Study on untargeted metabolomics of Codonopsis pilosula from different producing areas based on ultra-performance liquid chromatography tandem high resolution mass spectrometry / 药学学报

Lu Dangshen, a traditional authentic medicinal material of Codonopsis Radix is mainly produced in Shangdang (Changzhi) area of Shanxi Province. Baitiao Dangshen is mainly produced in Gansu Province. Codonopsis Radix contains many kinds of components such as phenylpropanoids, polyalkynes, alkaloids, terpenes, fatty acids, flavonoids, and so on. At present, the effect of producing areas on its chemical compositions has not been systematically studied. This study analyzed the differences of metabolites among Codonopsis pilosula from different producing areas by UPLC-HRMS. PCA, OPLS-DA coupled with Thermo mzcloud online and local databases were used to compare the overall differences of metabolites among Codonopsis pilosula from different producing areas, and the chemical constituents were identified to further screen and find out the different metabolites and analyze the metabolic pathways by information retrieval in HMDB, PubChem, Chemspider and KEGG databases. The results showed that 72 differential metabolites were identified in this study. There were 15 kinds of up-regulated and 57 kinds of down-regulated metabolites of Lu Dangshen compared with Baitiao Dangshen. The top 30 metabolic pathways were analyzed by KEGG enrichment, and the most important metabolic pathways were phenylpropanoid biosynthesis, which was demonstrated that phenylpropanoid biosynthesis pathway and related intermediate metabolites could be used as the characteristics of distinguishing Lu Dangshen from different habitats of Codonopsis pilosula. The present study provided a basis for analyzing the influence of producing areas on the chemical components of Codonopsis pilosula and reasonably evaluating the quality of Codonopsis Radix, and also provided a new idea for expounding the authenticity of Lu Dangshen.

Hepatocyte steatosis activates macrophage inflammatory response accelerating atherosclerosis development / 浙江大学学报·医学版

OBJECTIVES@#To investigate the mechanism of comorbidity between non-alcoholic fatty liver disease (NAFLD) and atherosclerosis (AS) based on metabolomics and network pharmacology.@*METHODS@#Six ApoE－/－ mice were fed with a high-fat diet for 16 weeks as a comorbid model of NAFLD and AS (model group). Normal diet was given to 6 wildtype C57BL/6J mice (control group). Serum samples were taken from both groups for a non-targeted metabolomics assay to identify differential metabolites. Network pharmacology was applied to explore the possible mechanistic effects of differential metabolites on AS and NAFLD. An in vitro comorbid cell model was constructed using NCTC1469 cells and RAW264.7 macrophage. Cellular lipid accumulation, cell viability, morphology and function of mitochondria were detected with oil red O staining, CCK-8 assay, transmission electron microscopy and JC-1 staining, respectively.@*RESULTS@#A total of 85 differential metabolites associated with comorbidity of NAFLD and AS were identified. The top 20 differential metabolites were subjected to network pharmacology analysis, which showed that the core targets of differential metabolites related to AS and NAFLD were STAT3, EGFR, MAPK14, PPARG, NFKB1, PTGS2, ESR1, PPARA, PTPN1 and SCD. The Kyoto Encyclopedia of Genes and Genomes showed the top 10 signaling pathways were PPAR signaling pathway, AGE-RAGE signaling pathway in diabetic complications, alcoholic liver disease, prolactin signaling pathway, insulin resistance, TNF signaling pathway, hepatitis B, the relax in signaling pathway, IL-17 signaling pathway and NAFLD. Experimental validation showed that lipid metabolism-related genes PPARG, PPARA, PTPN1, and SCD were significantly changed in hepatocyte models, and steatotic hepatocytes affected the expression of macrophage inflammation-related genes STAT3, NFKB1 and PTGS2; steatotic hepatocytes promoted the formation of foam cells and exacerbated the accumulation of lipids in foam cells; the disrupted morphology, impaired function, and increased reactive oxygen species production were observed in steatotic hepatocyte mitochondria, while the formation of foam cells aggravated mitochondrial damage.@*CONCLUSIONS@#Abnormal lipid metabolism and inflammatory response are distinctive features of comorbid AS and NAFLD. Hepatocyte steatosis causes mitochondrial damage, which leads to mitochondrial dysfunction, increased reactive oxygen species and activation of macrophage inflammatory response, resulting in the acceleration of AS development.

Bletilla striata polysaccharide improves toxic and side effects induced by 5-FU: an untargeted metabolomics study / 中国中药杂志

This study aimed to analyze the effect of Bletilla striata polysaccharide(BSP) on endogenous metabolites in serum of tumor-bearing mice treated with 5-fluorouracil(5-FU) by untargeted metabolomics techniques and explore the mechanism of BSP in alleviating the toxic and side effects induced by 5-FU. Male BALB/C mice were randomly divided into a normal group, a model group, a 5-FU group, and a 5-FU + BSP group, with eight mice in each group. Mouse colon cancer cells(CT26) were transplanted into the mice except for those in the normal group to construct the tumor-bearing mouse model by subcutaneous injection, and 5-FU chemotherapy and BSP treatment were carried out from the second day of modeling. The changes in body weight, diarrhea, and white blood cell count in the peripheral blood were recorded. The mice were sacrificed and sampled when the tumor weight of mice in the model group reached approximately 1 g. TUNEL staining was used to detect the cell apoptosis in the small intestine of each group. The proportions of hematopoietic stem cells and myeloid progenitor cells in bone marrow were measured by flow cytometry. Five serum samples were selected randomly from each group for untargeted metabolomics analysis. The results showed that BSP was not effective in inhibiting colon cancer in mice, but diarrhea, leukopenia, and weight loss caused by 5-FU chemotherapy were significantly improved after BSP intervention. In addition, apoptotic cells decreased in the small intestinal tissues and the percentages of hematopoietic stem cells and myeloid progenitor cells in bone marrow were significantly higher after BSP treatment. Metabolomics results showed that the toxic and side effects of 5-FU resulted in significant decrease in 29 metabolites and significant increase in 22 metabolites in mouse serum. Among them, 19 disordered metabolites showed a return to normal levels in the 5-FU+BSP group. The results of pathway enrichment indicated that metabolic pathways mainly involved pyrimidine metabolism, arachidonic acid metabolism, and steroid hormone biosynthesis. Therefore, BSP may ameliorate the toxic and side effects of 5-FU in the intestinal tract and bone marrow presumably by regulating nucleotide synthesis, inflammatory damage, and hormone production.

