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Rhubarb, a traditional herb, has been used in clinical practice for hundreds of years to cure constipation, but its mechanism is still not clear enough. Currently, growing evidence suggests that intestinal flora might be a potential target for the treatment of constipation. Thus, the aim of this study was to clarify the laxative effect of rhubarb via systematically analyzing the metagenome and metabolome of the gut microbiota. In this study, the laxative effects of rhubarb were investigated by loperamide-induced constipation in rats. The gut microbiota was determined by high-throughput sequencing of 16S rRNA gene. Ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry was used for fecal metabolomics analysis. The data showed that rhubarb could significantly shorten gastrointestinal transit time, increase fecal water content and defecation frequency, improve gastrointestinal hormone disruption, and protect the colon mucus layer. Analysis of 16S rRNA gene sequencing indicated that rhubarb could improve the disorder of intestinal microbiota in constipated rats. For example, beneficial bacteria such as Ligilactobacillus, Limosilalactobacillus, and Prevotellaceae UCG-001 were remarkably increased, and pathogens such as Escherichia-Shigella were significantly decreased after rhubarb treatment. Additionally, the fecal metabolic profiles of constipated rats were improved by rhubarb. After rhubarb treatment, metabolites such as chenodeoxycholic acid, cholic acid, prostaglandin F2α, and α-linolenic acid were markedly increased in constipation rats; in contrast, the metabolites such as lithocholic acid, calcidiol, and 10-hydroxystearic acid were notably reduced in constipation rats. Moreover, correlation analysis indicated a close relationship between intestinal flora, fecal metabolites, and biochemical indices associated with constipation. In conclusion, the amelioration of rhubarb in constipation might modulate the intestinal microflora and its metabolism. Moreover, the application of fecal metabolomics could provide a new strategy to uncover the mechanism of herbal medicines.Key points• Rhubarb could significantly improve gut microbiota disorder in constipation rats.• Rhubarb could markedly modulate the fecal metabolite profile of constipated rats.

This study was designed to investigate the protective effects and mechanisms of acteoside on DKD in diabetes male db/db mice and high glucose-induced HK-2 cells. The diabetes db/db mice were divided randomly into model group, metformin group, irbesartan group, and acteoside group. We observed the natural product of acteoside exhibiting a significant effect in renal protection through analyzing of biochemical indicators and endogenous metabolites, histopathological observations, and western blotting. HK-2 cells subjected to high glucose were used in invitro experiments. The molecular mechanisms of them were investigated by RT-PCR and western blot. Acteoside prevents high glucose-induced HK-2 cells and diabetes db/db mice by inhibiting NADPH/oxidase-TGF-β/Smad signaling pathway. Acteoside regulated the disturbed metabolic pathway of lipid metabolism, glyoxylate and dicarboxylate metabolism, and arachidonic acid metabolism. We discovered the natural product of acteoside exhibiting a significant effect in renal protection. This study paved the way for further exploration of pathogenesis, early diagnosis, and development of a new therapeutic agent for DKD.

Houpo Mahuang Decoction (HPMHD is one of the classic traditional Chinese prescriptions that has been used in the treatment of asthma. The therapeutic effects and mechanism of HPMHD in aggravated asthma remain to be explored, especially from the perspective of metabolomics and Transient Receptor Potential Vanilloid-1 (TRPV1)/CaTo investigate the therapeutic and metabolic regulatory effects and the underlying mechanism of HPMHD in asthmatic rats.The asthmatic rats were administered with the corresponding HPMHD (at dosages of 5.54, 11.07, 22.14 mg/kg). Then inflammatory cells in peripheral blood and bronchoalveolar lavage fluid (BALF) were counted, the levels of interleukin (IL)-4 and IL-13 in BALF were measured, and the changes in enhanced pause (Penh) and pathological damage of lung tissues were also detected to evaluate the protective effects of HPMHD. The serum metabolic profile of HPMHD in asthmatic rats was explored using Ultra-High-Performance Liquid Chromatography-mass spectrometer (UHPLC-MS), and the regulatory effects on TRPV1 and TJs of HPMHD in asthmatic rats were detected by Western blotting analysis. In vitro, 16HBE cells were stimulated with IL-4 plus SOHPMHD significantly attenuated the airway inflammation of asthmatic rats, and reduced the levels of inflammatory cells in peripheral blood and BALF as well as the levels of IL-4 plus IL-13 in BALF. In addition, the airway hyperresponsiveness and lung pathological damage were alleviated. Serum metabolomic analysis showed that 31 metabolites were differentially expressed among the normal saline-, model-, and HPMHD-treated rats. Pathway enrichment analysis showed that the metabolites were involved in 45 pathways, among which, TJs regulation-relevant pathway was associated with the CaThe current study indicates that HPMHD alleviates rat asthma and participates in the regulation of serum metabolism. The anti-asthma effects of HPMHD might be related to the protection of TJs by inhibiting the intracellular Ca

Graphical abstract, Highlights • Bruceine A displays potent anti-pancreatic cancer activity in vitro and in vivo. • Phosphoproteomic analyses identify bruceine A induces phosphorylation of p38α MAPK. • Octet system and microscale thermophoresis reveal p38α MAPK has high affinity for bruceine A. • Molecular simulations illustrate determinants of bruceine A binding with p38α MAPK., Pancreatic cancer remains one of the cancers with the poorest prognosis bearing an overall 5-year survival rate of about 5%. Efficient new chemotherapic drugs are still highly desired. Here, bruceine A, a quassinoid identified from the dried fruits of Brucea javanica (L.) Merr., displayed the most potent anti-proliferation activity against pancreatic cancer in vitro and in vivo. Phosphoproteomic analysis revealed p38α MAPK phosphorylation was involved in bruceine A’s action in MIA PaCa-2 cells. Utilizing fortebio octet system and microscale thermophoresis, we found p38α MAPK had high affinity for bruceine A. Molecular docking and molecular dynamic simulations showed that bruceine A widely bound to residues (Leu171, Ala172, Met179, Thr180, Val183) in P-loop of p38α MAPK. Key determinants of bruceine A binding with P-loop of p38α MAPK were 19-C Created by potrace 1.16, written by Peter Selinger 2001-2019 O, 22-CH3, 32-CH3, and 34-CH3. Taken together, our findings demonstrate that bruceine A binds directly to p38α MAPK, which can be used to probe the role of p38α MAPK phosphorylation in pancreatic cancer progression, and as a novel lead compound for pancreatic cancer therapy.

