentrations, as well as 12 types of human cell lines among normal, cancerous, and metastatic, has been performed. Compared with most fluorescent or colorimetric approaches which rely on the strong interaction between analytes and sensing elements, our array offers the advantage of both sensitivity and reversibility.

A simple and practical solvent-free preparation of polymaleimide.

Polymaleimide (PMAI) was synthesized by reacting polymaleic anhydride (PMA) with urea via a solvent-free reaction at 180 °C. The conversion of PMA could reach 95%. This method is simple, practical and environmentally-friendly. The structure of the resulting PMAI was characterized by ¹H-NMR and IR.

RNA interference targeting the platelet-derived growth factor receptor beta subunit ameliorates experimental hepatic fibrosis in rats.

BACKGROUND/AIMS: Platelet-derived growth factor (PDGF) is the strongest stimulator of the proliferation of hepatic stellate cells (HSCs). PDGF receptor beta subunit (PDGFR-beta) is acquired on HSCs proliferation induced by PDGF. In this study, we aim to investigate the effect of PDGFR-beta small interference RNA (siRNA) on experimental hepatic fibrosis. METHODS: We constructed a PDGFR-beta siRNA expression plasmid and investigated its effect on the activation of HSCs. Bromodeoxyuridine incorporation was performed to investigate the effect of PDGFR-beta siRNA on HSCs proliferation. A hydrodynamics-based transfection method was used to deliver PDGFR-beta siRNA to rats with hepatic fibrosis. The distribution of transgenes in the liver was observed by immunofluorescence. The antifibrogenic effect of PDGFR-beta siRNA was investigated pathologically. RESULTS: Platelet-derived growth factor receptor-beta subunit siRNA could significantly downregulate PDGFR-beta expression, suppress HSCs activation, block the mitogen-activated protein kinase signalling pathway and inhibit HSCs proliferation in vitro. PDGFR-beta siRNA expression plasmid could be delivered into activated HSCs by the hydrodynamics-based transfection method, and remarkably improve the liver function of the rat model induced by dimethylnitrosamine and bile duct ligation. Furthermore, the progression of fibrosis in the liver was significantly suppressed by PDGFR-beta siRNA in both animal models. CONCLUSIONS: Platelet-derived growth factor receptor-beta subunit siRNA may be presented as an effective antifibrogenic gene therapeutic method for hepatic fibrosis.

d-Glucosamine production from chitosan hydrolyzation over a glucose-derived solid acid catalyst.

A glucose-based solid acid catalyst (GSA) was synthesized by hydrothermal carbonization and its physicochemical properties were explored by various characterization techniques including IR, TG and SEM. In addition, its catalytic performance towards d-glucosamine formation from the hydrolysis of chitosan was extensively investigated to determine the effects of reaction parameters, such as reaction temperature, time and mass ratio of catalyst and reactants. The experimental results revealed that the yield of targeted product d-glucosamine could reach as high as 98.1% under optimal conditions (temperature: 110 °C; time: 6 h). After six catalytic cycles, no evident deactivation was observed, suggesting the satisfactory stability of the investigated solid acid catalyst. This might provide insight on the development of suitable catalyst systems for d-glucosamine formation to replace homogeneous catalysts.

Amphoteric starch derivatives as reusable flocculant for heavy-metal removal.

A pH-responsive amphoteric starch derivative (PRAS) bearing dual functional groups (amino and carboxyl groups) was prepared through etherification of starch with 2-chloro-4,6-diglycino-[1,3,5]-triazine. PRAS exhibits a reversible pH-response property in aqueous solution. The attractive property of PRAS is that it could be used as an effective flocculant for heavy metal-ion (e.g. Cu(ii) and Zn(ii)) removal from wastewater by changing pH. The transition of hydrophobicity-hydrophilicity would produce shrinkage of the polymer matrix, facilitating the release of heavy-metal ions from the saturated flocculant. As an ideal flocculant PRAS displayed outstanding stability and reproducibility, whose remove rate for Cu(ii) and Zn(ii) remained at 93% and 91% after three flocculation/regeneration cycles.

Effects of ribozyme targeting platelet-derived growth factor receptor beta subunit gene on the proliferation and apoptosis of hepatic stellate cells in vitro.

