Integration of untargeted and pseudotargeted metabolomics reveals specific markers for authentication and adulteration detection of Fritillariae Bulbus using tandem mass spectrometry and chemometrics.

Authentication and adulteration detection of closely related herbal medicines is a thorny issue in the quality control and market standardization of traditional Chinese medicine. Taking Fritillariae Bulbus (FB) as a case study, we herein proposed a three-step strategy that integrates mass spectrometry-based metabolomics and multivariate statistical analysis to identify specific markers, thereby accurately identifying FBs and determining the adulteration level. First, an ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry-based untargeted metabolomics method was employed to profile steroid alkaloids in five sorts of FB and screen potential differential markers. Then, the reliability of the screened markers was further verified by the distribution in different FB groups acquired from ultra-high performance liquid chromatography triple quadrupole mass spectrometry-based pseudotargeted metabolomics analysis. As a result, a total of 16 compounds were screened out to be the specific markers, which were successfully applied to distinguish five FBs by using discriminant analysis model. Besides, partial least squares regression models based on specific markers allowed accurate prediction of three sets of adulterated FBs. All the models afforded good linearity and good predictive ability with regression coefficient of prediction (R2p) > 0.9 and root mean square error of prediction (RMSEP) < 0.1. The reliable results of discriminant and quantitative analysis revealed that this proposed strategy could be potentially used to identify specific markers, which contributes to rapid chemical discrimination and adulteration detection of herbal medicines with close genetic relationship.

Quality Consistency Evaluation of Traditional Chinese Medicines: Current Status and Future Perspectives.

Quality consistency evaluation of traditional Chinese medicines (TCMs) is a crucial factor that determines the safe and effective application in clinical settings. However, TCMs exhibit diverse, heterogeneous, complex, and flexible chemical compositions, as well as variability in preparation processes. These characteristics pose greater challenges in researching the consistency of TCMs compared to chemically synthesized and biological drugs. Therefore, it is paramount to develop effective strategies for evaluating the quality consistency of TCMs. From the starting point of quality properties, this review explores the strategy used to evaluate quality consistency in terms of chemistry-based strategy (chemical consistency) and the biology-based strategy (bioequivalence). Among them, the chemistry-based strategy is the mainstream, and biology-based strategy complements the chemistry-based strategy each other. Furthermore, the emerging chemistry-biology strategies (overall evaluation) is discussed, including individually combining strategy and integration strategy. Finally, this review provides insights into the challenges and future perspectives in this field. By highlighting current status and trends in TCMs quality consistency, this review aims to contribute to establishment of generally applicable chemistry-biology integrated evaluation strategy for TCMs. This will facilitate the advancement toward a higher stage of overall quality evaluation.

Responses of biomass accumulation and nutrient utilization along a phosphorus supply gradient in Leymus chinensis.

Phosphorus (P) deficiencies are widespread in calcareous soils. The poor availability of nitrogen (N) and P in soils often restricts crop growth. However, the effects of P addition on plant growth and plant nutrient transport changes during the establishment of Leymus chinensis fields in Xinjiang are not clear. We investigated the responses of Leymus chinensis biomass and nutrient absorption and utilization to changes in soil N and P by adding P (0, 15.3, 30.6, and 45.9 kg P ha-1 year-1) with basally applied N fertilizer (150 kg N ha-1 year-1). The results showed that (a) Principal component analysis (PCA) of biomass, nutrient accumulation, soil available P, and soil available N during the different periods of Leymus chinensis growth showed that their cumulative contributions during the jointing and harvest periods reached 95.4% and 88%, respectively. (b) Phosphorus use efficiency (PUE) increased with the increase of P fertilizer gradient and then decreased and the maximum PUE was 13.14% under moderate P addition. The accumulation of biomass and nutrients in Leymus chinensis can be effectively improved by the addition of P fertilizer at 30.6 kg ha-1. Different P additions either moderately promoted or excessively inhibited Leymus chinensis growth and nutrient utilization.

An integrated strategy combining network toxicology and feature-based molecular networking for exploring hepatotoxic constituents and mechanism of Epimedii Folium-induced hepatotoxicity in vitro.