Ulcerative colitis (UC), a kind of inflammatory bowel disease, is generally characterized by chronic, persistent, relapsing, and nonspecific ulceration of the bowel, which is widespread in the world. Although the pathogenesis of UC is multifactorial, growing evidence has demonstrated that gut microbiota and its metabolites are closely related to the development of UC. Lizhong decoction (LZD), a well-known classical Chinese herbal prescription, has been used to clinically treat UC for long time, but its mechanism is not clear. In this study, 16S rRNA gene sequencing combining with untargeted metabolomics profiling was used to investigate how LZD worked. Results indicated that LZD could shape the gut microbiota structure and modify metabolic profiles. The abundance of opportunistic pathogens such as Clostridium sensu stricto 1, Enterobacter, and Escherichia-Shigella correlated with intestinal inflammation markedly decreased, while the levels of beneficial bacteria including Blautia, Muribaculaceae_norank, Prevotellaceae UCG-001, and Ruminiclostridium 9 elevated in various degrees. Additionally, fecal metabolite profiles reflecting microbial activities showed that adenosine, lysoPC(18:0), glycocholic acid, and deoxycholic acid notably decreased, while cholic acid, α-linolenic acid, stearidonic acid, and l-tryptophan significantly increased after LZD treatment. Hence, based on the systematic analysis of 16S rRNA gene sequencing and metabolomics of gut flora, the results provided a novel insight that microbiota and its metabolites might be potential targets for revealing the mechanism of LZD on amelioration of UC. Key Points • The potential mechanism of LZD on the amelioration of UC was firstly investigated. • LZD could significantly shape the structure of gut microbiota. • LZD could notably modulate the fecal metabolic profiling of UC mice.

Hui Medicine ZhaLi NuSi Prescription (ZLNS) is described in "Hui Hui Prescription," and it has been used to treat cerebral infarction in Hui Region, China. In this study, a rapid and reliable ultra-performance liquid chromatography coupled with mass spectrometry (UPLC-MS/MS) method was established and applied to simultaneously determine geniposidic acid, oxypaeoniflorin, hydroxysafflor yellow A, caffeic acid, magnoflorine, paeoniflorin, ferulic acid, β-ecdysterone, icariin, rhein, and baohuoside I in rat plasma. The pharmacokinetic parameters of these components and the influence of essential oils (EOs) on them were investigated in normal rats. The results showed that the pharmacokinetic parameters (AUC0 - t , AUC0 - ∞ , t1/2 , tmax , cmax ) of the aforementioned compounds were significantly changed after co-administering with ZLNS EO. The AUC values of oxypaeoniflorin, paeoniflorin, ferulic acid, and baohuoside I with EOs were decreased significantly. This is the first report for the comparative pharmacokinetic study of ZLNS bioactive components in normal rats, which may provide the basis for drug interaction study in vivo and insight into their clinical applications.

The seed of Ziziphus jujuba Mill. var. spinosa (Bunge) Hu ex H. F. Chou (ZSS) is often used as a traditional Chinese medicine for insomnia due to its sedative and hypnotic effects, but the mechanism underlying this effect has not been thoroughly elucidated. In this study, an insomnia model induced by intraperitoneal injection of DL-4-chlorophenylalanine suspension in Sprague-Dawley rats was adopted to investigate the therapeutic effect of ZSS extract. Metabolomics analyses of plasma and urine as well as 16S rRNA gene sequencing of the intestinal flora were performed. The relationships between the plasma and urine metabolites and the intestinal flora in insomnia rats were also analyzed. The results showed that changes in plasma and urine metabolites caused by insomnia were reversed after administration of ZSS, and these changes were mainly related to amino acid metabolism, especially phenylalanine metabolism. The results of 16S rRNA gene sequencing and short-chain fatty acid determination showed that the ZSS extract could reverse the imbalance of intestinal flora caused by insomnia and increase the contents of SCFAs in feces. All of these improvements are mainly related to the regulation of inflammation. Therefore, it is concluded that insomnia, which alters metabolic profiles and the intestinal flora, could be alleviated effectively by ZSS extract.

The rhizome of Polygonatum cyrtonema (Polygonati Rhizoma) is widely consumed as medicine-homology-food in Asia for its tonic effect, which can be enhanced by traditional steam-sun drying for nine cycles. However, the multi-constituents variation in this process was unclear, and the necessity of nine cycles should be further discussed. In this study, the multiple constituents, including saccharides, amino acids, nucleosides and bases, lipids, saponins, homoisoflavones and cinnamamides, in P. cyrtonema treated with sun drying, heated air drying, each cycle of steam-heated air drying, infrared drying and microwave drying were compared. The results showed that the content of total saccharides increased in samples from one to four cycles of steam-heated air drying (365.0 - 945.6 mg/g) and decreased from four to nine (945.6 - 288.0 mg/g). The content of fructose increased in samples from one to six cycles (29.9 - 234.7 mg/g) and decreased from six to nine (234.7 mg/g - 177.7 mg/g). The abundance of most phospholipids and free fatty acids increased continuously from one to nine cycles while most of the amino acids, nucleosides and bases showed continuous declining trend. Principal component analysis showed that the samples treated with one to four cycles were wider in distance than four to nine, indicating the chemical composition tending to be stable after fourth steaming. If taking total saccharides, fructose, and phospholipids as the major quality indicator, four cycles of steam-heated air drying processing should be the ideal postharvest processing method to obtain better taste, flavor and functionality. Samples treated with heated-air drying and infrared drying were far in distance from steaming ones by hierarchical cluster analysis, which means these processing methods were not suitable to replace the traditional steam-sun drying process. Collectively, the above results will not only provide novel processing methods that will obtain the high active ingredients for P. cyrtonema, but also shed light on the optimization of processing technology for the industrial production of medicinal crops which need nine cycles of steam-sun drying processing.

Trichosanthin (root of Trichosanthes kirilowii Maxim.) has received much attention, but T. kirilowii has received little systematic analysis. A comprehensive evaluation of flavones and triterpenes in nine different parts of T. kirilowii would, thus, be of value to fully understand the potential medicinal properties of T. kirilowii. A hierarchical extraction—simultaneous determination method was established, which based on different types of components had different response on the MS detector. The hierarchical extraction method extracted 3, 29-dibenzoyl rarounitriol solely, which response lowly on the MS detector in the mixture of one step extract, in order to obtain accurate results in the next UHPLC-MS/MS assay. By this method, ten flavones, three tetracyclic triterpenoids, and one pentacyclic triterpenoid in the extracts of T. kirilowii were determined. The results showed the level of flavonoids in leaves was higher than those in the other samples, tetracyclic triterpenoids mainly existed in root and root bark, and pentacyclic triterpenoid, especially 3, 29-dibenzoyl rarounitriol was concentrated in the seed kernel. According to principal component analysis, the F values of the leaves of all varieties were generally greater than those in other parts. The leaves might provide a supplementary source of flavonoids, and the root and root bark could be a good choice for tetracyclic triterpenoids. The analysis results revealed the distribution of flavones and triterpenes in different parts of T. kirilowii and provided reference for the research and rational utilization of T. kirilowii.

Background Indoxyl sulfate (IS) and p-cresyl sulfate (pCS) are two key protein-bound uremic toxins that accumulate in patients with end-stage renal disease. IS and pCS cannot be efficiently removed by conventional hemodialysis because they are highly bound to proteins. One promising means to optimize the removal of protein-bound uremic toxins involves using binding competitors to liberate uremic toxins from protein-binding partners. Purpose In this study, we try to identify potential binding competitors that can enhance the dialysis removal of IS and pCS in natural compounds of phytomedicine. Methods We employed microdialysis to evaluate whether Danhong injection (DHI) and its salvianolic acids can increase the free fractions of IS and pCS and thus improve their dialysis efficiency in vitro. Furthermore, we confirmed the positive effects of DHI and salvianolic acids in vivo on chronic kidney disease model rats in which IS and pCS had heavily accumulated. Results DHI significantly increased the dialysis efficiency of IS and pCS by 99.13% and 142.00% in vitro (10-fold dilution), respectively, and by 135.61% and 272.13% in vivo (4.16 ml/kg). Salvianolic acids including lithospermic acid (LA), salvianolic acid A (SaA), tanshinol (DSS), caffeic acid (CA), salvianolic acid B (SaB), protocatechuic aldehyde (PA) and rosmarinic acid (RA) significantly enhanced the dialysis removal of IS and pCS in a concentration-dependent manner. LA, the best competitor of the tested salvianolic acids, increased dialysis efficiency levels of IS and pCS by 197.23% and 198.31% in vitro (400 μM), respectively, and by 119.55% and 127.56% in vivo (24.69 mg/kg). Conclusion The removal of protein-bound uremic toxins IS and pCS using DHI or salvianolic acids as protein-bound competitors is superior to previously reported strategies and drugs and may contribute to clinical hemodialysis therapeutic practice.

