Epicoccum sorghinum Causing Leaf Spot on Polygonatum cyrtonema in China.

Polygonatum cyrtonema Hua (family Asparagaceae) is a traditional Chinese medicinal plant that is widely cultivated in various parts of China, including Hunan Province. In summer 2022, a leaf spot disease was observed in 10% of the P. cyrtonema plants (Huang jing) in 18 hectares of this crop in the Hongjiang District (27°18'4″N, 110°11'1″E) of Hunan Province. The initial symptoms of the disease were brown spots on young leaves, and adjacent tissues gradually changed from green to yellow. The entire leaf then became yellow, withered, and eventually exhibited a thn and black appearance. In total, 12 diseased plants from four sampling sites (three plants per site) were collected for laboratory analysis to address the concerns of P. cyrtonema growers. Symptomatic leaf samples were selected, and the leaf fragments containing infected parts of the plants were disinfected with 75% ethanol for 1 min, then immersed in 2.5% hypochlorite for 45 s. After disinfection, symptomatic leaf samples were rinsed three times with sterile water, placed on potato saccharose agar containing 50 µg/ml kanamycin and incubated at 25°C for 2 days. Subsequently, 12 fungal isolates were isolated from various leaf samples through hyphal tip transferring. Ten of the 12 fungal isolates had similar morphological features, and one of them (isolate hjh) was used as the representative isolate for the study. With a growth rate of 6.3 mm per day, its white colonies transformed into red concentric rings in five days; they gradually became black after 10 days of growth. The chlamydospores were round (4.0-9.9 × 3.1-9.3 µm, n = 30), whereas the conidia were ovate (8.0-12.1 × 3.2-6.5 µm, n = 30). The morphological features of the isolate hjh were similar to the features of Epicoccum spp. (Aveskamp et al. 2010). The internal transcribed spacer (ITS) region (including the partial ITS1 sequence and the 5.8S and ITS2 complete sequences), ß-tubulin (tub) gene, and large subunit (LSU) rRNA gene, were amplified from the isolate hjh using the primer pairs ITS5/ITS4, Bt2a/Bt2b, and LROR/LR5, respectively (Taguiam et al. 2021). BLASTn analysis showed that the ITS (OR253745), tub (OR253764), and LSU (OR253746) sequences generated from the isolate hjh were 98-99% similar to the sequences of E. sorghinum strains CBS 179.80 and CBS 627.68. Subsequently, the ITS, tub, and LSU sequences were combined using Sequence Matrix software; phylogenetic analysis via Bayesian and maximum likelihood methods (Vaidya et al. 2011; Li et al. 2021) classified the isolate hjh into the E. sorghinum clade. To fulfill Koch's postulates, pathogenicity tests were conducted on healthy (lesion-free and disease-free) 2-year-old P. cyrtonema plants. Three healthy plants were inoculated by spraying whole plant until run-off with a spore suspension of the isolate hjh (1 × 106 conidia/ml); Three other healthy plants were sprayed with sterile water as controls. The inoculated plants were incubated in a growth chamber at 25 ± 2°C with 85% humidity for 28 days(Chen et al. 2021). Leaves from the inoculated plants gradually became brown within 15 days. Finally, the plants died 28 days after inoculation. The control plants showed no symptoms throughout the experimental period. Isolates (isolate hjh1, hjh2 and hjh3) that were reisolated from the inoculated plants exhibited morphologically similar characteristics and molecularly identical to the original isolate hjh. To our knowledge, this is the first report of E. sorghinum causing leaf spot disease on P. cyrtonema. The results of this study may facilitate the production of P. cyrtonema in China.

Oral Supplements of Combined Bacillus licheniformis Zhengchangsheng® and Xylooligosaccharides Improve High-Fat Diet-Induced Obesity and Modulate the Gut Microbiota in Rats.

Gut dysbiosis induced by high-fat diet (HFD) may result in low-grade inflammation leading to diverse inflammatory diseases. The beneficial effects of probiotics and prebiotics on obesity have been reported previously. However, their benefits in promoting human health and the underlying mechanisms still need to be further characterized. This study is aimed at understanding how probiotic Bacillus licheniformis Zhengchangsheng® (BL) and prebiotic xylooligosaccharides (XOS) influence the health of a rat model with HF (60 kcal %) diet-induced obesity. Five groups of male Sprague Dawley (SD) rats were fed a normal fat diet (CON) or an HFD with or without BL and XOS supplementation for 3 weeks. Lipid profiles, inflammatory biomarkers, and microbiota composition were analyzed at the end of the experiment. Rats fed an HFD exhibited increased body weight and disordered lipid metabolism. In contrast, combined BL and XOS supplementation inhibited body weight gain and returned lipid metabolism to normal. Furthermore, BL and XOS administration changed the gut microbiota composition and modulated specific bacteria such as Prevotellaceae, Desulfovibrionaceae, and Ruminococcaceae. In addition, supplements of combined BL and XOS obviously reduced the serum LPS level, which was significantly related to microbial variations. Our findings suggest that modulation of the gut microbiota as a result of probiotic BL and prebiotic XOS supplementation has a positive effect on HFD-induced obesity in rats.

Preparation and in vitro-in vivo evaluation of intestinal retention pellets of Berberine chloride to enhance hypoglycemic and lipid-lowing efficacy.

