Fine-Tuning Electron-Donor Capability in the Basic Anion of Poly(ionic liquid) Frameworks for Revolutionizing Catalytic Synthesis of Ethyl Methyl Carbonate with Both Ultrahigh Catalytic Activity and Selectivity.

Ethyl methyl carbonate (EMC) is a crucial solvent extensively utilized in lithium-ion battery electrolytes; the transesterification of dimethyl carbonate (DMC) with ethanol is a pivotal reaction for EMC production. However, this reaction faces challenges due to the trade-off between catalytic activity and selectivity from the basic catalysts. In this issue, we report an innovative strategy through fine-tuning the electron-donor capability of the basic phenolate anion ([PhO]) in a novel poly(ionic liquid) (PIL) framework, as synthesized via an alkylation reaction between 1,3,5-tris(bromomethyl)benzene, biphenyldiimidazole, and N,N'-carbonyldiimidazole (CDI) to trigger targeted basicity that can directionally catalyze the transesterification of DMC with ethanol, so as to achieve both ultrahigh catalytic activity and selectivity toward EMC. By varying the substituent groups with electron-withdrawing and electron-donating effects on the phenolate anion, the PILs show expected changes in the catalytic performance, following well with the trend of charge density on these substituted phenolate anions. The optimized catalyst [CPIL-CDI][MeOPhO], induced by p-methoxyphenolate anions, allows an extraordinary EMC yield of 72.19% and an EMC selectivity of 91.48% under mild conditions without any process intensifications, suppressing all of the reported catalysts reported to date. Outcomes and approaches shown in this work have the potential to expedite the systematic design of cations and anions within PILs for industrial-scale EMC production through environmentally friendly transesterification processes.

Resin-Immobilized Palladium Acetate and Alcohol Dehydrogenase for Chemoenzymatic Enantioselective Synthesis of Chiral Diarylmethanols.

The enantioselective synthesis of chiral diarylmethanols is highly desirable in synthetic chemistry and the pharmaceutical industry, but it remains challenging, especially in terms of green and sustainable production. Herein, a resin-immobilized palladium acetate catalyst was fabricated with high activity, stability, and reusability in Suzuki cross-coupling reaction of acyl halides with boronic acids, and the coimmobilization of alcohol dehydrogenase and glucose dehydrogenase on resin supports was also conducted for asymmetric bioreduction of diaryl ketones. Experimental results revealed that the physicochemical properties of the resins and the immobilization modes played important roles in affecting their catalytic performances. These two catalysts enabled the construction of a chemoenzymatic cascade for the enantioselective synthesis of a series of chiral diarylmethanols in high yields (83-90%) and enantioselectivities (87-98% ee). In addition, the asymmetric synthesis of the antihistaminic and anticholinergic drugs (S)-neobenodine and (S)-carbinoxamine was also achieved from the chiral diarylmethanol precursors, demonstrating the synthetic utility of the chemoenzymatic cascade.

Ginseng-derived nanoparticles alleviate alcohol-induced liver injury by activating the Nrf2/HO-1 signalling pathway and inhibiting the NF-κB signalling pathway in vitro and in vivo.