Ethnopharmacological relevance As recorded in traditional Chinese medicine (TCM) theory, Genkwa Flos (YH) and Glycyrrhizae Radix et Rhizoma (GC) compose one herbal pair of the so-called “eighteen incompatible medicaments”, which indicate pairs of herbs that are mutually incompatible and that theoretically should not be applied simultaneously. However, the theory has been called into question due to a lack of evidence. Aims of study In this study, the incompatibility of YH and GC was investigated based on an assessment of the toxic effects of their combination by traditional safety methods and a modern metabonomic approach. Materials and methods Sprague-Dawley rats were used to evaluate the subacute toxicity of YH and YH-GC. The serum, urine, and several tissues were collected for biochemical analysis, histopathological examination, and metabonomic analysis. Results Rats exposed to a dose of 1.0 g/kg YH (3 times of the Chinese Pharmacopoeia maximum dose) exhibited toxicity of the heart, liver, kidney and testes, and rats exposed to a YH-GC combination (1.0 g/kg YH + 1.0 g/kg GC) exhibited similar hepatotoxicity, which aggravated renal and reproductive toxicity. Following this, a metabonomic study tentatively identified 14 potential biomarkers in the YH group and 10 potential biomarkers in the YH-GC group, and metabolic pathways were then constructed. YH disturbed the pathways of glycerophospholipid metabolism, primary bile acid biosynthesis, and sphingolipid metabolism, while YH-GC combination induced disruptions in phenylalanine, tyrosine and tryptophan biosynthesis, tyrosine metabolism, and glycerophospholipid metabolism. Conclusion The toxicities of YH and YH-GC combination above the Chinese Pharmacopoeia dose were obvious but different. Metabonomics combined with biochemical and histopathological methods can be applied to elucidate the toxicity mechanism of the YH-GC combination that caused liver, kidney and reproductive injuries in rats.

Zingiber officinale and Panax ginseng, as well-known traditional Chinese medicines, have been used together to clinically treat ulcerative colitis with synergistic effects for thousands of years. However, their compatibility mechanism remains unclear. In this study, the shift of gut microbiome and fecal metabolic profiles were monitored by 16S rRNA sequencing technology and ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry analysis, respectively, which aimed to reveal the synergistic mechanism of Zingiber officinale and Panax ginseng on the amelioration of ulcerative colitis. The results showed that the relative abundance of beneficial bacteria (such as Muribaculaceae_norank, Lachnospiraceae NK4A136 group and Akkermansia) was significantly increased and the abundance of pathogenic bacteria (such as Bacteroides, Parabacteroides and Desulfovibrio) was markedly decreased after the intervention of Zingiber officinale-Panax ginseng herb pair. And a total of 16 differential metabolites related to ulcerative colitis were identified by the metabolomics analysis, which were majorly associated with the metabolic pathways, including arachidonic acid metabolism, tryptophan metabolism, and steroid biosynthesis. Based on these findings, it was suggested that the regulation of the gut microbiota-metabolite axis might be a potential target for the synergistic mechanism of Zingiber officinale-Panax ginseng herb pair in the treatment of ulcerative colitis. Furthermore, the integrated analysis of microbiome and metabolomics used in this study could also serve as a useful template for exploring the mechanism of other drugs.

The incidence and mortality of bronchial asthma are increasing, and respiratory syncytial virus (RSV) is widely regarded as the common cause of clinical exacerbation of asthma. Ma-Xing-Gan-Shi decoction (MXGSD), a classic traditional Chinese medicine prescription, is well-known for treating respiratory diseases, while the mechanism of effecting on RSV-exacerbated asthma remains to be explored.In this study, we investigated the mechanism by which MXGSD exerts a protective effect on asthma exacerbated by RSV in vivo and in vitro.MXGSD is composed of four Chinese medicine, including Ephedra intermedia SchrenkC.A.Mey. (herbaceous stem, 27g), Prunus armeniaca L. (dry seed, 27g), Glycyrrhiza uralensis Fisch. (radix and rhizome, 18g), and Gypsum fibrosum (main component: CaSOThe results suggested that MXGSD could reduce the levels of inflammation cells, airway hyperresponsiveness, and pathological damage of lung tissue. TMT quantitative proteomics analysis and further experimental exploration revealed that MXGSD could reduce the levels of IL-4, IL-13, PGEMSGSD showed a protective effect on RSV-exacerbated asthma, which may be related to its regulation of TRPV1 expression and reduction of Th2 cytokines and neurogenic inflammatory mediators. It may provide an objective basis and reference for the clinical application of MXGSD.

Plant-microorganism interaction in the rhizosphere is thought to play an important role in the formation of soil fertility, transformation and absorption of nutrients, growth and development of medicinal plants, and accumulation of medicinal ingredients. Yet, the role that they play in the phthalides accumulation of Angelica sinensis (Oliv.) Diels remains unclear. In the present study, we report a correlative analysis between rhizosphere microorganisms and phthalides accumulation in A. sinensis from Gansu, China where was the major production areas. Meanwhile, Bacillus was explored the potential functions in the plant growth and phthalide accumulation. Results revealed that the common bacterial species detected in six samples comprised 1150 OTUs which were involved in 368 genera, and predominant taxa include Actinobacteria, Acidobacteria, and Proteobacteria. The average contents of the six phthalides were 4.0329 mg/g. The correlation analysis indicated that 20 high abundance strains showed positive or negative correlations with phthalides accumulation. Flavobacterium, Nitrospira, Gaiella, Bradyrhizobium, Mycobacterium, Bacillus, RB41, Blastococcus, Nocardioides, and Solirubrobacter may be the key strains that affect phthalides accumulation on the genus level. By the plant-bacterial co-culture and fermentation, Bacillus which were isolated from rhizosphere soils can promote the plant growth, biomass accumulation and increased the contents of the butylidenephthalide (36∼415%) while the ligustilide (12∼67%) was decreased. Altogether, there is an interaction between rhizosphere microorganisms and phthalides accumulation in A. sinensis, Bacillus could promote butylidenephthalide accumulation while inhibiting ligustilide accumulation.

