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The leaves of Lycium barbarum (LLB) have been utilized as crude drugs and functional tea for human health in China and Southeast Asia for thousands of years. To control its quality, a rapid and sensitive ultra-high-performance liquid chromatography coupled with triple quadrupole tandem mass spectrometry method was established and validated for the first time for simultaneous determination of 10 phenolic acids and flavonoids (including neochlorogenic acid, protocatechuic aldehyde, p-hydroxybenzoic acid, chlorogenic acid, cryptochlorogenic acid, caffeic acid, p-coumaric acid, ferulic acid, rutin and kaempferol-3-O-rutinoside) in LLB. The separation was performed on an Acquity UPLC C 18 chromatographic column (100 × 2.1 mm internal diameter, 1.7 μm particle size) with 0.1% formic acid in water (A)-acetonitrile (B) as the mobile phase under gradient elution. Multiple reaction monitoring mode was adopted to simultaneously monitor the target components. The developed method was fully validated in terms of linearity (r 2  ≥ 0.9860), precision (RSD ≤ 6.58%), repeatability (RSD ≤ 6.60%), stability (RSD ≤ 6.17%), recovery (95.56-108.06%, RSD ≤ 4.64%) and limit of detection (0.021-0.664 ng/mL) and limit of quantitation (0.069-2.210 ng/mL), and then successfully applied to evaluate the quality of 64 batches of LLB collected from 41 producing areas in four different provinces of China. The results showed that the LLB, especially collected from Inner Mongolia regions, were rich in the phenolic acids and flavonoids. Rutin, kaempferol-3-O-rutinoside and chlorogenic acid are the predominant compounds contained in LLB. The above findings will provide helpful information for the effective utilization of LLB., (© 2019 John Wiley & Sons, Ltd.)

Ethnopharmacological Relevance: The flower of Abelmoschus manihot (Linn.) Medicus (A. manihot), as a traditional Chinese Herbal medicine, was used widely in China with efficacy of inducing diuresis for treating strangurtia, and subdhing swelling and detoxicating. It has been reported that Huangkui capsule, prepared by the extract of the flower of A. manihot, can reduce the content of urinary protein, serum creatinine and serum urea nitrogen in nephropathy rats and processes renoprotective activity, while the action mechanism need to illuminate deeply., Aims of the Study: In this study, we investigated the protection effect of Huangkui capsule on tubulointerstitial fibrosis in chronic renal failure (CRF) rats and its mechanism against high glucose-induced epithelial to mesenchymal transition (EMT) in renal tubular epithelial cells (HK-2) of its bioactive components., Materials and Methods: The animals were divided into normal group, CRF model group and Huangkui capsule-treated group. Hematoxylin eosin (HE) staining and Masson staining were applied to observe pathological changes in renal tissue of different groups. Biochemical indicators including serum urea nitrogen (BUN), urine protein (UP) and serum creatinine (Scr) were measured according to the manufacturer's instructions of kits. HK-2 cell damaged model was established to access the protection effect and action mechanism of five main flavonoids from Huangkui capsule. The experimental cells were divided into eight groups: control group, model group, positive drug group and five main flavonoids treated groups. The dichlorodihydrofluorescein diacetate (DCFH-DA) assay was used to determine the reactive oxygen species (ROS) in different groups. Western blot was applied to analyze the expression of pathogenesis-related proteins in different groups., Results: The results stated that Huangkui capsule significantly inhibited the elevation of Scr, BUN, UP, the expression of α-smooth muscle actin (α-SMA), phosphorylation-extracellular signal-regulated kinase (p-ERK1/2), NADPH Oxidase 1, NADPH Oxidase 2 and NADPH Oxidase 4 in adenine-induced CRF rats. The main bioactive components of quercetin (QT), hyperoside (HY), isoquercitrin (IQT), gossypetin-8-O-β-D-glucuronide (GG) and quercetin-3'-O-glucoside (QG) at the dosage of 100µM, like NADPH oxidase inhibitor diphenyleneiodonium, exhibited a significant effect on inhibiting the expression of α-SMA, p-ERK1/2, NADPH Oxidase 1, NADPH Oxidase 2 and NADPH Oxidase 4 in high glucose-induced HK-2 cells, especially GG., Conclusions: These results demonstrated that Huangkui capsule and the flavonoids components prevent tubulointerstitial fibrosis in CRF rat involvement in the action mechanism of inhibiting NADPH oxidase/ROS/ERK pathway., (Copyright © 2017. Published by Elsevier B.V.)