Epimedii Folium (EF), a commonly used herbal medicine to treat osteoporosis, has caused serious concern due to potential hepatotoxicity. Until now, its intrinsic hepatotoxic mechanism and hepatotoxic ingredients remain unclear. Here, a novel high-throughput approach was designed to investigate the intrinsic hepatotoxic of EF. High-content screen imaging (HCS) and biochemical tests were first performed to obtain the cytotoxicity parameter matrix of 17 batch EF samples. EF-treated alpha mouse liver 12 (AML12) cells showed increased reactive oxygen species (ROS), reduced glutathione (GSH) and mitochondrial membrane potential (MMP), and apoptosis and cholestasis were further observed. Network toxicology predicted that EF-triggered hepatotoxiciy was involved in transcription factor (TF) activity. The FXR expression, screened by a TF PCR array, exhibited down-regulation following EF extract administration. Moreover, EF inhibited bile acid (BA) metabolism pathway in an FXR-dependent manner. Pearson correlation between the cytotoxicity parameter matrix and quantification feature table obtained from UHPLC-QTOF data of EF suggested 7 prenylated flavonoids possessed potent hepatotoxicities and their cytotoxicity order was further summarized. The transcriptional repression effects of them on FXR were also verified. Collectively, our findings indicate that FXR is probably responsible for EF-induced hepatotoxicity and prenylated flavonoids may be a major class of hepatotoxic constituents in EF.

Screening the effective components in treating dampness stagnancy due to spleen deficiency syndrome and elucidating the potential mechanism of Poria water extract.

Poria is an important medicine for inducing diuresis to drain dampness from the middle energizer. However, the specific effective components and the potential mechanism of Poria remain largely unknown. To identify the effective components and the mechanism of Poria water extract (PWE) to treat dampness stagnancy due to spleen deficiency syndrome (DSSD), a rat model of DSSD was established through weight-loaded forced swimming, intragastric ice-water stimulation, humid living environment, and alternate-day fasting for 21 days. After 14 days of treatment with PWE, the results indicated that PWE increased fecal moisture percentage, urine output, D-xylose level and weight; amylase, albumin, and total protein levels; and the swimming time of rats with DSSD to different extents. Eleven highly related components were screened out using the spectrum-effect relationship and LC-MS. Mechanistic studies revealed that PWE significantly increased the expression of serum motilin (MTL), gastrin (GAS), ADCY5/6, p-PKAα/ß/γ cat, and phosphorylated cAMP-response element binding protein in the stomach, and AQP3 expression in the colon. Moreover, it decreased the levels of serum ADH, the expression of AQP3 and AQP4 in the stomach, AQP1 and AQP3 in the duodenum, and AQP4 in the colon. PWE induced diuresis to drain dampness in rats with DSSD. Eleven main effective components were identified in PWE. They exerted therapeutic effect by regulating the AC-cAMP-AQP signaling pathway in the stomach, MTL and GAS levels in the serum, AQP1 and AQP3 expression in the duodenum, and AQP3 and AQP4 expression in the colon.

Revealing active components and action mechanism of Fritillariae Bulbus against non-small cell lung cancer through spectrum-effect relationship and proteomics.

BACKGROUND: Fritillariae Bulbus (FB) is widely used as a traditional medicine for the treatment of lung meridian diseases. It has been proved that FB has good anti-non-small cell lung cancer (NSCLC) activity. However, the active components and potential mechanism are still not clear. PURPOSE: To reveal the bioactive components of FB against NSCLC and potential mechanism through spectrum-effect relationship and proteomics. METHOD: First, the FB extract was chemically profiled by UHPLC-QTOF-MS and the inhibitory effect of FB extract on A549 cell viability was evaluated by Cell Counting Kit-8 assay. Second, orthogonal-partial least squares-regression analysis was applied to screen potential active compounds through correlating the chemical profile with corresponding inhibitory effect. Third, the anti-NSCLC activities of potential active components were further investigated in terms of cell proliferation, cell cycle and cell apoptosis in vitro and tumor growth in vivo. Finally, proteomics was utilized to reveal the underlying anti-NSCLC mechanism. RESULTS: Six potential active components including verticine, verticinone, zhebeirine, ebeiedinone, yibeissine and peimisine were screened out by spectrum-effect relationship. Among them, zhebeirine showed higher inhibitory effect on A549 cell viability with IC50 value of 36.93 µM and dosage-dependent inhibition of A549 xenograft tumor growth in nude mice. Proteomics and western blotting assays indicated that zhebeirine could arrest cell cycle by down-regulating the expressions of CDK1, CDK2, Cyclin A2, Cyclin B2 and inhibiting the phosphorylation of p53. Moreover, the proteins participating in p53 signaling pathway including PCNA, 14-3-3σ, CHEK1 were significantly decreased, which suggested that zhebeirine affected cell cycle progression through p53 signaling pathway. CONCLUSION: This study not only provides scientific evidence to support the clinical application of FB against NSCLC, but also demonstrates that zhebeirine is a promising anti-NSCLC lead compound deserving further studies.