BACKGROUND: Activation and proliferation of hepatic stellate cells (HSC) is essentially involved in the development and progression of hepatic fibrosis. The most potent growth factor for HSC is platelet-derived growth factor receptor (PDGF) and PDGF receptor beta subunit (PDGFR-beta) is the predominant signal transduction pathway of PDGF which is overexpressed in activated HSC. This study investigated the cleavage activity of hammerhead ribozyme targeting PDGFR-beta mRNA in HSC and the effect on biological characteristics of HSC. METHODS: Expression vector of anti-PDGFR-beta ribozyme was constructed and transfected into rat activated HSC with lipofectamin. The positive cell clones were gained by G418 selection. The expression of PDGFR-beta, alpha-smooth muscle actin, and typeI and type III collagen were detected by using Northern blot, Western blot and immunocytochemical staining, respectively. The cell proliferation was determined with MTT colorimetric assay. The cell apoptosis was analyzed by using flow cytometry, acridine orange fluorescence vital staining and transmission electron microscopy. RESULTS: The expression of PDGFR-beta at mRNA and protein level was markedly reduced in ribozyme-transfected HSC by 49% - 57% (P < 0.05 - 0.01). The proliferation and alpha-smooth muscle actin expression of ribozyme-transfected HSC were significantly decreased (P < 0.05 - 0.01), and the type I and type III collagen synthesis were also reduced (P < 0.01). In addition, the proliferative response of ribozyme-transfected HSC to PDGF BB was significantly inhibited. Otherwise, the apoptotic cells were significantly increased in ribozyme-transfected HSC (P < 0.01), and typical apoptotic cells could be found under transmission electron microscopy. CONCLUSIONS: The anti-PDGFR-beta ribozyme effectively cleaved the target RNA and significantly inhibited its expression, which blocked the signal transduction of PDGF at receptor level, inhibited HSC proliferation and collagen synthesis, and induced HSC apoptosis. These results suggest that inhibiting PDGFR-beta expression of HSC may be a new target for the therapy of liver fibrogenesis, and ribozyme may be a useful tool for inhibiting PDGFR-beta expression.

[Inhibition of proliferation of hepatic stellate cells by taurine is mediated via regulating cell cycle proteins].

OBJECTIVE: To explore the possible mechanism(s) of taurine-inhibiting the proliferation of hepatic stellate cells (HSC), this study investigated the effect of taurine on the HSC cell cycle and its regulatory protein expression. METHODS: Cell proliferation was assessed by MTT assay. Cell cycle was analyzed by flow cytometry. Cell cycle regulatory protein Cyclin D1 and P21waf1 expression were determined by immunocytochemistry and image-analysis system, and real-time quantitative PCR. RESULTS: HSC proliferation was markedly inhibited when HSC were treated with taurine at concentrations of 5, 10, 20, 30, 40 and 50 mmol/L for 48 hours, and the inhibition rates were 6.7%, 14.4%, 23.3%, 32.2%, 36.7% and 45.6% respectively (P < 0.05-0.01). In the flow cytometry analysis, it was found that taurine could block HSC in the G0/G1 phase from entering the S phase, resulting in more cells in the G0/G1 phase and fewer in the S phase. The percentage of the cells in the G0/G1 phase and the S phase at the dosage of 40 mmol/L were 68.2%+/-1.4% and 26.2+/-1.3% respectively, which was significantly different in comparison to the controls (56.2%+/-1.7% and 38.5%+/-0.8% respectively, P < 0.01). HSC expressed cyclin D1 and P21waf1. Taurine inhibited cyclin D1 expression and induced P21waf1 expression. The cyclin D1 protein and mRNA in the HSC treated with 40 mmol/L taurine were significantly reduced compared with the controls [protein (optical density value): 0.13+/-0.02 versus 0.18+/-0.02, P < 0.01; mRNA: 5776.7+/-3345.0 versus 18,400.6+/-1374.8 copies/10(6) GAPDH, P < 0.01]; and the P21waf1 protein and mRNA were markedly increased compared with the controls [protein (optical density value): 0.19+/-0.02 versus 0.14+/-0.01, P < 0.01; mRNA: 44,866.7+/-3910.7 versus 16,933.3+/-960.9 copies/10(6) GAPDH, P less than 0.05]. CONCLUSIONS: Cyclin D1 and P21waf1 were cell cycle regulatory proteins in HSC, and taurine can inhibit the HSC cyclin D1 expression and stimulate P21waf1 expression, facilitate arresting cells in G0/G1 phase, and suppress cell proliferation.

Effects of taurine on proliferation and apoptosis of hepatic stellate cells in vitro.