Flos Chrysanthemi, dried flower of Chrysanthemum morifolium Ramat, has drawn much attention recently owing to its potential beneficial health effects for human. Flos Chrysanthemi products are usually taken orally and metabolized by intestinal microflora. However, there has been no investigation of the comprehensive metabolic profile of the Flos Chrysanthemi extract by intestinal flora owing to its chemical complexity and the limitations of analytical methods. In this paper, a rapid, sensitive and automated analysis method, ultra-performance liquid chromatography/quadrupole time of flight mass spectrometry including MS E technology and automated data processing Metabolynx™ software, was developed and successfully applied for the biotransformation and metabolic profile of flavonoids in the Flos Chrysanthemi extract by intestinal flora from human and rat. A total of 32 metabolites were detected and tentatively identified in human and rat intestinal bacterial samples. These metabolites indicated that hydrolysis, hydroxylation, acetylation, methylation, hydrogenation and deoxygenation were the major conversion pathways of flavonoids in the Flos Chrysanthemi extract in vitro. Furthermore, the effects of the Flos Chrysanthemi extract on the growth of different intestinal bacteria were detected using an Emax precision microplate reader. Certain pathogenic bacteria such as Enterobacter, Enterococcus, Clostridium and Bacteroides were significantly inhibited by Flos Chrysanthemi, while commensal probiotics such as Lactobacillus and Bifidobacterium were moderately promoted. Our observation provided further evidence for the importance of intestinal bacteria in the metabolism and potential activity of the Flos Chrysanthemi extract. The results will also be helpful for the further pharmacokinetic study of Flos Chrysanthemi and to unravel how it works in vivo., (Copyright © 2016 John Wiley & Sons, Ltd.)

Scutellaria-coptis herb couple is widely used traditional Chinese medicine (TCM) in treating type 2 diabetes; however, the in vivo integrated metabolism of its main bioactive components in type 2 diabetic rats remains unknown. In this paper, ultra-performance liquid chromatography (UPLC) coupled to quadrupole time-of-flight (Q-TOF) and the MetaboLynx™ software combined with mass defect filtering (MDF) together provided unique high throughput capabilities for drug metabolism study with excellent MS mass accuracy and enhanced MS(E) data acquisition. This rapid automated analysis method was successfully applied for screening and identification of the absorbed and metabolized constituents after oral administration of scutellaria-coptis extract to rats. The results showed that a total of 14 metabolites of two parent compounds were detected and tentatively identified in vivo based on the characteristics of their protonated ions. Main parent components of scutellaria-coptis extract such as baicalin and berberine were absorbed into the blood circulation of the rats. Differences of metabolite classes were not observed between normal and type 2 diabetic rat plasma and urine samples. However, the concentrations of baicalin and methylated berberine in type 2 diabetic rat plasma were much higher than those in normal sample. While, the concentrations of these two compounds in type 2 diabetic rat urine were remarkably lower than those in normal sample. This helped maintain a high blood drug concentration which might be beneficial for the treatment of type 2 diabetes. Additionally, the developed method was simple and reliable, revealing that it could be used to rapid screen and propose the structures of active components responsible for pharmacological effects of scutellaria-coptis and to better clarify its action mechanism. This work suggests that the integrative metabolism approach makes a useful template for drug metabolism research of TCMs., (Copyright © 2014. Published by Elsevier B.V.)

Objective: To investigate the metabolic routes and metabolites of myricitrin, an important active ingredient of traditional herbal medicine, yielded by the isolated human intestinal bacteria, which have not been reported previously., Methods: Fresh human fecal samples were collected from a healthy female volunteer and about 100 different bacterial colonies were isolated. Ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry technique combined with Metabolynx™ software was used for analysis of the metabolic profile of myricitrin by the isolated human intestinal bacteria., Results: One hundred different bacterial colonies, which developed on plates, were picked up, and four of them were further identified by using the technique of 16S rRNA gene sequencing due to their relatively strong metabolic capacity toward myricitrin. Most of them belong to Escherichia. Parent compound and three metabolites (quercetin-3-O-rhamnoside, myricetin and quercetin) were detected in the isolated bacterial samples compared with blank samples. The metabolic pathways of myricitrin included deglycosylation and dehydroxylation., Conclusions: These metabolites suggested that myricitrin was first dehydroxylated to quercetin-3-O-rhamnoside and subsequently deglycosylated to quercetin. Additionally, myricitrin could also be deglycosylated to the aglycon myricetin. Moreover, those metabolites might influence the biological effect of myricitrin in vivo, which led to affect the clinical effects of the medicinal plants and traditional herb medicines.