Transcriptomics and metabolomics reveal the role of CYP1A2 in psoralen/isopsoralen-induced metabolic activation and hepatotoxicity.

Psoralen and isopsoralen are the pharmacologically important but hepatotoxic components in Psoraleae Fructus. The purpose of this study was to reveal the underlying mechanism of psoralen/isopsoralen-induced hepatotoxicity. Initially, we applied integrated analyses of transcriptomic and metabolomic profiles in mice treated with psoralen and isopsoralen, highlighting the xenobiotic metabolism by cytochromes P450 as a potential pathway. Then, with verifications of expression levels by qRT-PCR and western blot, affinities by molecular docking, and metabolic contributions by recombinant human CYP450 and mouse liver microsomes, CYP1A2 was screened out as the key metabolic enzyme. Afterwards, CYP1A2 induction and inhibition models in HepG2 cells and mice were established to verify the role of CYP1A2, demonstrating that induction of CYP1A2 aggravated the hepatotoxicity, and conversely inhibition alleviated the hepatotoxic effects. Additionally, we detected glutathione adducts with reactive intermediates of psoralen and isopsoralen generated by CYP1A2 metabolism in biosystems of recombinant human CYP1A2 and mouse liver microsomes, CYP1A2-overexpressed HepG2 cells, mice livers and the chemical reaction system using UPLC-Q-TOF-MS/MS. Ultimately, the high-content screening presented the cellular oxidative stress and relevant hepatotoxicity due to glutathione depletion by reactive intermediates. In brief, our findings illustrated that CYP1A2-mediated metabolic activation is responsible for the psoralen/isopsoralen-induced hepatotoxicity.

Baohuoside I inhibits FXR signaling pathway to interfere with bile acid homeostasis via targeting ER α degradation.

Epimedii folium (EF) is an effective herbal medicine in osteoporosis treatment, but the clinical utilization of EF has been limited due to potential hepatotoxicity. The previous studies identified that baohuoside I (BI), the main active component of EF, was relevant to EF-induced liver injury. However, the mechanisms of BI causing direct injury to hepatocytes remain unclear. Here, we reveal that BI inhibits FXR-mediated signaling pathway via targeting estrogen receptor α (ER α), leading to the accumulation of bile acids (BAs). Targeted bile acid analyses show BI alters the BA composition and distribution, resulting in impaired BA homeostasis. Mechanistically, BI induces FXR-dependent hepatotoxicity at transcriptional level. Additionally, ER α is predicted to bind to the FXR promoter region based on transcription factor binding sites databases and we further demonstrate that ER α positively regulates FXR promoter activity and affects the expression of target genes involved in BA metabolism. Importantly, we discover that ER α and its mediated FXR transcription regulation might be involved in BI-induced liver injury via ligand-dependent ER α degradation. Collectively, our findings indicate that FXR is a newly discovered target gene of ER α mediated BI-induced liver injury, and suggest BI may be responsible for EF-induced liver injury.

A Review of The Application of Spectroscopy to Flavonoids from Medicine and Food Homology Materials.

Medicinal and food homology materials are a group of drugs in herbal medicine that have nutritional value and can be used as functional food, with great potential for development and application. Flavonoids are one of the major groups of components in pharmaceutical and food materials that have been found to possess a variety of biological activities and pharmacological effects. More and more analytical techniques are being used in the study of flavonoid components of medicinal and food homology materials. Compared to traditional analytical methods, spectroscopic analysis has the advantages of being rapid, economical and free of chemical waste. It is therefore widely used for the identification and analysis of herbal components. This paper reviews the application of spectroscopic techniques in the study of flavonoid components in medicinal and food homology materials, including structure determination, content determination, quality identification, interaction studies, and the corresponding chemometrics. This review may provide some reference and assistance for future studies on the flavonoid composition of other medicinal and food homology materials.