Difference in liver injury induced by dictamnine between males and females: based on untargeted metabolomics / 中国中药杂志

In recent years, reports of adverse reactions related to traditional Chinese medicine(TCM) have been on the rise, especially some traditionally considered "non-toxic" TCM(such as Dictamni Cortex). This has aroused the concern of scholars. This study aims to explore the metabolomic mechanism underlying the difference in liver injury induced by dictamnine between males and females through the experiment on 4-week-old mice. The results showed that the serum biochemical indexes of liver function and organ coefficients were significantly increased by dictamnine(P<0.05), and hepatic alveolar steatosis was mainly observed in female mice. However, no histopathological changes were observed in the male mice. Furthermore, a total of 48 differential metabolites(such as tryptophan, corticosterone, and indole) related to the difference in liver injury between males and females were screened out by untargeted metabolomics and multivariate statistical analysis. According to the receiver operating characteristic(ROC) curve, 14 metabolites were highly correlated with the difference. Finally, pathway enrichment analysis indicated that disorders of metabolic pathways, such as tryptophan metabolism, steroid hormone biosynthesis, and ferroptosis(linoleic acid metabolism and arachidonic acid metabolism), may be the potential mechanism of the difference. Liver injury induced by dictamnine is significantly different between males and females, which may be caused by the disorders of tryptophan metabolism, steroid hormone biosynthesis, and ferroptosis pathways.

An Untargeted Urinary Metabolomics Strategy for Investigation of Therapeutical Mechanism of Schisandra chinensis on Complications of Diabetes Rats / 分析化学

An untargeted urinary metabonomics method based on ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry ( UPLC-Q-TOF-MS ) has been established to investigate the mechanism of Schisandra chinensis in treating diabetes and its complications. The urinary biomarkers related to the therapeutic effects of Schisandra chinensis on the diabetes rats were analyzed. In urine, 28 kinds of endogenous metabolites were identified as potential biomarkers, including 13 endogenous metabolites in positive ion mode, 15 endogenous metabolites in negative ion mode, and hippuric acid detected both in positive and negative ion modes. The results revealed that Schisandra chinensis mainly affected the pathways of pentose and glucuronate interconversions, riboflavin metabolism, pantothenate and CoA biosynthesis, arginine and proline metabolism, intestinal bacteria metabolism, ascorbate and aldarate metabolism and tryptophan metabolism in diabetic rats. Combined with biological analysis of these pathways, the therapeutic mechanism of Schisandra chinensis on diabetes and its complications was verified. Based on the biological function of each pathway, the effect of Schisandra chinensis on diabetic nephropathy is stronger. Moreover, it also has the effects of protecting liver, decreasing fat and antioxidant activity.

New approaches for identifying biologically active components of botanical natural products / 中国药理学与毒理学杂志

Countless studies have been devoted to the scientific evaluation of the safety and/or efficacy of botanical natural products. Investigators involved in such studies face a unique set of challenges. Natural products differ from their pharmaceutical counterparts in that they are typically complex mixtures, for which the identities and quantities of components present are not known. To further complicate matters, the composition of these mixtures will vary depending on source material and method of preparation. Investigators conducting clinical trials with complex botanical natural products must choose from a myriad of potential preparations, which may vary greatly in composition. In making such decisions, it is extremely useful to know which components of the mixture are most likely to be responsible for its purported biological activity (the ″active constituents″). The gold standard approach for identifying active constituents of botanical natural products is bioassay-guided fractionation, in which the mixture is subjected to successive rounds of purification and bioassays until an active compound is identified. Bioassay guided fractionation has historically played a critical role in drug discovery, but is, nonetheless, fraught with challenges. The process is biased towards the most abundant and easily isolatable mixture components, which may not be the most biologically active. Furthermore, if multiple compounds contribute either additively, antagonistically, or synergistically to the observed biological activity of the mixture, activity may be lost upon isolation. As a complementary strategy to bioassay-guided fractionation, our research group has developed untargeted metabolomics strategies to aid in the identification of bioactive mixture components. These strategies involve profiling botanical mixtures using ultraperformance chromatography coupled to high resolving power mass spectrometry. The resulting chemical data is then integrated with biological assay data using bioche?mometric data analysis strategies. Several case studies will be presented illustrating how this approach can be applied, including for the identification of compounds from the botanical green (Camellia sinensis) that inhibit drug metabolizing enzymes. Such studies are being conducted as part of the Center for Excellence in Natural Product Drug Interaction Studies (NaPDI), which is supported by a cooperative agreement with the National Center for Complementary and Integrative Health, a component of the National Institutes of Health.