BACKGROUND: Previous studies have confirmed the antioxidant and anti-inflammatory effects of active ginseng components that protect against liver injury. However, ginseng-derived nanoparticles (GDNPs), low-immunogenicity nanovesicles derived from ginseng, have not been reported to be hepatoprotective. PURPOSE: In this study, we investigated whether GDNPs could attenuate alcohol-induced liver injury in LO2 cells and mice by modulating oxidative stress and inflammatory pathways, thereby advancing the theoretical basis for the development of novel pharmacological treatments. STUDY DESIGN: Alcohol was used to construct in vitro and in vivo models of alcoholic liver injury. To explore the mechanisms by which GDNPs exert their protective effects against alcoholic liver injury, we examined the expression of oxidative stress-related genes and analysed inflammatory responses in vitro and in vivo. The experimental findings were verified using network pharmacology. METHODS: The composition of the GDNPs was analysed using liquid chromatography-mass spectrometry. GDNPs were extracted and purified using differential ultracentrifugation and sucrose density gradient centrifugation. In vitro models of alcoholic liver injury were established using LO2 cells, whereas C57BL/6 J mice were used as in vivo models. Oxidative stress, inflammation, and liver injury indicators were measured using appropriate kits. Levels of proteins associated with oxidative stress and inflammation were measured via western blot, while nuclear factor erythroid2-related factor 2 (Nrf2) and NF-κB protein expression was tested using immunofluorescence, immunohistochemistry, and flow cytometry. The levels of relevant transcription factors were determined using qPCR. Experimental haematoxylin and eosin staining was used to characterise the liver histological appearance and damage in mice. Network pharmacological analysis of GDNP mRNA sequencing of GDNPs was used to predict drug targets and disease associations using TCMSP. RESULTS: GDNPs primarily included 77 compounds, including organic acids and their derivatives, amino acids and their derivatives, sugars, terpenoids, and flavonoids. GDNPs have features that allow them to be taken up by LO2 cells and promote their proliferation. In vitro data indicated that GDNPs reduced the levels of alcohol-induced reactive oxygen species by activating the Nrf2/HO-1 signalling pathway, whilst inhibiting the NF-κB pathway and thereby reducing NO, tumour necrosis factor-α, and interleukin-1ß levels to alleviate inflammation. An in vivo model showed that GDNPs improved the liver parameters and pathology in mice with alcoholic liver injury. GDNPs activate the Nrf2/HO-1/Keap1 signalling pathway in a p62-dependent manner to exert antioxidant effects. Furthermore, the TLR4/NF-κB signalling pathway was involved in the in vivo anti-inflammatory effect. Network pharmacology also confirmed that the effects of GDNPs on liver disease were associated with oxidative stress and inflammation-related targets and pathways. CONCLUSION: This study showed for the first time that GDNPs can alleviate alcohol-induced liver damage by activating the Nrf2/HO1 signalling pathway and blocking the NF-κB signalling pathway, thus lowering oxidative stress and inflammatory responses. Hereby, we present the Nrf2/HO1 and NF-κB signalling pathways as potential targets and GDNPs as a novel therapeutic approach for the management of alcohol-induced liver damage.

Ginseng-derived nanoparticles alleviate inflammatory bowel disease via the TLR4/MAPK and p62/Nrf2/Keap1 pathways.

Inflammatory bowel disease (IBD) is closely linked to the homeostasis of the intestinal environment, and exosomes can be used to treat IBD due to their high biocompatibility and ability to be effectively absorbed by the intestinal tract. However, Ginseng-derived nanoparticles (GDNPs) have not been studied in this context and their mechanism of action remains unclear. Here, we investigated GDNPs ability to mediate intercellular communication in a complex inflammatory microenvironment in order to treat IBD. We found that GDNPs scavenge reactive oxygen species from immune cells and intestinal epithelial cells, inhibit the expression of pro-inflammatory factors, promote the proliferation and differentiation of intestinal stem cells, as well as enhancing the diversity of the intestinal flora. GDNPs significantly stabilise the intestinal barrier thereby promoting tissue repair. Overall, we proved that GDNPs can ameliorate inflammation and oxidative stress in vivo and in vitro, acting on the TLR4/MAPK and p62/Keap1/Nrf2 pathways, and exerting an anti-inflammatory and antioxidant effect. GDNPs mitigated IBD in mice by reducing inflammatory factors and improving the intestinal environment. This study offers new evidence of the potential therapeutic effects of GDNPs in the context of IBD, providing the conceptual ground for an alternative therapeutic strategy.

Timing of endoscopy for acute variceal bleeding in patients with cirrhosis (CHESS1905): A nationwide cohort study.