The development and validation of a predictive model for neonatal phototherapy outcome using admission indicators.

Delayed exchange transfusion therapy (ETT) after phototherapy failure for newborns with severe hyperbilirubinemia could lead to serious complications such as bilirubin encephalopathy (BE). In this current manuscript we developed and validated a model using admission data for early prediction of phototherapy failure. We retrospectively examined the medical records of 292 newborns with severe hyperbilirubinemia as the training cohort and another 52 neonates as the validation cohort. Logistic regression modeling was employed to create a predictive model with seven significant admission indicators, i.e., age, past medical history, presence of hemolysis, hemoglobin, neutrophil proportion, albumin (ALB), and total serum bilirubin (TSB). To validate the model, two other models with conventional indicators were created, one incorporating the admission indicators and phototherapy failure outcome and the other using TSB decrease after phototherapy failure as a variable and phototherapy outcome as an outcome indicator. The area under the curve (AUC) of the predictive model was 0.958 [95% confidence interval (CI): 0.924-0.993] and 0.961 (95% CI: 0.914-1.000) in the training and validation cohorts, respectively. Compared with the conventional models, the new model had better predictive power and greater value for clinical decision-making by providing a possibly earlier and more accurate prediction of phototherapy failure. More rapid clinical decision-making and interventions may potentially minimize occurrence of serious complications of severe neonatal hyperbilirubinemia.

Physiological and protein profiling analysis provides insight into the underlying molecular mechanism of potato tuber development regulated by jasmonic acid in vitro.

BACKGROUND: Jasmonates (JAs) are one of important phytohormones regulating potato tuber development. It is a complex process and the underlying molecular mechanism regulating tuber development by JAs is still limited. This study attempted to illuminate it through the potential proteomic dynamics information about tuber development in vitro regulated by exogenous JA. RESULTS: A combined analysis of physiological and iTRAQ (isobaric tags for relative and absolute quantification)-based proteomic approach was performed in tuber development in vitro under exogenous JA treatments (0, 0.5, 5 and 50 µΜ). Physiological results indicated that low JA concentration (especially 5 µM) promoted tuber development, whereas higher JA concentration (50 µM) showed inhibition effect. A total of 257 differentially expressed proteins (DEPs) were identified by iTRAQ, which provided a comprehensive overview on the functional protein profile changes of tuber development regulated by JA. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis indicated that low JA concentration (especially 5 µM) exhibited the promotion effects on tuber development in various cellular processes. Some cell wall polysaccharide synthesis and cytoskeleton formation-related proteins were up-regulated by JA to promote tuber cell expansion. Some primary carbon metabolism-related enzymes were up-regulated by JA to provide sufficient metabolism intermediates and energy for tuber development. And, a large number of protein biosynthesis, degradation and assembly-related were up-regulated by JA to promote tuber protein biosynthesis and maintain strict protein quality control during tuber development. CONCLUSIONS: This study is the first to integrate physiological and proteomic data to provide useful information about the JA-signaling response mechanism of potato tuber development in vitro. The results revealed that the levels of a number of proteins involved in various cellular processes were regulated by JA during tuber development. The proposed hypothetical model would explain the interaction of these DEPs that associated with tuber development in vitro regulated by JA.

Comprehensive chemical profiling of volatile constituents of Angong Niuhuang Pill in vitro and in vivo based on gas chromatography coupled with mass spectrometry.

