Pharmacokinetics and metabolite identification of 23-hydroxybetulinic acid in rats by using liquid chromatography-mass spectrometry method.

23-hydroxybetulinic acid (23-HA), a main bioactive component isolated from Pulsatilla chinensis (Bunge) Regel, exhibits various pharmacological activities, such as antimelanoma, antileukemia, anti-colon cancer, and antihepatotoxicity. Although the main active ingredient anemoside B4 (AB4) from this plant has been well studied, research on its active metabolite 23-HA is limited. In the present study, a validated HPLC-QQQ-MS/MS method was established for the quantification of 23-HA in rat plasma. Pharmacokinetics analysis showed that the absorption and elimination of 23-HA in rats were rapid, with an oral bioavailability as 12.9 %. After oral administration with 50 mg/kg 23-HA for SD rats, the plasma, urine, feces, and bile samples were collected and analyzed by UPLC-Q Exactive Plus MS and HPLC-QQQ-MS/MS. Seventeen metabolites of 23-HA were identified, and its major metabolic pathways included oxidation, hydration, sulfation, and glucuronidation. This study highlights the first detailed investigation of 23-HA's pharmacokinetics in rats along with its metabolism in vivo, and will provide robust evidence for further research and clinical application of 23-HA.

Sinoflavonoids NJ and NK, anti-inflammatory prenylated flavonoids from the fruits of Podophyllum hexandrum Royle.

Two new prenylated flavonoids named sinoflavonoids NJ and NK (1-2), along with ten known compounds were isolated from the fruits of Podophyllum hexandrum Royle. The chemical structures were determined through NMR spectroscopic data and MS analysis. Sinoflavonoid NJ (1) with an unusual 5,11-dioxabenzo[b]fluoren-10-one skeleton was firstly reported from Berberidaceae. The isolated flavonoids were tested with LPS-induced RAW 264.7 mouse macrophages model for their anti-inflammatory activity. Sinoflavonoid NJ (1) showed the most potent inhibition on nitric oxide production with IC50 value as 0.06 µM.

Six Undescribed Capnosane-Type Macrocyclic Diterpenoids from South China Sea Soft Coral Sarcophyton crassocaule: Structural Determination and Biological Evaluation.

Six undescribed capnosane-type macrocyclic diterpenes sarcocrassolins A-F (1-6) and one related known analog pavidolide D (7) were isolated from Sarcophyton crassocaule, a soft coral collected off the Nansha Islands, in the South China Sea. Their complete structures, relative configurations and absolute configurations were established through comprehensive spectroscopic analysis, quantum mechanical nuclear magnetic resonance (QM-NMR) and single-crystal X-ray diffraction. Sarcocrassolins D (4) and E (5) showed inhibitory activity against lipopolysaccharide (LPS)-stimulated inflammatory responses in RAW264.7 cells with IC50 values of 76.8 ± 8.0 µM and 93.0 ± 3.8 µM, respectively.

Endophytic Microbes from Medicinal Plants in Fenghuang Mountain as a Source of Antibiotics.

One of the largest concerns with world health today is still antibiotic resistance, which is making it imperative to find efficient alternatives as soon as possible. It has been demonstrated that microbes are reliable sources for the creation of therapeutic antibiotics. This research intends to investigate the endophytic microorganisms from several medicinal plants in Fenghuang Mountain (Jiangsu Province, China) and to discover new antibiotics from their secondary metabolites. A total of 269 endophytic strains were isolated from nine distinct medicinal plants. Taxonomic analysis revealed that there were 20 distinct species among these endophytes, with Streptomyces being the most common genus. Three of the target strains were chosen for scale-up fermentation after preliminary screening of antibacterial activities and the metabolomics investigation using LC-MS. These strains were Penicillium sp. NX-S-6, Streptomyces sp. YHLB-L-2 and Streptomyces sp. ZLBB-S-6. Twenty-three secondary metabolites (1-23), including a new sorbicillin analogue (1), were produced as a result of antibacterial activity-guided isolation. Through spectroscopic analysis using MS and NMR, the structures of yield compounds were clarified. According to antibacterial data, S. aureus or B. subtilis were inhibited to varying degrees by sorrentanone (3), emodic acid (8), GKK1032 B (10), linoleic acid (14), toyocamycin (17) and quinomycin A (21). The most effective antimicrobial agent against S. aureus, B. subtilis, E. coli and A. baumannii was quinomycin A (21). In addition, quinomycin A showed strong antifungal activity against Aspergillus fumigatus, Cryptococcus neoformans, and two clinical isolated strains Aspergillus fumigatus #176 and #339, with MIC as 16, 4, 16 and 16 µg/mL, respectively. This is the first time that bioprospecting of actinobacteria and their secondary metabolites from medicinal plants in Fenghuang Mountain was reported. The finding demonstrates the potential of endophytic microbes in medical plants to produce a variety of natural products. Endophytic microbes will be an important source for new antibiotics.