BACKGROUND: Endoscopy plays an important role in the management of acute variceal bleeding (AVB) in patients with cirrhosis. This study aimed at determining the optimal endoscopy timing for cirrhotic AVB. METHODS: Patients with cirrhosis with AVB across 34 university hospitals in 30 cities from February 2013 to May 2020 who underwent endoscopy within 24 hours were included in this study. Patients were divided into an urgent endoscopy group (endoscopy <6 h after admission) and an early endoscopy group (endoscopy 6-24 h after admission). Multivariable analysis was performed to identify risk factors for treatment failure. Primary outcome was the incidence of 5-day treatment failure. Secondary outcomes included in-hospital mortality, need for intensive care unit, and length of hospital stay. A propensity score matching analysis was performed. In addition, we performed an analysis, in which we compared the 5-day treatment failure incidence and the in-hospital mortality among patients with endoscopy performed at <12 hours and 12-24 hours. RESULTS: A total of 3319 patients were enrolled: 2383 in the urgent endoscopy group and 936 in the early endoscopy group. After propensity score matching, on multivariable analysis, Child-Pugh class was identified as an independent risk factor for 5-day treatment failure (HR, 1.61; 95% CI: 1.09-2.37). The incidence of 5-day treatment failure was 3.0% in the urgent endoscopy group and 2.9% in the early group ( p = 0.90). The in-hospital mortality was 1.9% in the urgent endoscopy group and 1.2% in the early endoscopy group ( p = 0.26). The incidence of need for intensive care unit was 18.2% in the urgent endoscopy group and 21.4% in the early endoscopy group ( p = 0.11). The mean length of hospital stay was 17.9 days in the urgent endoscopy group and 12.9 days in the early endoscopy group ( p < 0.05). The incidence of 5-day treatment failure in the <12-hour group was 2.3% and 2.2% in the 12-24 hours group ( p = 0.85). The in-hospital mortality was 2.2% in the <12-hour group and 0.5% in the 12-24 hours group ( p < 0.05). CONCLUSIONS: The data suggest that performance of endoscopy within 6-12 or within 24 hours of presentation among patients with cirrhosis with AVB led to similar treatment failure outcomes.

Ginseng-derived nanoparticles induce skin cell proliferation and promote wound healing.

Background: Past studies suggested that ginseng extracts and ginseng-derived molecules exerted significant regulatory effects on skin. However, no reports have described the effects of ginseng-derived nanoparticles (GDNPs) on skin cell proliferation and wound healing. In this study, we investigated whether GDNPs regulate the proliferation of skin cells and promote wound healing in a mouse model. Methods: GDNPs were separated and purified via differential centrifugation and sucrose/D2O gradient ultracentrifugation. GDNP uptake, cell proliferation and cell cycle progression were measured by confocal microscopy, CCK-8 assay and flow cytometry, respectively. Cell migration and angiogenic effects were assessed by the wound scratch assay and tube formation assay, respectively. ELISA was used to detect extracellular matrix secretion. The relevant signaling pathway was confirmed by western blotting. The effects of GDNPs on skin wound healing were assessed by wound observation, HE staining, and western blotting. Results: GDNPs possessed the essential features of exosomes, and they were accumulated by skin cells. Treatment with GDNPs notably enhanced the proliferation of HaCaT, BJ and HUVECs. GDNPs also enhanced the migration in HaCaT cells and HUVECs and angiogenesis in HUVECs. GDNPs increased the secretion of MMP-1, fibronectin-1, elastin-1, and COL1A1 in all three cell lines. GDNPs regulated cell proliferation through the ERK and AKT/ mTOR pathways. Furthermore, GDNPs facilitated skin wound healing and decreased inflammation in a mouse skin wound model. Conclusion: GDNPs can promote skin wound healing through the ERK and AKT/mTOR pathways. GDNPs thus represent an alternative treatment for chronic skin wounds.

Benzoyl-xanthone derivative induces apoptosis in MCF-7 cells by binding TRAF6.