BACKGROUND: Angong Niuhuang Pill (ANP), a renowned precious traditional Chinese medicine prescription, is extensively utilized for the clinical treatment of stroke, meningitis and encephalorrhagia in China. As a classic resuscitation-inducing aromatic prescription, ANP has been investigated for its pharmacological effects in recent years, while the volatile composition in ANP still lacks comprehensive elucidation. METHOD: To better explore the volatile constituents in ANP, a qualitative analysis method was developed based on gas chromatography coupled with mass spectrometry. Furthermore, a validated quantitative method was established to determine 21 main compounds in 8 batches of commercially available ANP samples by gas chromatography-tandem mass spectrometry. The quantitative data were successively subjected to Pearson correlation coefficient analysis. Additionally, the absorbed volatile constituents in rat plasma after single oral administration of ANP have also been characterized. RESULTS: A total of 93 volatile constituents including 29 sesquiterpenoids, 28 monoterpenoids, 13 fatty acids and their esters, 7 alkanes, 6 ketones, 3 phenols, 3 aldehydes, 2 benzoate esters, and 2 other types, were preliminarily characterized, which primarily originated from Borneolum, Moschus, Curcumae Radix, and Gardeniae Fructus. D-Borneol, isoborneol and muscone were the top three abundant ingredients (> 600 µg/g) in 8 batches of ANP samples. Subsequently, the average Pearson correlation coefficient of the contents of 21 analytes was 0.993, inferring the high batch-to-batch similarity among 8 batches. After oral administration of ANP, D-borneol, isoborneol, muscone and camphor were the main volatile constituents absorbed in the rat plasma. CONCLUSION: This research may be helpful for the comprehensive quality control study of ANP, and provide for guarantee the clinical efficacy of ANP.

Integrated 16S rDNA, metabolomics, and TNF-α/NF-κB signaling pathway analyses to explain the modulatory effect of Poria cocos aqueous extract on anxiety-like behavior.

BACKGROUND: Poria cocos is an ancient medicine and modern functional food, which exerts excellent effects on anxiety, although its mechanism is unknown. PURPOSE: To explore the mechanisms of the aqueous extract of P. cocos (PCD) in ameliorating anxiety-like behavior caused by chronic sleep deprivation (CSD). METHODS: PCD chemical composition was analyzed by UPLC-QTOF-MS/MS. A CSD rat model was established over 21 days. We examined the effects and mechanisms after 10 days of CSD using open-field tests (OFTs), enzyme-linked immunosorbent assays, 16S rDNA, non-targeted metabolomics, and Western blot analyses. RESULTS: Sixty-two triterpenoids were identified in PCD. CSD-induced anxiety-like behavior was significantly attenuated by PCD treatment. PCD improved hypothalamic neurotransmitters, decreased proinflammatory cytokines, and depressed the proteins expression of tumor necrosis factor (TNF)-α/nuclear factor (NF)-κB signaling pathway. The full-length 16S rDNA sequence of bacterial cells was also sequenced by high-throughput analysis. CSD caused significant changes in the intestinal flora. PCD improved the species diversity and bacterial abundance in the intestines of rats with anxiety. Metabolomics analysis indicated that 12 PCD-related metabolites in serum and 32 PCD-related metabolites in feces were identified, respectively. Metabolite analysis in serum, PCD treatment affected taurine, hypotaurine, cysteine, methionine, glycine, serine, and threonine metabolism, among others. Metabolite analysis in feces showed significant effects of PCD treatment on the metabolism of vitamin B6, tyrosine, drugs, and glycerophospholipid. Additionally, the correlation analysis of heatmaps showed a tight relationship between inflammatory factors, metabolic parameters, and gut microbial phylotypes. CONCLUSIONS: PCD relieved anxiety by regulating intestinal flora, regulating metabolic disorders, and inhibiting inflammatory pathways in chronic sleep-deprived rats.

Psoralen and isopsoralen from Psoraleae Fructus aroused hepatotoxicity via induction of aryl hydrocarbon receptor-mediated CYP1A2 expression.