Background: The frankincense and myrrh can be used in traditional Chinese medicine (TCM) clinic with the treatment of stroke, while its mechanism is still unclear. The aim of this study was to investigate the effects of boswellic acid (BA) and myrrh sesquiterpene (MS) compositions on ischemic stroke in rats. Methods: The ischemic stroke rat model induced by ischemia-reperfusion (I/R) after transient middle cerebral artery occlusion (MCAO) was adopted and treated with boswellic acid and myrrh sesquiterpenes with proportions compatibility (BA-MS) of 10:1, 5:1 and 20:1. Biochemical indexes, histopathological analysis, metabolomics analysis of plasma and urine, as well as 16S rDNA sequencing of gut microbiota and short chain fatty acids were investigated. Results: BA and MS could obviously significantly ameliorated neurological deficits scores, reduced cerebral infarction area in I/R, and improve the damage of brains and lungs of MCAO rats. Additionally, it could significantly reduce the expression of neuroinflammatory factors (IL-1β, nNOS, BDNF, NGF) and angiogenic growth factors (ET-1, PAI-1, PDGF, VEGF, VWF), and significantly increase the expression of nerve indicators (TDP-43, NeuN) and angiogenic growth factors (Ang Ⅱ, TGF-β1, bFGF). After the administration of BA-MS, the metabolites of ischemic stroke in plasma and urine of ischemic stroke of rats were reversed, mainly correlated with the metabolic pathway of linoleic acid metabolism (impact value =1), and the gut microbiota and SCFAs metabolism of cecal contents were also improved. Conclusions: BA-MS can improve brain injury of MCAO rats via Ang/Tie and TLR4 signaling pathway, improve metabolism disorder by regulating linoleic acid metabolism pathway, and ameliorate gut microbes and their metabolic disorders.

Rehmanniae Radix Preparata (RR) and Corni Fructus (CF), well-known traditional Chinese medicines (TCMs), were generally used together in the clinical practices to treat chronic kidney disease (CKD) with synergistic effects for thousands of years, but their combination mechanism remains largely unknown so far. Recent evidences have implicated intestinal flora as potential targets for the therapy of CKD. In this study, the CKD rat model was induced by adenine. The levels of proteinuria, serum creatine (SCr), blood urea nitrogen (BUN) and creatinine clearance (Ccr) were used to assess the cooperation effect of RR and CF. Furthermore, high-throughput 16S ribosomal RNA (rRNA) gene sequencing combined with fecal metabonomics based on UPLC-Q-TOF-MS/MS were applied to explore the variations of intestinal flora and their metabolic profiles. 16S rRNA gene sequencing data indicated that CKD rats treated with RR, CF and RC showed the differences in the composition of gut microbiota. The abundance of beneficial bacteria including Ruminococcaceae UCG-014, Ruminococcus 1, Prevotellaceae_NK3B31_group, Lachnospiraceae NK4A136 group and Lachnospiraceae UCG-001 were elevated in various degrees, while the opportunistic pathogen such as Desulfovibrio was markedly decreased after the treatment. Moreover, fecal metabolite profiles revealed 15 different metabolites associated with CKD. These metabolites were mainly involved in the related metabolic pathways such as amino acid metabolism, bile acids metabolism and glycerophospholipid metabolism. The results implied that gut flora and their metabolites might play a vital role in the progress of CKD, which provided a potential target for the development of novel drugs for the therapy of CKD.

Goji berries are used as functional food for hundreds of years in Asia, Europe, North America and Austria, and are popular for nutritive properties in global. Commercial Goji berries are mainly produced in Ningxia, Xinjiang, Gansu, Qinghai and Inner Mongolia of China. However, the Goji berries produced in these regions exhibited different appearance and taste. Thus, characterization of the nutritional components in Goji berries produced in these regions could provide the guidance for application of them. In this study, 94 samples were collected, and a total of 20 amino acids, 17 nucleosides and nucleobases, 4 sugars and protein were determined by UHPLC-MS/MS, HPLC-ELSD or UV, and the variation was illustrated through heatmap clustering analysis, PCA and PLS-DA. The results showed that Goji berries from Xinjiang were rich in protein than the samples from other regions; those from Gansu and Ningxia were rich in amino acids, nucleosides and nucleobases; and those from Jiuquan of Gansu and Qinghai were rich in sugars. Heatmap clustering and PCA analysis results showed that all the samples exhibited a significant spatial aggregation, and the producing regions located along the Yellow River (belonging to the Hetao plain) produced Goji berries with the similar chemical profile. Additionally, PLS-DA analysis results showed that fructose and glucose were the predominant markers to distinguish Goji berries from different producing regions.

Starch nanoparticles (SNPs) with different morphology and particle size can be prepared by modulating the reaction conditions over SNPs preparation. This study was to synthesize various SNPs by using ultrasound assisted nanoprecipitation method, and characterized by particle size analysis, SEM and XRD performing. SNPs were successfully produced via nanoprecipitation and the particle size were controlled in the range of 95 to 150 nm. Moreover, variously different morphologies were obtained when using corn, potato or Trichosanthes kirilowii pulp (TKP) starch to produce nanoparticles, including fiber, flake and film. Results shown film TKP SNPs demonstrated an improved urea adsorption rate to 135.60 mg/g with the highest qm at 1.00 mg/mL. SNPs can be developed using ultrasound assisted nanoprecipitation method and the particle size together with surface morphology can be varied according to the source of starch and preparation method, while surface morphology is the key factor in altering adsorption performance.

Ethnopharmacological relevance Mulberry leaves have been used as traditional hypoglycemic medicine-food plant for thousand years in China. According to traditional Chinese medicine theory, type 2 diabetes mellitus (T2DM) belongs to the category of XiaoKe. Presently, the research of mulberry leaf hypoglycemic and lipid-lowering direction is mature, but the curative effects of alkaloids, flavonoids, polysaccharides, and other bioactive ingredients and the related mechanism is still unclear. Aim of the study This paper aims to study the efficacy and mechanism of alkaloids, flavonoids, polysaccharides, and other bioactive components in mulberry leaves in the treatment of T2DM individually. Materials and methods The determination of levels of fasting blood glucose (FBG), triglyceride (TG) and total cholesterol (T-Cho), and pyruvate kinase (PK), hexokinase (HK), and alanine aminotransferase (ALT/GPT) of in plasma of diabetic mice. Urine metabolomics was analyzed by UPLC-QTOF/MS to evaluate differential metabolites from multiple metabolic pathways. The glucose uptake of HepG2 cells and 3T3-L1 cells. Expression of Caspase-3 and caspase-9, inflammatory injury and p38MAPK/NF-κB signaling pathway in GLUTag cells. Results Our study revealed alkaloids, flavonoids, and polysaccharides in mulberry leaf could increase the levels of PK, HK, and ALT/GPT, and decrease the levels of TG and T-Cho significantly, and regulate glucose, amino acid, and lipid metabolism. Furthermore, 1-deoxynojirimycin (DNJ) and isoquercitrin (QG) both could increase glucose uptake and promote differentiation of HepG2 cells, increase PPARγ, C/EBPα and SREBP-l expression in 3T3-L1 cells, and inhibit AGEs-induced injury and apoptosis in GLUTag cells, reduce the expression of proteins related to AGEs/RAGE and p38MAPK/NF-κB pathway. Notably, isoquercitrin exhibited more pronounced anti-diabetic efficacy. Conclusions The alkaloids, flavonoids, and polysaccharides from mulberry leaf exhibited hypoglycemic activity through the regulation of glucose, amino acid, and lipid metabolism. 1-DNJ and QG increased glucose uptake and promoted differentiation of HepG2 cells, increased PPARγ, C/EBPα and SREBP-l expression in 3T3-L1 cells, and inhibited AGEs-induced injury and apoptosis in GLUTag cells via the AGEs/RAGE and p38 MAPK/NF-κB pathway.