TNF receptor-associated factor 6 (TRAF6) has been reported to be associated with the development of cancer. Nevertheless, the exact role of TRAF6 in cancer remains unclear. The purpose of the present study was to explore the mechanism of 2-benzoyl-3-hydroxy-4-methyl-9H-xanthen-9-one leading to the inhibition of the activation of AKT and TGF-ß-activated kinase 1 (TAK1), and to the apoptosis of MCF-7 cells. Using a computational docking program and examination of AKT and TAK1 level changes, a new small molecule was identified, 2-benzoyl-3-hydroxy-4-methyl-9H-xanthen-9-one, which competitively bound to TRAF6. Next, the effect of this new compound on MCF-7 cells' biological behavior was studied in vitro. MTT assays were used to investigate cell viability; flow cytometry and invasion assays were performed to detect early apoptosis and invasion in MCF-7 cells, respectively. Immunoprecipitation, western blotting and caspase-3/9 activity assays were carried out to explore changes in protein expression. Briefly, the present data indicated that 2-benzoyl-3-hydroxy-4-methyl-9H-xanthen-9-one could suppress proliferation, induce early apoptosis and inhibit invasion in MCF-7 cells by suppressing the expression of Bcl-2 and promoting the expression of Bax, caspase-9, and caspase-3. These findings indicated that 2-benzoyl-3-hydroxy-4-methyl-9H-xanthen-9-one could induce apoptosis by inhibiting the activation of AKT and TAK1, and affecting the Bcl-2/Bax-caspase-9-caspase-3 pathway by competitively binding with TRAF6.

Ginseng root extract attenuates inflammation by inhibiting the MAPK/NF-κB signaling pathway and activating autophagy and p62-Nrf2-Keap1 signaling in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Panax ginseng C.A. Meyer is a type of herbal plant that has frequently been used in many Asian countries to treat a variety of diseases. Ginseng is considered to exhibit anti-inflammatory and anti-oxidative pharmacological effects. However, the specific mechanism is still not entirely understood. AIM OF THE STUDY: In this study, we investigated if ginseng extract could attenuate inflammation and oxidative stress in RAW264.7 cells and in dextran sulfate sodium (DSS)-induced colitis mouse model. MATERIALS AND METHODS: RAW264.7 cells and LPS were used to develop inflammatory and oxidative cell models. C57/6J male mice and DSS were used to construct the animal models. O2-, mitochondria number, and mitochondrial membrane potential were analyzed using fluorescent probes and fluorescence microscopy. Reactive oxygen species and nitric oxide generation were detected with probes and microplate readers. The secreted amounts of inflammatory cytokines were measured by enzyme-linked immunosorbent assay kits. Protein expression levels in the cytoplasm and nucleus were measured by western blotting analyses. Quantitative real-time PCR (qRT-PCR) was used to determine the changes in mRNA levels. Autophagosome accumulation was analyzed by transmission electron microscopy. A p62-specific siRNA was used to evaluate the effect of p62 on the anti-oxidative function of ginseng root extract (GRE). Asperuloside and SP600125 were used to confirm the involvement of the MAPK/NF-κB signaling pathway. RESULTS: We performed a systematic analysis of the anti-inflammatory, anti-oxidative, and autophagy regulatory mechanisms of GRE in LPS-treated RAW264.7 cells. GRE considerably reduced the levels of nitric oxide, TNF-α, and IL-6 secreted by LPS-treated cells. GRE treatments dose-dependently upregulated IL-10 mRNA levels and decreased IL-6 and IL-1ß mRNA levels in LPS-treated cells. Similar to the NF-κB and JNK inhibitors, GRE treatment significantly inhibited NF-κB activity and phosphorylation of MAPKs (JNK, ERK-1/2, and p38). Additionally, GRE treatment remarkably decreased LPS-triggered reactive oxygen species production and mitochondrial dysfunction by motivating Nrf2 nuclear translocation by enhancing phosphorylated p62. Knockdown of p62 resulted in the loss of GRE anti-oxidative ability. Autophagy was strongly induced by GRE via the Akt-mTOR signaling pathway, relieving excessive oxidation, mitochondrial dysfunction, and inflammation, while enhancing Beclin-1, LC3 II, and Atg7 protein expression. Furthermore, GRE alleviated the degree of injury, inflammatory cytokine production, and regulated the relative signaling pathway in DSS-induced colitis. CONCLUSIONS: GRE can exert both anti-inflammatory and anti-oxidative functions by targeting the MAPK/NF-κB and p62-Nrf2-Keap1 pathways, as well as autophagy, in vitro and vivo.