ETHNOPHARMACOLOGICAL RELEVANCE: Psoraleae Fructus (PF), a traditional Chinese medicine, has long been used to treat diseases such as cancer, osteoporosis and leukoderma. Psoralen and isopsoralen are main bioactive ingredients of PF with anti-tumor, anti-inflammatory, estrogen-like neuroprotection, etc., meanwhile they are also representative hepatotoxic components of PF. Hepatic CYP1A2 has been reported to be the important metabolic enzymes involved in psoralen and isopsoralen-induced hepatotoxicity. However, the relationship between the hepatotoxicity and CYP1A2 expression, and the underlying mechanism of regulating CYP1A2 expression remain unclear. AIM OF STUDY: The aim of this study was to explore the associated mechanism between psoralen or isopsoralen induced hepatotoxicity and activated aryl hydrocarbon receptor (AhR)-mediated transcriptional induction of CYP1A2 in vitro and in vivo. MATERIALS AND METHODS: Psoralen and isopsoralen at different doses were treated on HepG2 cells (10, 25, 50, 100, 200 µM for 2, 12, 24, 36, 48 h) and mice (20, 80, 160 mg/kg for 3, 7, 14 days) for different time, to assess the correlation of induced hepatotoxicity and CYP1A2 mRNA and protein expression in vivo and in vitro, as well as the effect on CYP1A2 enzyme activity evaluated by phenacetin metabolism. In addition, the potential mechanism of the regulation of CYP1A2 expression mediated by AhR was explored through nucleocytoplasmic shuttling, immunofluorescence, cellular thermal shift assay and molecular docking, etc. RESULTS: Psoralen and isopsoralen induced cytotoxicity in HepG2 cells, and hepatomegaly, biochemicals disorder and tissue pathological impairment in mice, respectively in dose- and time-dependent manners. Simultaneously accompanied with elevated levels of CYP1A2 mRNA and protein in the same trend, and the CYP1A2 activity was remarkably inhibited in vitro but significantly elevated overall in vivo. Besides, psoralen and isopsoralen bound to AhR and activated translocation of AhR from the cytoplasm to the nucleus, leading to the transcriptional induction of target gene CYP1A2. CONCLUSIONS: Hepatotoxicities in HepG2 cells and mice aroused by psoralen and isopsoralen were related to the induction of CYP1A2 expression and activity, whose underlying mechanism might be psoralen or isopsoralen activated AhR translocation and induced increase of CYP1A2 transcriptional expression. Hopefully, these finding are conductive to propose an alert about the combined usage of psoralen or isopsoralen and AhR ligands or CYP1A2 substrates in clinical practice.

Poria cocos water-soluble polysaccharide modulates anxiety-like behavior induced by sleep deprivation by regulating the gut dysbiosis, metabolic disorders and TNF-α/NF-κB signaling pathway.

Poria cocos (P. cocos) has been traditionally used as folk medicine and functional food in China for more than 2000 years. The water-soluble polysaccharide is the main component of P. cocos decoction. The effects and mechanisms of the water-soluble polysaccharide from P. cocos (PCWP) were investigated in chronic sleep deprivation (CSD)-induced anxiety in rats. CSD induced anxiety, gut dysbiosis, and inflammatory responses, and reduced neurotransmitter levels, whereas PCWP intervention ameliorated anxiety-like behaviors, increased the levels of 5-hydroxytryptamine, dopamine, norepinephrine, and γ-aminobutyric acid in the hypothalamus, regulated gastrointestinal peptide levels, reduced inflammatory factors, and inhibited the tumor necrosis factor (TNF)-α/nuclear factor (NF)-κB signaling pathway in rats with CSD. The changes in the intestinal flora composition were determined using 16S rDNA sequencing, and indicated that PCWP significantly improved species richness and diversity in the intestinal flora of rats with anxiety, and adjusted the abundance of the following dysregulated bacteria closer to that of the normal group: Rikenellaceae_RC9_gut_group, Ruminococcus, Prevotellaceae_UCG-001, Prevotellaceae_NK3B31_group, Fusicatenibacter. Metabolomics was used to analyze fecal samples to identify significantly altered metabolites in the PCWP-treated groups. Thirty-eight PCWP-related metabolites and four metabolic pathways such as sphingolipid metabolism, taurine and hypotaurine metabolism, vitamin B6 metabolism, and glycerophospholipid metabolism were explored. The results of serum metabolomics showed that 26 biomarkers were significantly changed after PCWP intervention compared with the model group. The regulatory effects of metabolic pathway enrichment on sphingolipid, phenylalanine, and taurine and hypotaurine metabolism, and validation results showed that PCWP intervention regulated the activity of enzymes involved in the above metabolic pathways. A strong correlation between intestinal bacteria and potential biomarkers was found. Our findings present new evidence supporting the potential effect of PCWP in preventing the progression of anxiety by inhibiting the TNF-α/NF-κB signaling pathway, alleviating metabolic disorders, and ameliorating the gut microflora imbalance.

Precise identification of Celosia argentea seed and its five adulterants by multiple morphological and chemical means.