Dammarane-type triterpenoid saponins isolated from Gynostemma pentaphyllum ameliorate liver fibrosis via agonizing PP2Cα and inhibiting deposition of extracellular matrix.

Gypenosides, structurally analogous to ginsenosides and derived from a sustainable source, are recognized as the principal active compounds found in Gynostemma pentaphyllum, a Chinese medicinal plant used in the treatment of the metabolic syndrome. By bioactive tracking isolation of the plants collected from different regions across China, we obtained four new gypenosides (1-4), together with nine known gypenosides (5-13), from the methanol extract of the plant. The structures of new gypenosides were elucidated by one-dimensional (1D) and two-dimensional (2D) nuclear magnetic resonance (NMR) spectra, complemented by chemical degradation experiments. Through comprehensive evaluation involving COL1A1 promoter assays and PP2Cα activity assays, we established a definitive structure-activity relationship for these dammarane-type triterpenoids, affirming the indispensability of the C-3 saccharide chain and C-17 lactone ring in effectively impeding extracellular matrix (ECM) deposition within hepatic stellate cells. Further in vivo study on the CCl4-induced liver damage mouse model corroborated that compound 5 significantly ameliorated the process of hepatic fibrosis by oral administration. These results underscore the potential of dammarane-type triterpenoids as prospective anti-fibrotic leads and highlight their prevalence as key molecular frameworks in the therapeutic intervention of chronic hepatic disorders.

Gypenosides Synergistically Reduce the Extracellular Matrix of Hepatic Stellate Cells and Ameliorate Hepatic Fibrosis in Mice.

Liver fibrosis resulting from chronic liver damage is becoming one of the major threats to health worldwide. Active saponin constituents isolated from Gynostemma pentaphyllum were found to possess a protective effect in liver diseases. Here, we obtained a naturally abundant gypenoside, XLVI, and evaluated its liver protection activity in both animal and cellular models. The results showed that it ameliorated acute and chronic liver injuries and lightened the process of fibrogenesis in vivo. XLVI can inhibit TGF-ß-induced activation of hepatic stellate cells and ECM deposition in vitro. The underlying mechanism study verified that it upregulated the protein expression of protein phosphatase 2C alpha and strengthened the vitality of the phosphatase together with a PP2Cα agonist gypenoside NPLC0393. These results shed new light on the molecular mechanisms and the potential therapeutic function of the traditional herb Gynostemma pentaphyllum in the treatment of liver fibrosis.

Rapid evaluation of Radix Paeoniae Alba and its processed products by near-infrared spectroscopy combined with multivariate algorithms.

It is well known that the processing method of herbal medicine has a complex impact on the active components and clinical efficacy, which is difficult to measure. As a representative herb medicine with diverse processing methods, Radix Paeoniae Alba (RPA) and its processed products differ greatly in clinical efficacy. However, in some cases, different processed products are confused for use in clinical practice. Therefore, it is necessary to strictly control the quality of RPA and its processed products. Giving that the time-consuming and laborious operation of traditional quality control methods, a comprehensive strategy of near-infrared (NIR) spectroscopy combined with multivariate algorithms was proposed. This strategy has the advantages of being rapid and non-destructive, not only qualitatively distinguishing RPA and various processed products but also enabling quantitative prediction of five bioactive components. Qualitatively, the subspace clustering algorithm successfully differentiated RPA and three processed products, with an accuracy rate of 97.1%; quantitatively, interval combination optimization (ICO), competitive adaptive reweighted sampling (CARS), and competitive adaptive reweighted sampling combined with successive projections algorithm (CARS-SPA) were used to optimize the PLS model, and satisfactory results were obtained in terms of wavelength selection. In conclusion, it is feasible to use NIR spectroscopy to rapidly evaluate the effect of processing methods on the quality of RPA, which provides a meaningful reference for quality control of other herbal medicines with numerous processing methods.