Detachable string magnetically controlled capsule endoscopy for detecting high-risk varices in compensated advanced chronic liver disease (CHESS1801): A prospective multicenter study.

BACKGROUND: Gastroesophageal varices is a serious complication of compensated advanced chronic liver disease (cACLD). Primary prophylaxis to reduce the risk of variceal hemorrhage is recommended if high-risk varices (HRV) are detected. We performed this study to compare the accuracy, patients' satisfaction and safety of detection of HRV by detachable string magnetically controlled capsule endoscopy (DS-MCCE) with esophagogastroduodenoscopy (EGD) as the reference. METHODS: We prospectively recruited participants with cACLD from 12 university hospitals (11 in China and one in the United Kingdom) between November 2018 and December 2019 (ClinicalTrials.gov, NCT03749954). All participants underwent DS-MCCE, followed by EGD within a week in a blinded fashion. Following endoscopy, and on the same day, participants were asked to fill in a satisfaction questionnaire regarding their experience. FINDINGS: A total of 105 eligible participants were enrolled. With EGD as the reference standard, the concordance index, sensitivity, specificity, positive predictive value, negative predictive value, positive likelihood ratio, and negative likelihood ratio of DS-MCCE in diagnosis of HRV were 0•90 (95% confidence interval [CI]: 0•83-0•95), 92% (95% CI: 78-98%), 88% (95% CI: 78-95%), 80% (95% CI: 70-92%), 95% (95% CI: 90-100%), 7•91 (95% CI: 4•10-15•30), and 0•09 (95% CI: 0•03-0•30), respectively. The kappa score of 0•78 (95% CI: 0•65-0•90) suggested substantial agreement between DS-MCCE and EGD. Moreover, in participants undergoing EGD without sedation, the satisfaction of DS-MCCE was significantly better than that of EGD (p < 0•0001, d = 1•15 [95%CI: 0•88-1•42]). All participants confirmed the excretion of the capsule, and no adverse events occurred. INTERPRETATION: DS-MCCE is an accurate alternative to EGD for detecting HRV in cACLD, which is safe and associated with better satisfaction. FUNDING: A full list of funding can be found in the Funding Support section.

Compatibility and Efficacy of the Parasitoid Eretmocerus hayati and the Entomopathogenic Fungus Cordyceps javanica for Biological Control of Whitefly Bemisia tabaci.

Biological control is an effective method for whitefly management compared to the potential problems caused by chemical control, including environmental pollution and the development of resistance. Combined use of insect parasitoids and entomopathogenic fungi has shown high efficiency in Bemisia tabaci control. Here, we assessed the impacts of an entomopathogenic fungus, Cordyceps javanica, on the parasitism rate of a dominant whitefly parasitoid, Eretmocerus hayati, and for the first time also compared their separate and combined potential in the suppression of B. tabaci under semi-field conditions. Six conidial concentrations of C. javanica (1 × 103, 1 × 104, 1 × 105, 1 × 106, 1 × 107 and 1 × 108 conidia/mL) were used to assess its pathogenicity to the pupae and adults of E. hayati. Results showed that the mortality of E. hayati increased with higher concentrations of C. javanica, but these higher concentrations of fungus had low pathogenicity to both the E. hayati pupae (2.00-28.00% mortality) and adults (2.67-34.00% mortality) relative to their pathogenicity to B. tabaci nymphs (33.33-92.68%). Bioassay results indicated that C. javanica was harmless (LC50 = 3.91 × 1010) and slightly harmful (LC50 = 5.56 × 109) to the pupae and adults of E. hayati respectively on the basis of IOBC criteria, and that E. hayati could parasitize all nymphal instars of B. tabaci that were pretreated with C. javanica, with its rate of parasitism being highest on second-instar nymphs (62.03%). Interestingly, the parasitoids from second and third-instar B. tabaci nymphs infected with C. javanica had progeny with increased longevity and developmental periods. Moreover, experimental data from 15 day semi-field studies indicate that combined application of C. javanica and E. hayati suppresses B. tabaci with higher efficiency than individual applications of both agents. Therefore, combined applications of C. javanica (1 × 108 conidia/mL) and E. hayati is a more effective and compatible biological control strategy for management of B. tabaci than using either of them individually.