Chronic kidney disease (CKD) is a worldwide public health problem. Uremic retention solutes such as indoxyl sulfate (IS) and p-cresyl sulfate (PCS) are accumulated in CKD patients and are associated with the incidence of CKD progression. Amino acids are the major precursors of uremic retention solutes in gut. The dynamic change of amino acid metabolism in the gut during CKD progression has not been reported previously. In this paper, we studied the dynamic change of gut IS/PCS precursor and amino acid metabolism profile during CKD progression in 5/6 nephrectomized (5/6Nx) rats model. The related gut microbiota and metagenome profile was also studied. Rat plasma, urine and feces were collected at different time points after nephrectomization. Plasma IS and PCS, fecal indole (the precursor of IS), p-cresol (the precursor of PCS) and 19 kinds of amino acids were analyzed by LC-MS. During CKD progression, 5/6 Nx rats showed increased plasma IS, PCS concentration and increased fecal indole, p-cresol concentration. 5/6 Nx rats also showed disordered gut amino acids metabolism profile which became more significant along with the progession of CKD. The abundance of some specific gut bacteria also changed significantly in 5/6 Nx rats. The 16S rDNA sequencing data of gut microbiota was further analyzed by an online tool PICRUSt, a large-scale computational method for metagenomes prediction with 16S rDNA sequencing data. The content of each gene was compared between the two groups by Mann-Whitney U test, and then the significantly regulated genes in 5/6 Nx group were subjected to KEGG website. The amino acid metabolism related genes were picked out. Most of these genes are more abundant in 5/6 Nx groups. Our study showed that gut amino acids metabolism profile was disordered with CKD progression, which was highly related to the gut microbiota dysbiosis and metagenome change. And that regulation of gut amino acids metabolism pathway may be a possible way to intervene the progression of CKD.

Aims Rehmanniae Radix Preparata (RR) and Corni Fructus (CF) are commonly used together for the treatment of chronic kidney disease (CKD) in the clinical practices for thousands of years. However, little information on their synergy mechanism is available. Main methods In this study, an integrated approach combining ultra performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS)-based metabonomics and network pharmacology was adopted to elucidate the cooperation mechanism of RR and CF on the amelioration of CKD. Furthermore, the targets from network pharmacology and metabolism pathways were jointly analyzed. Finally, the activities of key metabolic enzymes were experimentally validated by ELISA. Key findings Metabolic profiling indicated that the metabolic disturbance in plasma was markedly alleviated after treatment. Nine putative biomarkers mainly involving in phenylalanine, tyrosine and tryptophan biosynthesis and tyrosine metabolism were identified. Moreover, the compound-target-pathway network of RR and CF for CKD treatment was constructed by network pharmacology, which was related to tyrosine metabolism and arginine and proline metabolism. The results were partly consistent with the findings of plasma metabolomics. Significance In conclusion, this study solidly supported and enhanced current understanding of the synergy effects of RR and CF on CKD. Meanwhile, it also confirmed the feasibility of combining metabolomics and network pharmacology to identify active components and elucidate the pharmacological effects of traditional Chinese medicines (TCMs).

Pancreatic cancer is one of the most aggressive cancers with a poor prognosis and 5-year low survival rate. In the present study, we report that bruceine A, a quassinoid found in Brucea javanica (L.) Merr. has a strong antitumor activity against human pancreatic cancer cells both in vitro and in vivo. Human proteome microarray reveals that 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 4 (PFKFB4) is the candidate target of bruceine A and both fluorescence measurement and microscale thermophoresis suggest bruceine A binds to PFKFB4. Bruceine A suppresses glycolysis by inhibiting PFKFB4, leading to cell cycle arrest and apoptosis in MIA PaCa-2 cells. Furthermore, glycogen synthase kinase-3 β (GSK3β) is identified as a downstream target of PFKFB4 and an PFKFB4-interacting protein. Moreover, bruceine A induces cell growth inhibition and apoptosis through GSK3β, which is dysregulated in pancreatic cancer and closely related to the prognosis. In all, these findings suggest that bruceine A inhibits human pancreatic cancer cell growth via PFKFB4/GSK3β-mediated glycolysis, and it may serve as a potent agent for curing human pancreatic cancer.

The blood brain barrier (BBB) maintains the basic stability of the brain tissue under physiological conditions, while destroys and exaggerates brain edema and inflammatory response after ischemic stroke. In this study, we researched S-oxiracetam (S-ORC), a nootropic drug, alleviates BBB dysfunction and protects against ischemic stroke in rats. Middle cerebral artery occlusion(MCAO)/reperfusion in rats is applied to mimic ischemic stroke. One hour after reperfusion, rats are administered intravenously with different dose (0.12, 0.24, or 0.48 g/kg) of S-ORC for continuative three days. Seventy-two hours after MCAO, TTC staining, hematoxylin and eosin (H&E) staining, brain water content, immunohistochemical staining, EB extravasation, western blot are provided to evaluate the protective effect and possible mechanism of S-ORC on BBB dysfunction. Furthermore, brain concentration of verapamil (P-glycoprotein substrate) and atenolol (paracellular transport marker) were assayed by UPLC-MS/MS co administration with or without S-ORC. The results show that post-treatment of S-ORC decreases cerebral infarct size, lessens brain edema, inhibits neutrophil infiltration and cytokines releasing. Furthermore, S-ORC treatment decreases EB leakage, downregulates MMP-9, upregulates occludin and claudin-5, and decreases brain concentration of verapamil and atenolol after MCAO surgery. In conclusion, the present study demonstrates that post-treatment of S-ORC alleviates BBB dysfunction by regulating tight junction proteins (TJPs), upregulating P-glycoprotein function, and protects against ischemic stroke as result.

The roots of Paeoniae lactiflora. Pall. are widely consumed as crude drugs in Asian countries due to their remarkable beneficial health effects. The present research was undertaken to illuminate the dynamic changes in metabolites and enzymes and facilitate selection of the harvesting time when the herb can provide optimum health benefits. P. lactiflora. roots were analyzed at 12 stages of growth for monoterpenoid glycosides, phenols, nucleosides, nucleobases, amino acids, and polysaccharides by high-performance liquid chromatography with photodiode array detector, ultra-high pressure liquid chromatography coupled with tandem mass spectrometry, and UV spectrophotometry. The enzyme activities of plant β-glucosidases and esterases were determined by UV methods. The total content of monoterpenoid glycosides and phenols peaked in December. For nucleosides and nucleobases, the highest content appeared in April. The maximum phasic accumulation of the total amino acids took place in March, and the content of total polysaccharides reached a peak value in September. December, April, and March were selected as the appropriate harvesting times for producing natural medicinal or health food products. Plant β-glucosidases and esterases showed the highest activity in December and May, respectively. When the activity of β-glucosidase increased, esterase activity decreased, while the contents of oxypaeoniflora and paeoniflorin increased. When esterase activity increased, the contents of benzoylpaeoniflorin, paeoniflorin, and gallic acid decreased. In conclusion, the results from the present study would be useful in determination of the suitable time for harvesting P. lactiflora. roots for medicinal purposes.