Celosia argentea seed (CAS) has been used as a traditional Chinese medicine for the treatment of liver damage and eye diseases. CAS is easily falsely harvested and misused, five species from Amaranthaceae family have frequently found to be involved in the adulteration and misapplication, namely Celosia cristata seed (CCS), Amaranthus tricolor seed (ATS), Amaranthus retroflexus seed (ARS), Amaranthus cruentus seed (ACS), and Amaranthus spinosus seed (ASS). For the purpose of identification, multiple morphological means including stereomicroscopy, scanning electron microscopy, normal light and polarized light microscopy were comprehensively employed. As a result, micromorphological and microscopic characteristics were extracted and a diagnostic key to CAS and its five adulterants was proposed for the first time. With respect to the genetically closely related species, viz. CAS and CCS, chemical means were developed to achieve the goal of precise identification. Firstly, triterpenoid saponins in CAS and CCS were fully characterized by an HPLC-QTOF-MS/MS method, a total of 20 triterpenoid saponins including 9 novel members were identified. Secondly, the HPLC-ELSD specific chromatogram was established, in which 12 common peaks were assigned. Finally, after a careful comparison, the peak area ratio of two triterpenoid saponins was discovered as interspecies discriminant marker. In conclusion, CAS and its five adulterants can be precisely identified by multiple morphological and chemical means.

Long-term oral administration of Epimedii Folium induced cholestasis in mice by interfering with bile acid transport.

ETHNOPHARMACOLOGICAL RELEVANCE: Epimedii Folium (EF) is a common traditional Chinese medicine that functions as a tonifying kidney yang to strengthen bones and muscles and dispel wind dampness (limb pain, lethargy, nausea, anorexia, and loose stools). Several studies have reported the potential risk of cholestatic liver damage from EF use; however, there have been few investigations of EF-induced cholestasis, particularly the underlying mechanisms. AIMS OF THE STUDY: The purpose of this study was to evaluate the risk of EF-induced cholestasis in vivo and to explore the mechanisms of action. MATERIALS AND METHODS: ICR mice were orally administered a water extract of EF (WEF) in doses of 6.5 and 19.5 g/kg/day for 14 weeks. Liver-to-body weight ratios, body weight, histopathological examination, and biochemical analyses were performed to assess WEF-induced cholestasis in the mice. Genes associated with bile acid (BA) metabolism and transport, including sodium taurocholate cotransporting polypeptide (NTCP), cytochrome P450 8B1 (CYP8B1), bile-salt export pump (BSEP), multidrug resistance P-glycoproteins 1 (MDR1), and farnesoid X receptor (FXR), were measured at the transcript and protein levels to investigate the potential mechanisms through which cholestasis is aroused by EF. RESULTS: After administration of WEF for 14 weeks, mice in the high-dose WEF group showed poor health with an increased liver-to-body weight ratio as well as higher serum aminotransferase, alkaline phosphatase, direct bilirubin, and total BA levels. Compared with the control group, mRNA expression of NTCP and cholesterol 7a-hydroxylase (CYP7A1) increased, and levels of BSEP, MDR1, multidrug resistance-associated protein 2, and multidrug resistance-associated protein 3 decreased in the WEF-treated group. NTCP, BSEP, MDR1, and CYP8B1 showed similar mRNA and protein expression trends. CONCLUSION: We demonstrated that the long-term oral administration of WEF causes cholestatic liver injury in mice, which is consistent with reported clinical cases. Furthermore, we found that the destruction of BA metabolism and transport is involved in WEF-induced cholestasis. The fine-scale molecular mechanisms of WEF-induced cholestasis and the active compounds of EF need further study.

Quality consistency evaluation between dispensing granules and traditional decoction of Gardeniae Fructus based on chemical similarity and bioequivalence.

Dispensing granules of Chinese herbal medicines are gaining more and more recognition. Despite this, how to evaluate the quality consistency between traditional decoction and its corresponding dispensing granules is a challenging task. In this work, we attempted to propose a comprehensive strategy through in vitro and in vivo comparisons to overcome this challenge, taking Gardeniae Fructus as a typical case. On one hand, HPLC fingerprinting and multi-component quantification were performed to evaluate chemical similarity. On the other hand, pharmacokinetic profiling was conducted to estimate bioequivalence in terms of concentration-time curve and key pharmacokinetic parameters. The in vitro and in vivo comparison results demonstrated that there were no significant differences between these two dosage forms. This proposed strategy is applicable not only for quality consistency evaluation between dispensing granules and traditional decoction but also for broader application scenarios.