Polymer Constructed Through the Formation of Carbon-Carbon Triple Bonds: Reductive Coupling Polymerization of Bis(benzylic gem-tribromide)s.

Ethynylene-bridged polymers are currently synthesized by alkyne metathesis polymerization or Pd-catalyzed coupling between diethynylarene and dihaloarene. We report the formation of C≡C linkages in reductive coupling polymerization of bis(benzylic gem-tribromide)s promoted by Cu/polyamine under mild conditions. The polymer backbone was constructed through cascade formation of (Br)C═C(Br) bonds and C≡C bonds. This protocol provides a new method for synthesis of ethynylene-bridged polymers using monomers without alkynyl groups.

A strategy for fusion expression and preparation of functional glucagon-like peptide-1 (GLP-1) analogue by introducing an enterokinase cleavage site.

KGLP-1, a 31-amino acid glucagon-like peptide-1 (GLP-1) analogue, has a great therapeutic potential for anti-diabetes. In this work, a strategy for expression and purification of functional KGLP-1 peptide has been established. KGLP-1 cDNA was fused with glutathione S-transferase (GST), with an enterokinase cleavage site in the fusion junction. The recombinant fusion protein GST-KGLP-1 was affinity purified via the GST-tag, and then digested with enterokinase. The resulting GST part as well as the enzymes were eliminated by ultra-filtration followed by size exclusion chromatograph. The yield of purified KGLP-1 was approximately 12.1 mg/L, with purity of 96.18 %. The recombinant KGLP-1 was shown to have similar bioactivity as native GLP-1 when evaluated in a Chinese hamster ovary cell line expressing a GLP-1 receptor-egfp reporter gene.

Biomechanical evaluation of porous bioactive ceramics after implantation: micro CT-based three-dimensional finite element analysis.

Hydroxyapatite ceramics have been widely investigated for bone regeneration due to their high biocompatibility. However, few studies focus on their mechanical characteristics after implantation. In this study, the finite element (FE) method was used to evaluate the mechanical properties of a fully interconnected porous hydroxyapatite (IPHA) over time of implantation. Based on the micro-CT images obtained from the experiments dealing with IPHA implanted into rabbit femoral condyles, three-dimensional FE models of IPHA (1, 5, 12, 24, and 48 weeks after implantation) were developed. FE analysis indicated that the elastic modulus gradually increased from 1 week and reached the peak value at 24 weeks, and then it kept at high level until 48 weeks postoperatively. In addition, as a local biomechanical response, strain energy density became to distribute evenly over time after the implantation. Results confirmed that the mechanical properties of IPHA are strongly correlated to bone ingrowth. The efficiency of the proposed numerical approach was validated in combination with experimental studies, and the feasibility of applying this approach to study such implanted porous bioceramics was proved.

Structural insight into substrate specificity of human intestinal maltase-glucoamylase.

Human maltase-glucoamylase (MGAM) hydrolyzes linear alpha-1,4-linked oligosaccharide substrates, playing a crucial role in the production of glucose in the human lumen and acting as an efficient drug target for type 2 diabetes and obesity. The amino- and carboxyl-terminal portions of MGAM (MGAM-N and MGAM-C) carry out the same catalytic reaction but have different substrate specificities. In this study, we report crystal structures of MGAM-C alone at a resolution of 3.1 Å, and in complex with its inhibitor acarbose at a resolution of 2.9 Å. Structural studies, combined with biochemical analysis, revealed that a segment of 21 amino acids in the active site of MGAM-C forms additional sugar subsites (+ 2 and + 3 subsites), accounting for the preference for longer substrates of MAGM-C compared with that of MGAM-N. Moreover, we discovered that a single mutation of Trp1251 to tyrosine in MGAM-C imparts a novel catalytic ability to digest branched alpha-1,6-linked oligosaccharides. These results provide important information for understanding the substrate specificity of alpha-glucosidases during the process of terminal starch digestion, and for designing more efficient drugs to control type 2 diabetes or obesity.