Delisheng injection (DLS), composed of venenum bufonis (VB), Chinese blister beetle (CB), red ginseng (RG) and Astragalus membranaceus (AM), is a Chinese medicine injection with good efficacy for the treatment of different types of cancers, while the side effects of DLS, such as vascular irritation and inflammation, restrict its wide clinical applications. Based on our previous study, VB is a main contributor for the vascular irritation of DLS. To reveal the possible mechanism involved in VB-induced vascular irritation is of practical importance. Moreover, to find a way to alleviate VB-induced vascular irritation is of great necessity. In our present study, venenum bufonis liposome (VBL) was prepared by thin film dispersion method to attenuate VB-induced vascular irritation. Vascular irritation of VB and its liposome were evaluated in a rabbit model. Rabbits were divided into three groups: (1) the control group (K); (2) the VB group (VB); (3) the VBL group (VBL). Animals administrated with VB or VBL 0.12 mg kg−1 were sacrificed at 6 h. Inflammatory cytokines and TLR/MAPK/NF-κB pathway-related proteins were examined, respectively. Our results showed that VB significantly increased serum proinflammatory cytokines, including interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α), while it led to decreased anti-inflammatory cytokines interleukin-10 (IL-10) after intravenous administration. VB-induced vascular irritation was further confirmed by histopathological evaluation. Moreover, the results obtained from western blot and immunohistochemical analysis revealed that VB activated TLR/MAPK/NF-κB signaling pathway involving TLR2, TLR4, MyD88, p-Erk, p-JNK, p-P38, p-IκBα and p-NF-κB p65 protein. VB liposome decreased the VB-induced overexpression of TLR2, TLR4, MyD88, p-Erk, p-JNK, p-P38, p-IκBα and p-NF-κB p65. The anticancer activity of VB and VBL was also evaluated, and VBL still showed anticancer activity both in vivo and in vitro. Taken together, our finding demonstrated that VB induced vascular irritation was related to the regulation of the TLR/MAPK/NF-κB signaling pathway, and liposome encapsulation significantly attenuated VB-induced vascular irritation while maintaining its anticancer activity. The results of this study have important practical significance in guiding the safety clinical application of Chinese medicine injection.

A highly sensitive method using ultra-high-pressure liquid chromatography coupled with linear ion trap-Orbitrap tandem mass spectrometry (UHPLC-LTQ-Orbitrap-MS) has been developed and validated for the simultaneous identification and quantification of ginkgolic acids and semi-quantification of their metabolites in rat plasma. For the five selected ginkgolic acids, the method was found to be with good linearities (r0.9991), good intra- and inter-day precisions (RSD15%), and good accuracies (RE, from -10.33% to 4.92%) as well. Extraction recoveries, matrix effects and stabilities for rat plasm samples were within the required limits. The validated method was successfully applied to investigate the pharmacokinetics of the five ginkgolic acids in rat plasma after oral administration of 3 dosage groups (900mg/kg, 300mg/kg and 100mg/kg). Meanwhile, six metabolites of GA (15:1) and GA (17:1) were identified by comparison of MS data with reported values. The results of validation in terms of linear ranges, precisions and stabilities were established for semi-quantification of metabolites. The curves of relative changes of these metabolites during the metabolic process were constructed by plotting the peak area ratios of metabolites to salicylic acid (internal standard, IS), respectively. Double peaks were observed in all 3 dose groups. Different type of metabolites and different dosage of each metabolite both resulted in different T

Frankincense and myrrha (FM), commonly used as a classical herbal pair, have a wide range of clinical applications and definite anti-inflammatory activity. However, anti-neuroinflammation effects and mechanisms are not clear. In this study, we adopted a lipopolysaccharide (LPS)-induced microglial (BV2) cell model and a network pharmacology method to reveal the anti-neuroinflammatory effects and mechanisms of boswellic acid (BA) and myrrha sesquiterpenes (MS) with different proportions of compatibility. The data showed that the different ratios of BA and MS had different degrees of inhibition of interleukin-1&beta, (IL-1&beta, ), IL-6, and inducible nitric oxide synthase (iNOS) mRNA expression, down-regulated the phosphor-nuclear factor kappa B/nuclear factor kappa B (p-NF-ҡB)/(NF-ҡB), phosphorylated protein kinase b/protein kinase b (p-AKT/AKT), and Toll-like receptor 4 (TLR4) protein expression levels, and increased phospho-PI3 kinase (p-PI3K) protein expression levels. When the ratios of BA and MS were 10:1, 5:1, and 20:1, better effective efficacy was exhibited. According to the correlation analysis between the effect index and bioactive substances, it was suggested that 2-methoxy-5-acetoxy -fruranogermacr-1(10)-en-6-one (Compound 1), 3&alpha, acetyloxylanosta-8,24-dien-21-oic acid (Compound 2), 11-keto-boswellic acid (Compound 3), and 3-acetyl-11-keto-&beta, boswellic acid (Compound 4) made important contributions to the treatment of neuroinflammation. Furthermore, based on the network pharmacological analysis, it was found that these four active compounds acted on 31 targets related to neuroinflammation and were involved in 32 signaling pathways which mainly related to the immune system, cardiovascular system, and nervous system, suggesting that BA and MS could be used to treat neuroinflammation.

The spontaneous db/db mice were used to elucidate the biological effects and mechanisms of Rehmannia glutinosa leaves total glycoside (DHY) on kidney injury through biochemical indicators, kidney pathological section analysis, metabolic profiling, intestinal flora analysis and in vitro Human renal tubular epithelial (HK-2) cell model induced by high glucose. It was found that DHY can decrease the blood sugar level (insulin, INS; fasting blood glucose, FBG), blood lipid level (Total Cholesterol, T-CHO; Triglyceride, TG) significantly and improve kidney injury level (blood urea nitrogen, BUN; urine microalbumin, mALB; serum creatinine, Scr). It can also alleviate kidney tubular epithelial cell oedema and reduce interstitial connective tissue hyperplasia of the injury kidney induced by high glucose. 13 endogenous metabolites were identified in serum, which involved of ether lipid metabolism, sphingolipid metabolism, glyoxylic acid and dicarboxylic acid metabolism and arachidonic acid metabolism. High glucose can also lead to the disorder of intestinal flora, especially Firmicutes and Bacteroides. Meanwhile, DHY also inhibited the expression of α-SMA, TGF- β1, Smad3 and Smad4 in the kidney tissues of db/db mice and HK-2 cells. To sum up, DHY may restore the dysfunctional intestinal flora to normal and regulate glycolipid level of db/db mice as well as TGF-β/Smad signalling pathway regulation to improve early kidney damage caused by diabetes.