[Rapid identification of raw and sulfur-fumigated Paeoniae Radix Alba based on Heracles NEO ultra-fast gas phase electronic nose].

Since the current identification method for Paeoniae Radix Alba is complex in operation and long time-consuming with high requirements for technicians, the present study employed Heracles NEO ultra-fast gas phase electronic nose(E-nose) technology to identify raw and sulfur-fumigated Paeoniae Radix Alba decoction pieces in order to establish a rapid identification method for sulfur-fumigated Paeoniae Radix Alba. The odors of raw Paeoniae Radix Alba and its sulfur-fumigated products were analyzed by Heracles NEO ultra-fast gas phase E-nose to obtain the odor chromatographic information. The chemometric model was established, and the data were processed by principal component analysis(PCA), discriminant function analysis(DFA), soft independent modeling of class analogy(SIMCA), and partial least squares discriminant analysis(PLS-DA). The differential compounds of raw and sulfur-fumigated samples were qualitatively analyzed based on the Kovats retention index and Arochembase. As revealed by the comparison of gas chromatograms of raw and sulfur-fumigated Paeoniae Radix Alba, the heights of several peaks in the chromatograms before and after sulfur fumigation changed significantly. The peak(No.8) produced by ethylbenzene disappeared completely due to sulfonation reaction in the process of sulfur fumigation, indicating that ethylbenzene may be the key component in the identification of Paeoniae Radix Alba and its sulfur-fumigated products. In PCA, DFA, SIMCA, and PLS-DA models, the two types of samples were separated into two different regions, indicating that the established models can clearly distinguish between raw and sulfur-fumigated Paeoniae Radix Alba. The results showed that Heracles NEO ultra-fast gas phase E-nose technology could realize the rapid identification of raw and sulfur-fumigated Paeoniae Radix Alba, which provides a new method and idea for the rapid identification of sulfur-fumigated Chinese medicine.

Hygromycin A derivatives isolated from Streptomyces sp. PC-22 in the rhizosphere soil of Pulsatilla chinensis.

On the basis of the one strain-many compounds (OSMAC) strategy, two new hygromycin A derivatives (3, 4), together with six known compounds were isolated from a medicinal plant inter rhizospheric Streptomyces in Pulsatilla chinensis. The structures of 3 and 4 were elucidated using NMR and HRESIMS analyses. A plausible biosynthetic pathway for these compounds was discussed. All the compounds were evaluated for their antimicrobial and cytotoxic activities. Compound 5 exhibited potent inhibitory activity against S. aureus and B. subtilis with the MICs of 16 and 8 µg ml-1, while 4 showed weak inhibitory activity against S. aureus.

Terragines F-G produced by endophytic Bacillus sp. SH-1.2-ROOT-18 from Dendrobium officinale.

Two new terragine analogs (1‒2) with special succinimide and aminopentane moieties were isolated from the fermentation broth of Bacillus sp. SH-1.2-ROOT-18, an endophyte previously discovered from the root of Dendrobium officinale. The structures were elucidated base on comprehensive 1 D/2D NMR and MS data analysis. Complete NMR assignments for the first reported naturally occurring metabolite 3 was also provided.[Formula: see text].

Enzymatic Cß-H Functionalization of l-Arg and l-Leu in Nonribosomally Derived Peptidyl Natural Products: A Tale of Two Oxidoreductases.

Muraymycins are peptidyl nucleoside antibiotics that contain two Cß-modified amino acids, (2S,3S)-capreomycidine and (2S,3S)-ß-OH-Leu. The former is also a component of chymostatins, which are aldehyde-containing peptidic protease inhibitors that─like muraymycin─are derived from nonribosomal peptide synthetases (NRPSs). Using feeding experiments and in vitro characterization of 12 recombinant proteins, the biosynthetic mechanism for both nonproteinogenic amino acids is now defined. The formation of (2S,3S)-capreomycidine is shown to involve an FAD-dependent dehydrogenase:cyclase that requires an NRPS-bound pathway intermediate as a substrate. This cryptic dehydrogenation strategy is both temporally and mechanistically distinct in comparison to the biosynthesis of other capreomycidine diastereomers, which has previously been shown to proceed by Cß-hydroxylation of free l-Arg catalyzed by a member of the nonheme Fe2+- and α-ketoglutarate (αKG)-dependent dioxygenase family and (eventually) a dehydration-mediated cyclization process catalyzed by a distinct enzyme(s). Contrary to our initial expectation, the sole nonheme Fe2+- and αKG-dependent dioxygenase candidate Mur15 encoded within the muraymycin gene cluster is instead demonstrated to catalyze specific Cß hydroxylation of the Leu residue to generate (2S,3S)-ß-OH-Leu that is found in most muraymycin congeners. Importantly, and in contrast to known l-Arg-Cß-hydroxylases, the Mur15-catalyzed reaction occurs after the NRPS-mediated assembly of the peptide scaffold. This late-stage functionalization affords the opportunity to exploit Mur15 as a biocatalyst, proof of concept of which is provided.