Ethnopharmacological relevance Traditional Chinese Medicines (TCMs) are increasingly used in combination with Western medicine. Scutellaria baicalensis Georgi (Lamiaceae) is a widely used TCM in treating various diseases. However, the in vivo metabolism of its main bioactive flavonoids, baicalin, baicalein, wogonoside and wogonin, needs further study. Materials and methods A systematic method based on ultra-performance liquid chromatography/quadrupole-time-of-flight mass spectrometry (UPLC-Q-TOF/MS) technique combined with Metabolynx TM software was developed to speculate the metabolites and excretion profiles of the main flavonoids in S. baicalensis extract in rats bile, urine and feces samples after oral administration of the extract. Results Four parent components and a total of 15 metabolites were tentatively detected in vivo . All metabolites were detected including sulfate and glucuronide conjugates, hydroxylated, methylated, acetylated and deoxygenated products. Twelve metabolites were from the rat urine, five from the feces and two from the bile. Among them, several products were reported firstly. Conclusion The research provided useful information for further study of the pharmacology and mechanism of action of S. baicalensis extract in vivo and a proposed method which could develop an integrated template approach to analyze screening and identification of biological samples after oral administration of TCMs. [ABSTRACT FROM AUTHOR]

Identification of multicomponent in traditional Chinese medicine (TCM) is complex and time-consuming. The inspection of the full-scan mass chromatograms was usually performed manually, which is labor-intensive. It is difficult to distinguish low response signals from complex chemical background. Furthermore, this process is typically based on earlier knowledge of the chemical composition of TCM, and those molecules that have not been characterized earlier were thus ignored. In this paper, a strategy using UPLC-MS combined with pattern recognition analysis was developed to simplify and quicken the identification of multicomponent in Abelmoschus manihot (L.) Medik. First, complex signals obtained by UPLC-MS were processed using automated data mining algorithm and further processed with multivariate chemometric methods. Multicomponent in Abelmoschus manihot L. can be clearly displayed in S- and VIP-plot. Using this method, 320 peaks which present in Abelmoschus manihot L. were detected. In the next step, accurate mass spectra of the characteristic markers acquired by QTOF MS were used to estimate their elemental formulae and enable structure identification. By searching in METLIN database, 41 components were tentatively identified in Abelmoschus manihot L. Our results showed that UPLC-MS based-pattern recognition analysis approach can be used to quickly identify TCM multicomponent and for standardization of herbal preparations. [ABSTRACT FROM AUTHOR]

The root of Astragalus membranaceus var. mongholicus is one of the most popular herbal medicines worldwide. In order to increase the yield of underground roots of A.membranaceus var. mongholicus, its flowers (AMF) have often been removed in their flowering stage, which produces the flowers as waste being discarded. To explore its phytochemicals and potential value for utilization, the antioxidant activities of extracts from AMF were evaluated by a free radical scavenging assay and reducing power assay. The total phenols and flavonoids, as well as the individual compounds, in different extracts of AMF were also investigated. The results showed that the extract ME obtained from AMF through macroporous resins separation exhibited strong antioxidant activities, which were close to those of positive control BHT. ME was rich in phenolic acids and flavonoids, and the contents reached 108.42 mg gallic acid equivalents/g and 265.70 mg rutin equivalents/g, respectively. A total of 31 compounds, including four phenolic acids, nineteen flavonoids, three isoflavones, two pterocarpans, and three saponins, were identified using UPLC-QTOF-MS in ME. Quantitative analysis of sixteen components in the extracts of AMF showed that flavonoids were the predominant constituents, especially for the compounds of hyperoside, rutin, and isorhamnetin-3-O-β-d-glucoside. [ABSTRACT FROM AUTHOR]