Study of the inhibition of two human maltase-glucoamylases catalytic domains by different α-glucosidase inhibitors.

In humans, both the N-terminal catalytic domain (NtMGAM) and the C-terminal catalytic domain (CtMGAM) of small intestinal maltase glucoamylase (MGAM) are α-glycosidases that catalyze the hydrolysis of α-(1→4) glycosidic linkages in the process of starch digestion, and are considered to be the main therapeutic targets for type 2 diabetes. In this work, recombinant human CtMGAM has been cloned for the first time, and this, combined with the expression of NtMGAM in Pichia pastoris, made it possible for us to study the catalytic mechanism of MGAM in a well-defined system. The enzymatic kinetic assays of the two catalytic domains suggest that CtMGAM has the higher affinity for longer maltose oligosaccharides. Kinetic studies of commercially-available drugs such as 1-deoxynojirimycin (DNJ), miglitol, voglibose, and acarbose along with a series of acarviosine-containing oligosaccharides we isolated from Streptomyces coelicoflavus against NtMGAM, CtMGAM, and human pancreatic α-amylase (HPA) provide us an overall profile of the inhibitory ability of these inhibitors. Of all the inhibitors used in this paper, DNJ was the most effective inhibitor against MGAM; the K(i) values for the two catalytic domains were 1.41 and 2.04 µM for NtMGAM and CtMGAM, respectively. Acarviostatins 2-03 and 3-03 were the best inhibitors against HPA with relatively high inhibitory activity against CtMGAM. The acarviostatins 2-03 and 3-03 inhibition constants, K(i), for HPA were 15 and 14.3 nM, and those for CtMGAM were 6.02 and 6.08 µM, respectively. These results suggest that NtMGAM and CtMGAM differ in their substrate specificities and inhibitor tolerance despite their structural relationship.

[High-throughput screening of human pancreatic alpha-amylase inhibitors].

OBJECTIVE: Targeting the important enzyme in human glucose metabolic pathway, we established a high throughput screening model for human pancreatic alpha-amylase inhibitors. METHODS: Pichia pastoris expression system was used to clone and express the human pancreatic alpha-amylase; we established the alpha-amylase inhibitor screening model using the catalytic properties of enzyme; this model was applied in screening of actinomycete' metabolites; the taxonomic status of positive strains were analyzed by constructing 16S rRNA phylogenetic tree. RESULTS: We cloned and expressed the intact gene of human pancreatic alpha-amylase successfully; the high-throughput screening model of alpha-amylase inhibitors was established; nearly 2000 actinomycete' metabolites were screened, 14 alpha-amylase inhibitor producing strains were obtained finally, and showed taxonomically rich diversity. CONCLUSION: The alpha-amylase inhibitor high-throughput screening model had high practical value for developing new hypoglycemic drugs.

Structures of human pancreatic α-amylase in complex with acarviostatins: Implications for drug design against type II diabetes.

Human pancreatic α-amylase (HPA) catalyzes the hydrolysis of α-d-(1,4) glycosidic linkages in starch and is one of the major therapeutic targets for type II diabetes. Several acarviostatins isolated from Streptomyces coelicoflavus var. nankaiensis previously showed more potent inhibition of HPA than acarbose, which has been successfully used in clinical therapy. However, the molecular mechanisms by which acarviostatins inhibit HPA remains elusive. Here we determined crystal structures of HPA in complexes with a series of acarviostatin inhibitors (I03, II03, III03, and IV03). Structural analyses showed that acarviostatin I03 undergoes a series of hydrolysis and condensation reactions in the HPA active site, similar to acarbose, while acarviostatins II03, III03, and IV03 likely undergo only hydrolysis reactions. On the basis of structural analysis combined with kinetic assays, we demonstrate that the final modified product with seven sugar rings is best suited for occupying the full active site and shows the most efficient inhibition of HPA. Our high resolution structures reported here identify first time an interaction between an inhibitor and subsite-4 of the HPA active site, which we show makes a significant contribution to the inhibitory effect. Our results provide important information for the design of new drugs for the treatment of type II diabetes or obesity.