Cytotoxic Guaianolide Sesquiterpenoids from Ainsliaea fragrans.

Twelve guaianolide-type sesquiterpene oligomers with diverse structures were isolated from the whole plants of Ainsliaea fragrans, including a novel trimer (1) and two new dimers (2, 3). The chemical structures of the new compounds were elucidated through spectroscopic data interpretation and computational calculations. Ainsfragolide (1) is an unusual guaianolide sesquiterpene trimer generated with a novel C-C linkage at C2'-C15″, which may be biosynthesized prospectively through a further Michael addition. Cytotoxicity results showed that ainsfragolide (1) was the most potent compound against five cancer cell lines with IC50 values in the range of 0.4-8.3 µM.

Metabolite identification of iridin in rats by using UHPLC-MS/MS and pharmacokinetic study of its metabolite irigenin.

Iridin, one of the main bioactive components isolated from Belamcanda chinensis (L.) DC, exerts various pharmacological activities, such as anti-inflammation, antioxidant, and antitumor. However, the metabolism and pharmacokinetics of iridin are still unknown. After 100 mg/kg administration of iridin orally, the plasma, urine, and fecal bio-samples from Sprague-Dawley (SD) rats were collected and detected by ultrahigh-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). The pharmacokinetics of the major metabolite irigenin (aglycon of iridin) and a total of thirteen metabolites of iridin were identified, including five metabolites in plasma, ten metabolites in urine, and six metabolites in feces. The most principal metabolic pathway of iridin was glucuronidation after demethylation and was mediated by UDP-glucuronosyltransferases (UGTs) 1A7, 1A8, 1A9 and 1A10. This study highlights the first-time investigation of the metabolism of iridin in vivo, and the pharmacokinetics of irigenin (the major metabolite of iridin) in rats. These results provide robust evidence for further research and clinical application of iridin.

Jatrophacine, a 4,5-seco-rhamnofolane diterpenoid with potent anti-inflammatory activity from Jatropha curcas.

A new diterpenoid named jatrophacine (1), with an unusual 4,5-seco- rhamnofolane skeleton, was isolated from the roots of Jatropha curcas, together with eleven known diterpenoids. The structure of the new compound was elucidated through a detailed analysis of its 1 D- and 2 D-NMR spectra. The X-ray structure of jatrophol (2) is also presented. Anti-inflammatory activity with LPS-induced RAW 264.7 macrophages revealed that compound 1 strongly inhibited the production of nitric oxide (IC50 = 0.53 µM).

Dendrocrepidamine, a novel octahydroindolizine alkaloid from the roots of Dendrobium crepidatum.

A novel octahydroindolizine alkaloid, named dendrocrepidamine (1) with an unusual 18,19,19'-cyclopropanone-dendrocrepine skeleton, was isolated from the ethanol extract of the roots of Dendrobium crepidatum, along with six known compounds (2-7). The structure of 1 was elucidated through HR-ESIMS, NMR spectroscopic data and computational calculations. All compounds were examined for their inhibitory effects on nitric oxide (NO) production induced by lipopolysaccharide (LPS) in RAW264.7 cells with IC50 values in the range of 3.04-54.89 µM. In vivo, crepidatin (6) (80, 40 and 10 mg/kg) showed a significant protective effect against LPS-induced acute lung injury (ALI) in mice.

The crystal structure of AbsH3: A putative flavin adenine dinucleotide-dependent reductase in the abyssomicin biosynthesis pathway.

Natural products and natural product-derived compounds have been widely used for pharmaceuticals for many years, and the search for new natural products that may have interesting activity is ongoing. Abyssomicins are natural product molecules that have antibiotic activity via inhibition of the folate synthesis pathway in microbiota. These compounds also appear to undergo a required [4 + 2] cycloaddition in their biosynthetic pathway. Here we report the structure of an flavin adenine dinucleotide-dependent reductase, AbsH3, from the biosynthetic gene cluster of novel abyssomicins found in Streptomyces sp. LC-6-2.