BACKGROUND: Hepatic fibrosis, a common response to chronic liver injury, is characterized by increased production of extracellular matrix components, whose major part is produced by hepatic stellate cells (HSCs). Taurine is a sulfur containing beta-amino acid rich in human body, and our previous experiments showed that it can inhibit the deposition of the extracellular matrix in the damaged liver. This work was to investigate the effects of taurine on proliferation and apoptosis of HSC and its possible mechanism. METHODS: Cell proliferation was detected by the thiazole blue (MTT) colorimetric assay. Cell apoptosis and cell cycle were assessed via flow cytometry. The morphology of apoptotic cells was observed by phase-contrast fluorescent micrography after orange acridine staining, and the cAMP content was measured by radioimmunoassay. The expression of c-jun and c-fos was determined by the combination of immunocytochemistry and image analysis software. RESULTS: Taurine dose-dependently inhibited the proliferation of HSCs at the concentration of 5-50 mmol/L, resulting in more cells in the G0/G1 phase and fewer in the S phase. Taurine markedly increased the synthesis of cAMP and suppressed the gene expression of c-jun and c-fos (P<0.01) in addition to the inhibition of the proliferative effect of platelet-derived growth factor BB on HSC. However, taurine had no effect on induction of cell apoptosis. CONCLUSIONS: Taurine can significantly inhibit the proliferation of HSC, causing a G0/G1-phase arrest. This effect on HSC proliferation is associated with the enhancement of the synthesis of cAMP and inhibition of the gene expression of c-jun and c-fos. However it can not induce the apoptosis of HSC.

[Effect of lovastatin on proliferation and extracellular matrix secretion of rat hepatic stellate cells in vitro].

OBJECTIVE: To investigate the effect of lovastatin on proliferation and extracellular matrix secretion of hepatic stellate cells in vitro. METHODS: Rat hepatic stellate cells were incubated with different concentration of lovastatin and geranyl geranypyrophosphate. Cell proliferation was assessed by MTT colorimetric assay. Cell cycle was analysed by flow cytometry. Type IV collagen and laminin were determined by ELISA, and c-jun and c-fos expression by immunocytochemistry and computer video text analysis system. RESULTS: Addition of 0.1 to 50 micromol/L lovastatin into culture medium had no toxicity to hepatic stellate cells, but could significantly inhibit hepatic stellate cell proliferation and provoke G0/G1 phase arrest in dose-dependent manner, and could also markedly inhibit the c-jun and c-fos expression and type IV collagen and laminin secretion, which could partly be antagonized by geranyl geranypyrophosphate. CONCLUSIONS: Lovastatin can significantly inhibit hepatic stellate cell proliferation and type IV collagen and laminin secretion, which might be partly related to its inhibitory effect on geranyl geranypyrophosphate formation.

[Effect of ribozyme against platelet-derived growth factor receptor beta subunit mRNA on the biological characters of hepatic stellate cells].

OBJECTIVE: To study the cleavage activity of hammerhead ribozyme targeting at platelet-derived growth factor receptor beta subunit (PDGFR- beta) mRNA in hepatic stellate cells (HSCs) and its effect on the biological characters of HSCs. METHODS: Expression vector of anti-PDGFR- beta ribozyme was constructed and transfected into rat-derived HSC-T6 cells with lipofectin. The positive cell clones were gained by G418 selection. The expression of PDGFR- beta, alpha-smooth muscle actin (alpha-SMA), and type I and type III collagen was detected by means of northern blot, Western blot and immunocytochemical staining respectively. The cell proliferation was determined with MTT colorimetric assay. The cell apoptosis was demonstrated with flow cytometry, acridine orange fluorescence vital staining and transmission electron microscopy. RESULTS: The expression of PDGFR- beta at mRNA and protein level was markedly reduced in ribozyme-transfected HSCs only 43% to 51% of that in control cells (t > or = 3.957, P < 0.05), and alpha-SMA expression level, type I and type III collagen synthesis ability were also reduced (t > or = 6.790, P < 0.01). The proliferation of ribozyme-transfected HSCs was significantly decreased (t > or = 3.858, P < 0.05), and the proliferation response to PDGF BB was markedly inhibited. However the apoptotic rate was significantly increased in ribozyme-transfected HSCs (chi2 > or = 14.157, P < 0.01), and typical apoptotic cells could be found under transmission electron microscopy. CONCLUSIONS: The anti-PDGFR- beta ribozyme can be expressed stably in HSCs, cleave the target RNA effectively, inhibit HSCs proliferation and collagen synthesis, and induce HSC apoptosis. The results suggest that inhibiting PDGFR- beta expression in HSCs may be a new therapy for liver fibrosis.