A novel and generally applicable approach was established to hierarchically identify the bioactive components of a medicinal herb by preparative high-performance liquid chromatography (prep-HPLC) and a selective knock-out strategy. In this study, the targeted components of an herbal medicine were separated and knocked out using prep-HPLC. Subsequently, the contributions of the different target components to the overall effect of the medicinal herb were comparatively evaluated and differentiated by a heat map and a 3D score plot. This approach was successfully applied to investigate the bioactive constituents of safflower. The contributions of 11 components to the overall effect of safflower were as follows: anhydrosafflor yellow B ( 10) > 6-hydroxykaempferol 3,6-di- O - β - d -glucoside ( 8 ) > hydroxysafflor yellow A ( 3 ) > kaempferol 3- O - β -rutinoside ( 11 ) > 6-hydroxykaempferol 3- O - β -rutinoside ( 9 ) > 6-hydroxykaempferol 3,6-di- O - β - d -glucoside-7- O - β - d -glucuronide ( 4 ) > 6-hydroxyapigenin 6- O - β - d -glucoside-7- O - β - d -glucuronide ( 6 ) > cytidine ( 1 ) > 6-hydroxykaempferol 3- O - β -rutinoside-6- O - β - d -glucoside ( 7 ) > 6-hydroxykaempferol 3,6,7-tri- O - β - d -glucoside ( 5 ) > adenosine ( 2 ). These results demonstrate that quinochalcone C -glycosides ( 3 and 10 ) and some flavonoid glycosides containing C7-OH (such as 8 , 9 and 11 ) made a greater contribution to the overall effect of safflower than the other components that were knocked out. The results provided an important reference for improving quality control and further development of safflower products. And this approach should also be useful for investigating the bioactive constituents of other medicinal herbs. [ABSTRACT FROM AUTHOR]

Buddleoside (also known as linarin) as the major flavonoid in Chrysanthemum morifolium Ramat., has been reported to possess a wide range of pharmacological activities. The human intestinal microbiota might have an important impact on drug metabolism and ultimately on the drug oral bioavailability. However, the interaction of the buddleoside with human intestinal bacteria remains unknown. In this study, the conversion of buddleoside by different bacteria from human feces was firstly investigated. A reliable, sensitive and rapid analytical method, ultra performance liquid chromatography was established and successfully applied to investigate the metabolites and metabolic profile of buddleoside by human intestinal bacteria. Among the isolated bacteria, four strains including Escherichia sp. 4, Escherichia sp. 34, Enterococcus sp. 45 and Bacillus sp. 46 showed more powerful conversion capability. Based on the accurate mass data and the characteristic MS n product ions, the parent and six metabolites were detected and tentatively identified compared with blank samples. The metabolites were produced by four main metabolic pathways including deglycosylation, acetylation, methylation and hydroxylation. Buddleoside could be firstly converted to its aglycon acacetin (M2) by the majority of the isolated intestinal bacteria. Subsequently, M2 was further metabolize to its methylated (M3), acetylated (M4), hydroxylated (M5) and hydrogenated product (M6). However, acacetin-7-glucosid (M1) was obtained only from the minor bacterial samples like Bacillus sp. 46. To further explain the metabolism of buddleoside, the β- d -glucosidase and α- l -rhamnosidase activities of four strains were analyzed. Bacillus sp. 46 could only produce α- l -rhamnosidase, while the other three strains showed two kinds of enzyme activities. Furthermore, the activities of α- l -rhamnosidase and β- d -glucosidase reached the highest level at 12–18 h and 10–12 h, respectively. The metabolic routes and metabolites of buddleoside produced by human intestinal microflora were firstly reported in this paper. The results will be very helpful for the further investigation of the pharmacokinetic research of buddleoside and to unravel how it works in vivo . [ABSTRACT FROM AUTHOR]

Abstract: Ethnopharmacological relevance:: Abelmoschus manihot (L.) Medik. var. manihot is one of the most commonly used Chinese medicines and has played an important role in treating chronic glomerulonephritis and diabetic nephropathy. Aim of the study:: Metabolites identification of traditional Chinese medicine (TCM) is a complex and time-consuming process due to the complicity of TCM and subsequent large number of detected ions. In this paper, UPLC–MS combined with pattern recognition analysis approach were used to simplify and quicken the identification of the metabolites of Abelmoschus Manihot. Materials and methods:: Rat urine samples were collected before (as control sample) and after Abelmoschus Manihot administration. Pattern recognition analysis method was used to differentiate components between Abelmoschus Manihot-treated group and its controlled comparison. These components could be considered as Abelmoschus Manihot-related metabolites in vivo. Results:: LC–MS based metabolomics could be an advanced tool to help us find metabolites with regards to its capacity of processing large datasets, differentiating and classifying of sample groups, as well as its indiscriminative nature of biomarker and metabolite identification. Using this method, seven metabolites were identified, which are flavonoid aglycone glucuronidation, sulfatation, and methylation metabolites. Conclusion:: Our results showed that UPLC–MS based- pattern recognition analysis approach can be used to quickly identify Abelmoschus Manihot related metabolites in biological fluids. Furthermore, this work demonstrates the potential application of combining the UPLC–MS approach with the metabolomics approach in identifying the metabolites of TCM. [Copyright &y& Elsevier]