Huoshanmycins A‒C, New Polyketide Dimers Produced by Endophytic Streptomyces sp. HS-3-L-1 From Dendrobium huoshanense.

Three new polyketide dimers named huoshanmycins A‒C (1-3) were isolated from a plant endophytic Streptomyces sp. HS-3-L-1 in the leaf of Dendrobium huoshanense, which was collected from the Cultivation base in Jiuxianzun Huoshanshihu Co., Ltd. The dimeric structures of huoshanmycins were composed of unusual polyketides SEK43, SEK15, or UWM4, with a unique methylene linkage. Their structures were elucidated through comprehensive 1D-/2D-NMR and HRESIMS spectroscopic data analysis. The cytotoxicity against MV4-11 human leukemia cell by the Cell Counting Kit-8 (CCK8) method was evaluated using isolated compounds with triptolide as positive control (IC50: 1.1 ± 0.4 µM). Huoshanmycins A and B (1, 2) displayed moderate cytotoxicity with IC50 values of 32.9 ± 7.2 and 33.2 ± 6.1 µM, respectively.

The Anti-colitis Effect of Schisandra chinensis Polysaccharide Is Associated With the Regulation of the Composition and Metabolism of Gut Microbiota.

Background: The pathogenesis of inflammatory bowel disease (IBD) is linked to an intricate association of environmental, microbial, and host-related factors. Polysaccharide affects host immunity by regulating the composition and metabolism of gut microbiota is the common mechanism of disease resistance. However, the efficacy and mechanism of Schisandra chinensis polysaccharide (SCP) in the treatment of inflammatory bowel disease have not been studied. Objective: To explore the effect and mechanism of SCP on dextran sodium sulfate (DSS) - induced ulcerative colitis (UC) in mice. Materials/Methods: In this study, we established a mouse model of UC, and used SCP for treatment intervention. The biochemical indexes related to inflammation were determined by ELISA kit, and the therapeutic effect of SCP on UC was clarified. Then, 16S rDNA sequencing was used to study the effect of SCP on the composition and diversity of gut microbiota. At the same time, GC-MS was used to determine the content of short chain fatty acids in intestinal contents. Finally, the relationship among gut microbiota, short chain fatty acids and inflammatory factors was analyzed, and to comprehensively explain the effect and mechanism of SCP on UC. Results: The results showed that SCP could significantly improve the physiological state of UC mice and regulate the level of inflammatory factors to normal levels. Meanwhile, SCP could significantly regulate the imbalance of gut microbiota and increase the content of SCFAs. In addition, the results of the correlation between gut microbiota and SCFAs showed that butyric acid, isobutyric acid and valeric acid had the highest correlation with gut microbiota. Conclusion: In conclusion, this research showed that SCP can inhibit inflammatory bowel disease by regulating the composition and metabolism of gut microbiota, and indicating that SCP may be used as adjuvant therapy for IBD patients.

The triterpenoid sapogenin (2α-OH-Protopanoxadiol) ameliorates metabolic syndrome via the intestinal FXR/GLP-1 axis through gut microbiota remodelling.

Gypenosides, extracts of Gynostemma yixingense, have been traditionally prescribed to improve metabolic syndrome in Asian folk and local traditional medicine hospitals. However, the mechanism of its action remains unclarified. In this work, our results indicated that chronic administration of 2α-OH-protopanoxadiol (GP2), a metabolite of gypenosides in vivo, protected mice from high-fat diet-induced obesity and improved glucose tolerance by improving intestinal L-cell function. Mechanistically, GP2 treatment inhibited the enzymatic activity of bile salt hydrolase and modulated the proportions of the gut microbiota, which led to an increase in the accumulation of tauro-ß-muricholic acid (TßMCA) in the intestine. TßMCA induced GLP-1 production and secretion by reducing the transcriptional activity of nuclear receptor farnesoid X receptor (FXR). Transplantation of GP2-remodelled fecal microbiota into antibiotic-treated mice also increased the intestinal TßMCA content and improved intestinal L-cell function. These findings demonstrate that GP2 ameliorates metabolic syndrome at least partly through the intestinal FXR/GLP-1 axis via gut microbiota remodelling and also suggest that GP2 may serve as a promising oral therapeutic agent for metabolic syndrome.