Scutellariae radix (Scutellaria baicalensis Georgi, SR) and coptidis rhizoma (Coptis chinensis Franch, CR) are both widely used traditional Chinese medicines and have been used together to treat T2DM with synergistic effects in the clinical practices for thousands of years, but their combination mechanism is not clear. Accumulating evidences have implicated gut microbiota as important targets for the therapy of T2DM. Thus, this study aimed to unravel the cooperation mechanism of SR and CR on the amelioration of T2DM based on the systematic analysis of metagenome and metabolome of gut microbiota. Bacterial communities were analyzed based on high-throughput 16S rRNA gene sequencing. Furthermore, ultra high-performance liquid chromatography coupled with quadrupole time of flight mass spectrometry (UPLC-Q-TOF-MS) was used to analyze variations of microbial metabolites in feces and the contents of short chain fatty acids (SCFAs) in the cecum were determined by a gaschromatography-flame ionization detector (GC-FID). 16S rRNA gene sequencing results revealed that T2DM rats treated with SR, CR, and the combination of SR and CR (SC) exhibited changes in the composition of the gut microbiota. The SCFAs-producing bacteria such as Bacteroidales S24-7 group_norank, [Eubacterium] nodatum group, Parasutterella, Prevotellaceae UCG-001, Ruminiclostridium, and Ruminiclostridium 9 in T2DM rats were notably enriched after treatment with SR, CR, and their combination. In contrast, secondary bile acid-producing bacteria such as Escherichia-Shigella strongly decreased in numbers. The perturbance of metabolic profiling in T2DM rats was obviously improved after treatment, exhibiting a lower level of secondary bile acids and a numerical increase of microbially derived SCFAs. Moreover, the correlation analysis illustrated a close relationship among gut microbiota, its metabolites, and T2DM-related indexes. The findings indicated that the crosstalk between microbiota-derived metabolites and the host played an important role in the progress of T2DM and might provide a novel insight regarding gut microbiota and its metabolites as potential new targets of traditional Chinese medicines. Furthermore, this work also suggested that the integration of various omics methods and bioinformatics made a useful template for drug mechanism research.

Trichosanthes kirilowii Maxim. is one of the original plants for traditional Chinese medicines Trichosanthis Fructus, Trichosanthis Semen, Trichosanthis Pericarpium and Trichosanthis Radix. Amino acids, nucleosides and carbohydrates are usually considered to have nutritional value and health-care efficacy. In this study, methods involving high-performance liquid chromatography coupled with evaporative light scattering detector (HPLC-ELSD), UV-visible spectrophotometry and ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS) were established for quantifying carbohydrates (fructose, glucose, stachyose, raffinose and polysaccharide), fourteen nucleosides and twenty one amino acids. Moreover, sixty-three samples from nine different parts, including pericarp, seed, fruit pulp, stem, leaf, main root, main root bark, lateral root and lateral root bark of T. kirilowii from different cultivated varieties were examined. The established methods were validated with good linearity, precision, repeatability, stability, and recovery. The results showed that the average content of total amino acids in roots (15.39 mg/g) and root barks (16.38 mg/g) were relatively higher than for others. Contents of nucleosides in all parts of T. kirilowii were below 1.5 mg/g. For carbohydrates, fruit pulp has a higher content than others for glucose (22.91%), fructose (20.63%) and polysaccharides (27.29%). By using partial least-squared discriminate analysis (PLS-DA), Variable importance in the projection (VIP) plots and analysis of variance (ANOVA) analysis, the characteristic components of the different organs (fruit, stems and leaves, roots) were found. This analysis suggested there were potential medicinal and nutritive health care values in various parts of the T. kirilowii, which provided valuable information for the development and utilization of T. kirilowii.

The ambiguity of dose-effect relationship of many traditional Chinese medicines (TCMs) has always influenced their rational use in TCM clinic. Rhubarb, a preferred representative of cathartic TCM, is currently widely used that results in a diversity of its dosage. The aim of this study was to use an integrated metabolomics strategy to simultaneously reveal dose-effect relationship and therapeutic mechanisms of different efficacy of rhubarb in constipation rats. Six doses of rhubarb (0.135, 0.27, 0.81, 1.35, 4.05, and 8.1 g/kg) were examined to elucidate the laxative and fire-purging effects by pathological sections and UPLC-Q-TOF/MSE. The results showed that there existed serious lesions in the stomach and colon of model rats. And conditions were basically improved to some extent in rhubarb-treated groups. Through relative distance calculation based on metabolomics score plots, it suggested that the effective dose threshold (EC20–EC80 range) of rhubarb was from 0.31 to 4.5 g/kg (corresponding to 3.44–50.00 g in the clinic) in rat serum and 0.29–2.1 g/kg (corresponding to 3.22–23.33 g in the clinic) in feces. Then, 33 potential biomarkers were identified in total. Functional pathway analysis revealed that the alterations of these biomarkers were associated with 15 metabolic pathways, mainly including arachidonic acid metabolism, glycerophospholipid metabolism, steroid biosynthesis, primary bile acid biosynthesis and sphingolipid metabolism. Of note, different doses of rhubarb could alleviate endogenous disorders to varying degrees through regulating multiple perturbed pathways to the normal state, which might be in a dose-dependent manner and involved in therapeutic mechanisms. To sum up, integrated serum and fecal metabolomics obtained that rhubarb ranging from 0.31 to 2.1 g/kg is safe and effective for constipation treatment. Also, our findings showed that the robust metabolomics techniques would be promising to be more accurately used in the dose-effect studies of complex TCM, and to clarify syndrome pathogenesis and action mechanisms in Chinese medicine.

Epilepsy is a common clinical syndrome characterized by sudden and recurrent attacks and temporary central nervous system dysfunction caused by excessive discharge of neurons in the brain. Amber, a fossilized organic substance formed by the resins of conifers and leguminous plants, was prescribed to tranquilize the mind in China. In this paper, the antiepileptic effect of amber was evaluated by a pentylenetetrazole (PTZ)-induced epileptic model. An untargeted metabolomics approach was applied to investigate metabolic changes in the epileptic model, which was based on HILIC-UHPLC-MS/MS multivariate statistical analysis and metabolism network analysis. The outcome of this study suggested that 35 endogenous metabolites showed marked perturbations. Moreover, four metabolism pathways were mainly involved in epilepsy. After treatment by amber, the endogenous metabolites had a marked tendency to revert back to the situation of the control group which was consistent with phenobarbital. This study characterized the pentylenetetrazole-induced epileptic model and provided new evidence for the sedative effect of amber.

Xiexin Decoction (XXD), a traditional Chinese medicine prescription composed of Rhei rhizome (RR), Scutellaria radix (SR) and Coptidis rhizome (CR), has been used to cure diabetes in clinical practices for thousands of years, but its mechanism is not clear. Our previous study indicated that XXD could significantly ameliorate the symptom of type 2 diabetes mellitus (T2DM) rats by shifting the composition of gut microbiota. However, the effect of XXD on the metabolic activity of gut microbiota is not clarified. In this study, the underlying mechanism of XXD on the amelioration of T2DM was explored by fecal metabolic profiling analysis based on ultra performance liquid chromatography coupled with quadrupole time-of-fight mass spectrometry (UPLC-Q-TOF/MS). The disordered metabolic profiles in T2DM rats were notably improved by XXD. Ten potential biomarkers, which were mainly involved in arachidonic acid metabolism, amino acid metabolism, bile acid metabolism, glycolysis and gluconeogenesis, were identified. Furthermore, these metabolites were closely related to SCFAs-producing and anti-inflammatory gut microflora. After XXD intervention, these biomarkers restored to the normal level at some extent. This study not only revealed potential biomarkers and related pathways in T2DM rats affected by XXD, but also provided a novel insight to uncover how traditional herb medicines worked from fecal metabolomics.