Berberine chloride (BBR) is a pharmacokinetic profile of drug with poor bioavailability but good therapeutic efficacy, which is closely related to the discovery of BBR intestinal target. The major aim of this paper is to develop BBR intestinal retention type sustained-release pellets and evaluate their in vivo and in vitro behaviors base on the aspect of local action on intestinal tract. Here, wet milling technology is used to improve dissolution and dissolution rate of BBR by decreasing the particle size and increasing the wettability. The pellets are prepared by liquid layer deposition technology, and then the core pellets are coated with Eudragit® L30D-55 and Eudragit® NE30D aqueous dispersion. The prepared pellets show high drug loading capacity, and the drug loading up to 93%. Meanwhile, it possesses significant sustained drug release effect in purified water which is expected to improve the pharmacokinetic behavior of BBR. The pharmacokinetics results demonstrate that the half-life of BBR was increased significantly from 24 h to 36 h and the inter- and intra-subject variability are decreased compared to commercial BBR tablets. The retention test results indicate that the pellet size and Eudragit® NE30D plays an important role in retention time of the pellet, and it is found that the pellets with small particle size and high Eudragit® NE30D coating content can stay longer in the intestine than the pellets with large particle size. All in all, BBR intestinal retention type pellets are prepared successfully in this study, and the pellets show satisfactory in vivo and in vitro behaviors.

Development and application of a UPLC-MS/MS method for simultaneous determination of fenofibric acid and berberine in rat plasma: application to the drug-drug pharmacokinetic interaction study of fenofibrate combined with berberine after oral administration in rats.

With the purpose of carrying out pharmacokinetic interaction studies ofnberberine (BBR) and fenofibrate (FBT), an UPLC-MS/MS method has been developed and validated. The analytes, BBR and fenofibric acid (FBA, metabolite of FBT) and the internal standard, tetrahydropalmatine, were extracted with dichloromethane-diethyl ether (3:2, v/v) and separated on an Agilent Eclipse XDB C18 column using a mobile phase composed of acetonitrile and water. With positive ion electrospray ionization, the analytes were monitored on a triple quadrupole mass spectrometer in multiple reaction monitoring mode. Linear calibration curves were obtained over the concentration ranges of 0.1-100.0 ng/mL for BBR and 10.0-50,000.0 ng/mL for FBA. For BBR and FBA, the intra- and inter-day precisions were <11.5 and 11.9%, respectively. The accuracy was within 11.7% and 11.3%. The mean recoveries of BBR at three concentrations of 0.2, 20.0, 80.0 ng/mL were >85.6%, and those of FBA at three concentrations of 20.0, 2500.0, 40,000.0 ng/mL were >87.9%. Consequently, the proposed method was applied to the pharmacokinetic interaction study of FBT combined with BBR after oral administration in rats and was proved to be sensitive, specific and reliable to analyze BBR and FBA in biological samples simultaneously. Copyright © 2016 John Wiley & Sons, Ltd.

Validated UPLC-MS/MS method for simultaneous determination of simvastatin, simvastatin hydroxy acid and berberine in rat plasma: Application to the drug-drug pharmacokinetic interaction study of simvastatin combined with berberine after oral administration in rats.

A rapid and sensitive liquid chromatography-tandem mass spectrometric (LC-MS/MS) assay method was developed and validated for simultaneous quantification of simvastatin (SV), its metabolite simvastatin hydroxy acid (SVA) and berberine (BBR) in rat plasma. Separation was performed on Poroshell 120 EC-C18 column (4.6×50mm, 2.7µm) using gradient elution by mobile phase containing acetonitrile and 10mM ammonium acetate (pH 4.5). Polarity switch (positive-negative-positive ionization mode) was performed in a total run time of 4.0min. The lower limits of quantification (LLOQ) for SV, SVA and BBR were 0.10, 0.20 and 0.10ng/mL, respectively. The response function was established for concentration range of 0.10-100ng/mL for SV and BBR and 0.20-3000ng/mL for SVA, with a coefficient of correlation of >0.99 for all the compounds. The proposed method was applied to the drug-drug pharmacokinetic interaction study of SV combined with BBR after oral administration in rats.

Pharmacokinetics of single and multiple oral doses of arbidol in healthy Chinese volunteers.

BACKGROUND: Arbidol is licensed in Russia and China for prophylaxis and treatment of influenza A and B. This study was to assess the pharmacokinetics of single and multiple doses of arbidol in healthy Chinese volunteers. METHODS: This was a single-center, open-label, two-phase study conducted in 12 subjects. In single-dose phase, subjects were randomized to receive single doses of 0.2, 0.4 and 0.8 g of arbidol in a crossover design with a 7-day washout period between administration. In the multiple-dose phase, subjects received 0.2 g 3 times a day for 7 days. Serial blood samples were collected at predefined time points. Plasma concentrations were determined with a validated HPLC method. Safety assessments were conducted throughout the study. RESULTS: After administration of single doses of 0.2, 0.4 and 0.8 g, geometric mean estimates for arbidol Cmax were 0.70, 1.24, and 2.16 mg/l and the mean of AUClast were 3.27, 5.81 and 12.72 mg×h/l, respectively. The AUClast and Cmax showed dose proportionality. After administration of multiple doses, the mean of Cmax,ss of arbidol was 0.41 mg/l and the mean accumulation ratio is ~ 1.12. Compared with single-dose phase, arbidol exhibited lower Cmax and prolonged plasma concentration profiles. CONCLUSIONS: In healthy Chinese subjects, single dosing of arbidol resulted in linear plasma pharmacokinetics. Arbidol exhibited little accumulation with repeated administration. Compared with single doses, multiple oral doses showed somewhat different pharmacokinetics and tissue distribution patterns. Sex did not appear to affect the pharmacokinetic properties of arbidol.