Extracts of Poria cocos improve functional dyspepsia via regulating brain-gut peptides, immunity and repairing of gastrointestinal mucosa.

BACKGROUND: Poria cocos (Schw.) Wolf (PC), a fungus, has been used for more than 2000 years as a food and medicine in China. It has a very good therapeutic effect for functional dyspepsia (FD). However, the material basis and mechanism of PC on FD were not reported. PURPOSE: To investigate the function and potential mechanisms of PC including its three extracts (triterpenoid, PCT; water-soluble polysaccharide, PCWP; acidic polysaccharide, PCAP) on FD. STUDY DESIGN: The study explored the therapeutic effect of PC and its three extracts on FD in rats for the first time and discussed its mechanisms based on brain-gut peptides, immunity and repair of the gastrointestinal mucosa. METHODS: The chemical components of PC extracts were analyzed and quantified using ultra high performance liquid chromatography coupled with quadrupole time of flight mass spectrometry (UPLC-Q-TOF-MS) and gel permeation chromatography coupled with size exclusion chromatography (GPC/SEC). The FD rat models were established using weight-loaded forced swimming and alternate-day fasting for 42 days. After 14 days of treatment, the effect and mechanisms were investigated using ELISA, histopathology, immunohistochemistry as well as Western blot. RESULTS: Seventy-seven triterpenoids in PCT were identified. PCWP was primarily composed of component A (Mw: 3.831 × 107 Da), component B (Mw: 5.650 × 106 Da) and component C (Mw: 113,117 Da). PCAP was a homogeneous composition with an average Mw of 74,320 Da. PCT, PCWP and PCAP alleviated the symptoms of FD. These extracts promoted the repair of gastrointestinal mucosa and regulated the balance between the T helper cell (Th)1/Th2 axis and the Th17/Treg axis. PCT and PCWP regulated brain-gut peptides more effectively, PCWP and PCAP enhanced immunity more effectively. Further study demonstrated that these extracts may have enhanced immunity via the Toll-like receptor (TLR) and c-Jun N-terminal kinase (JNK) signaling pathways. CONCLUSIONS: PC extracts showed therapeutic effects on FD rats, and the mechanism of action involved multiple pathways. PCAP, which is often discarded in traditional applications, was effective. Our study provides new ideas for the application and development of PC extracts.

Discovering the main "reinforce kidney to strengthening Yang" active components of salt Morinda officinalis based on the spectrum-effect relationship combined with chemometric methods.

Morinda officinalis, a well-known traditional herbal medicine in China, is used to treat deficiency of kidney-yang syndrome. Although this medicine has the property of "reinforcing kidney to strengthening Yang," the chemical constituents responsible for this effect remain to be elucidated. Here, we aimed to identify the main active compounds responsible for reinforcing kidney to strengthening Yang, based on spectrum-effect relationships combined with chemometrics. We used the UPLC-diode array detection method to establish the chromatography fingerprint of M. officinalis. Hydrocortisone-induced and adenine-induced kidney-yang deficiency patterns were established to evaluate the efficacy of M. officinalis. Serum triiodothyronine, free thyroxine, thyrotropin, testosterone, cortisol, luteinizing hormone, follicle-stimulating hormone, corticotropin-releasing hormone, and adrenocorticotropic hormone levels were determined as pharmacodynamic indices. Analytic hierarchy process was used to determine the weight of each index to the total pharmacodynamic contribution. Lastly, the spectrum-effect between the fingerprint and the pharmacological effects were established using grey relational analysis and partial least squares. Our findings indicated that peaks 1, 2, 3, 5, 6, 7, 8, 9, 11, 13, 15, 17, and 20 might represent the main components that positively correlated to the total effect, of which four were identified by comparison with reference standards. The identified components were monotropein (peak 1), deacetyl asperulosidic acid (peak 3), asperulosidic acid (peak 8), and asperuloside (peak 9). Our results suggest that the "reinforce kidney to strengthening Yang" effects were attributable to the combined effects of the multiple chemical components of M. officinalis and provide a valuable method to identify the active "reinforce kidney to strengthening Yang" components of M. officinalis and establish the quality control of M. officinalis.