Ethnopharmacological relevance Cough variant asthma (CVA) is characterized with its long-lasting cough symptom on clinic. The mechanism of CVA is related to chronic persistent airway inflammation, airway hyperresponsiveness, etc. The traditional Chinese prescription has achieved good curative effect on CVA treatment through reducing cough counts, decreasing airway hyperresponsiveness and alleviating airway inflammation. The mechanism is associated with reducing IL4, IL-13, NGF and CGRP levels, as well as down-regulating TRPA1/TRPV1/TRPV5 channels in both lung and brain tissues. Aim of the study The Chinese prescription, San'ao decoction with scorpio and bombyx batryticatus (SSB), is well known in treating cough in asthmatic patients. In this study, the anti-tussive and anti-asthmatic role of SSB, as well as its mechanism on CVA mice model were explored and evaluated via alleviating airway inflammation and regulation of TRP channels. Materials and methods The major chemical components in SSB were detected and analyzed by UPLC-QTOF-MS under an optimized chromatographic and MS condition. 60 BALB/c mice were randomly divided into six groups: normal group, model group, dexamethasone group (0.1178 mg/kg/d), SSB high dose group (9.74 g/kg/d), SSB middle dose group (4.87 g/kg/d) and SSB low dose group (2.435 g/kg/d). The cough variant asthma mice model was established by ovalbumin sensitization and challenge. The protective role of SSB on CVA mice model was studied through inducing cough counts by capsaicin, assessing inflammatory cells in peripheral blood and bronchoalveolar lavage fluid (BALF), measuring airway responsiveness, detecting histopathological changes in lung tissues, analyzing cytokines and neuropeptides levels in BALF, as well as examining the mRNA and protein expressions of TRPA1, TRPV1 and TRPV5 in both lung and brain tissues. Results 17 signal peaks of the chemical components in SSB were identified by using UPLC-QTOF-MS. SSB (especially the high dose and middle dose), showed significantly effects on mice model by reducing mice cough counts (P ＜ 0.01), decreasing eosinophil (EOS) counts in blood (P ＜ 0.01) and inflammatory cell numbers in BALF (P ＜ 0.01), decreasing airway hyperresponsiveness (P ＜ 0.05), reducing the levels of IL-4 (P ＜ 0.05), IL-13 (P ＜ 0.01), NGF (P ＜ 0.01) and CGRP (P ＜ 0.01) in BALF, as well as down regulating the mRNA and protein expressions of TRPA1, TRPV1 and TRPV5 in both lung and brain tissues (P ＜ 0.01). Conclusions SSB showed anti-tussive and anti-asthmatic effects on cough variant asthma mice model by reducing cough counts, improving lung function, alleviating lung injury and airway inflammation. The mechanism of SSB might be associated with the regulation of cytokines and neuropeptides in BALF, as well as the regulation of TRPA1, TRPV1, TRPV5 channels in both lung and brain tissues.

The drug combination of Danhong injection (DHI) and low-dose aspirin (ASA) was frequently applied for the treatment of cardiovascular and cerebrovascular diseases. Due to the drug interactions, a lot of potential benefits and risks might exist side by side in the course of combination therapy. However, there had been no studies of interaction between DHI and ASA. Metabolomics was a powerful tool to explore endogenous biomarkers and metabolic pathways. In present study, metabolic profiling with ultra-high-performance liquid chromatography coupled to quadrupole time of flight mass spectrometry (UHPLC-QTOF/MS) coupled with multivariate statistical analysis was performed to provide insight into understanding the interaction between DHI and low-dose ASA. Eleven potential biomarkers of three types were identified and seven metabolic pathways were constructed. The results showed that the interaction between DHI and low-dose ASA during synergistic treatment indeed affected some key endogenous biomarkers and metabolic pathways, which could not happen when DHI or low-dose ASA was used alone. The quality and quantity of endogenous metabolite were both influenced by interaction between DHI and low-dose ASA. In details, the amount of flavin mononucleotide, L-2, 4-diaminobutyric acid (DABA) and 4-aminohippuric acid were significantly increased. On the contrary, the amount of 3-methyluridine, 4, 6-dihydroxyquinoline, cortolone-3-glucuronide, and serotonin were significantly decreased. Furthermore, O-phosphotyrosine, 3-methyl-2-butenal, indoxyl sulfate and dolichyl diphosphate were disappeared in urine. As to metabolic pathways, riboflavin metabolism, pentose and glucuronate interconversions, and tryptophan metabolism were all significantly influenced. The emerging alterations of biomarkers and metabolic pathways were associated with a lot of drugs and diseases based on literature researches, which might influence the co-administration of other drugs or the treatments of relevant diseases. Our paper presented some hints to uncover the mechanism of interaction between DHI and low-dose ASA, which would provide some references for application of DHI and low-dose ASA combination.

This research was designed to study metabonomic characteristics of the toxicity induced by yuanhuapine, a major bioactive diterpenoid in a well-known traditional Chinese medicine-Genkwa Flos. General observation, blood biochemistry and histopathological examination were used to reflect yuanhuapine-induced toxicity. Urine samples from rats in control and yuanhuapine treated rats were analyzed by ultra-performance liquid chromatography tandem quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS). Pattern recognition methods including principal components analysis (PCA), partial least-squared discriminant analysis (PLS-DA), orthogonal partial least-squared discriminant analysis (OPLS-DA) and computational system analysis were integrated to obtain comprehensive metabonomic profiling and pathways of the biological data sets. The results suggested that yuanhuapine could induce intestinal and liver damage. And 14 endogenous metabolites as biomarkers related to the amino acids metabolism, lipids metabolism, carbohydrate metabolism and gut microflora were significantly changed in the urine of yuanhuapine treated rats, which were firstly constructed the metabolomic feature profiling and metabolite interaction network of yuanhuapine-induced injury using pattern recognition methods and Ingenuity Pathway Analysis (IPA) approach. The present study showed that yuanhuapine-induced intestinal and hepatic toxicity were correlated with disturbance of amino acids metabolism, lipids metabolism, carbohydrate metabolism and gut microflora.

A simple, sensitive and fast method for the simultaneous quantitation of 15 flavonoids in licorice based on an ultra high performance liquid chromatography coupled with triple quadrupole electrospray tandem mass spectrometry had been established and validated in this study. The analysis was performed on an ACQUITY HSS T3 column with gradient elution using a mobile phase consisted of A (0.1% formic acid in water)/B (acetonitrile) at a flow rate of 0.4 mL/min. Satisfactory separation of these compounds was obtained in less than 9 min. All calibration curves showed good linearity (r(2) = 0.9940) during the test ranges. The precision, repeatability, accuracy, limit of detection and limit of quantification were also fully investigated. The validated method was successfully applied for the simultaneous quantitation of 15 flavonoids in 106 licorice samples which contained 83 batches of G. uralensis, 14 batches of G. glabra and 9 batches of G. inflata. Furthermore, multivariate statistical analysis (using principal components analysis) was performed to classify the samples based on the contents of the 15 analyzed compounds. The results showed that all of these licorice samples were rich in flavonoids, although their contents were obviously various, and the proposed method could serve as a prerequisite for quality control of licorice products.