Influence of aeration on CH4, N2O and NH3 emissions during aerobic composting of a chicken manure and high C/N waste mixture.

Co-composting of chicken manure, straw and dry grasses was investigated in a forced aeration system to estimate the effect of aeration rates on NH(3), CH(4) and N(2)O emissions and compost quality. Continuous measurements of gas emissions were carried out and detailed gas emission patterns were obtained using an intermittent-aeration of 30 min on/30 min off at rates of 0.01 (A1), 0.1 (A2) and 0.2 (A3) m(3)min(-1)m(-3). Concentrations of CH(4) and N(2)O at the low aeration rate (A1) were significantly greater than those at the other two rates, but there was no significant difference between the A2 and A3 treatments. CH(4) and N(2)O emissions for this mixture could be controlled when the composting process was aerobic and ammonia emissions were reduced at a lower aeration rate. Comparison of CH(4), N(2)O, NH(3) emissions and compost quality showed that the aeration rate of the A2 treatment was superior to the other two aeration rates.

[Establishment and application of a high-throughput model for screening alpha-glucosidase inhibitors].

OBJECTIVE: Targeted at the important enzyme in human glucose metabolic pathway, the purpose of this paper is to establish alpha-glucosidase inhibitors high throughput screening model. METHODS: Pichia pastoris expression system was used to clone and express the human alpha-maltase glucosidase. Using the catalytic properties of enzyme to establish alpha-glucosidase inhibitor screening model. This model was applied in screening of actinomycete metabolites library. The taxonomic status of positive strains were analyzed by constructing 16S rRNA phylogenetic tree. RESULTS: The N-terminal catalytic domain of human alpha-maltase glucosidase was successfully cloned and expressed for the first time. The high-throughput screening model of alpha-glucosidase inhibitors was established. A natural product library containing metabolites from nearly 2000 actinomycetes was screened, 20 alpha-maltase glucosidase inhibitor producing strains were obtained finally, of which, 19 strains initially identified as Streptomyces, and showed taxonomically rich diversity. CONCLUSION: The alpha-glucosidase inhibitor high-throughput screening model has high practical value, this work laid the foundation for developing new hypoglycemic drugs.

Impact of struvite crystallization on nitrogen losses during composting of pig manure and cornstalk.

An absorbent mixture of magnesium hydroxide (Mg(OH)(2)) and phosphoric acid (H(3)PO(4)) was added to compost mixtures of pig manure with cornstalk in different molar ratios (T1, 1:1; T2, 1:2; T3, 1:3) in order to examine its effect on controlling ammonia losses during composting. Based on the principle of struvite precipitation, and with an unamended trial as control (CK), an in-vessel composting experiment was conducted in fermenters (60L with forced aeration) in which the absorbent mixture was added with proportions of 3.8%, 7.3% and 8.9% of dry weight for T1, T2 and T3, respectively. The results showed that the total nitrogen loss was reduced from 35% to 12%, 5% and 1% of initial N mass, respectively. In the final compost, the total nitrogen content in T1, T2 and T3 was improved by 10, 14, 12gkg(-1), and NH(4)(+)-N in T1, T2 and T3 was improved by 8, 9, and 10gkg(-1), respectively, compared with the unamended trial. The results of the germination index test showed that the maturity of treatment T2 was best among the four treatments in the final compost, followed by T1, CK and T3. The results of X-ray diffraction (XRD) confirmed the formation of magnesium ammonium phosphate hexahydrate (MgNH(4)PO(4).6H(2)O:MAP) in the T1, T2 and T3 compost. Based on these results, the adsorbent mixture of Mg(OH)(2)+H(3)PO(4) could control nitrogen loss effectively during composting via struvite crystallization. However, an excess of phosphoric acid (1:3) had a negative influence on composting properties. The pH value decreased which led to reduced microorganism activity, and which finally resulted in reduced biodegradation of the organic matter.