| Objective:Taxifolin is a dihydroflavonol compound.The previous research of the group found that it has antioxidant activity in vitro.This study is based on liquid chromatography-high resolution mass spectrometry?LC-MS?techniques to optimize metabolomics and lipidomics analysis methods.Through the metabonomic analysis and lipidomics analysis of serum and liver tissue,we explored the mechanism of Taxifolin protecting liver from subacute liver injury induced by carbon tetrachloride?CCl4?in rats,and evaluated the pharmacodynamics of Taxifolin more systematically and scientifically.Method:?1?Animal model preparation:Healthy male Wistar rats were random Ly divided into blank group,model group,Taxifolin high?45 mg/m L?,medium?30 mg/m L?,and low?15 mg/m L?dose administration group and silymarin positive control group?20mg/m L?.Subacute liver injury models were established by intragastric administration of50%CCl4 at a dose of 1 m L/kg,twice a week.At the same time,the blank group and the model group were given distilled water daily,and the Taxifolin group and the positive control group were given corresponding drugs daily.The dosage was 1mL/100g,once a day.The interval of two times of gastric administration was more than4 hours for 4 weeks.Blood and liver were collected to detect the levels of alanine aminotransferase?ALT?,aspartate aminotransferase?AST?and triglyceride?TG?in serum.The changes of liver tissue in rats were observed by HE.?2?Metabolomics study:Based on LC-MS,serum and liver tissue pretreatment methods and LC/MS analysis conditions were investigated and optimized,and endogenous sources for serum and liver tissues were established.metabolomics analysis of small molecule metabolite analysis,method validation of the final optimization method.By comparing the differences of metabolic profiles in serum and liver between model group rats and blank group rats of rats,the differential metabolites were marked,and the regulation of different metabolites by Taxifolin was analyzed to explain its mechanism of liver protection.?3?Lipidomics study:Based on LC-MS,serum and liver tissue pretreatment methods and LC/MS analysis conditions were investigated and optimized,and established for serum and liver tissue.lipidomics analysis of sourced small molecule lipid metabolites was performed to validate the method of final optimization.By comparing the differences of lipid metabolites in serum and liver between model rats and blank rats,the differential metabolites were labeled in the lipid metabolism network,and the regulation of Taxifolin on lipid differential metabolites was analyzed to reveal the mechanism of liver protection.Results:?1?Successful copy of rat subacute liver injury model showed that ALT and AST increased,ALT and AST decreased in the middle and low dose groups compared with the model group,and the lowering effect was the most significant in the middle dose group,suggesting that Taxifolin had protective effect on liver,while ALT and AST increased in the high dose group,suggesting that Taxifolin had protective effect on liver in a certain dose range.Too much will aggravate the damage to the liver.The liver morphology of rats was observed by HE.The results showed that the liver of model group was severely damaged and fat balloon-like changes appeared in a large area.The liver damage of Taxifolin medium and low dose group was lighter than that of model group,and the liver damage of middle dose group was the least,while that of high dose group was not significantly different from that of model group.?2?To optimize and validate the metabolomics research method,and to analyze endogenous small molecule metabolites in serum and liver tissues by this method.Serum metabolomics study found that 31 different metabolites were identified in the model group compared with the blank group,mainly phospholipids,amino acids,fatty acids,bile acids,energy metabolism related substances and other small molecular compounds.In the regulation of these differential metabolites,it was found that the middle dose group had the most number of callback compounds and had a callback effect on 12 differential metabolites,involving phospholipid metabolism,amino acid metabolism,fatty acid metabolism,bile acid metabolism and energy metabolism pathway.Liver tissue metabolomics studies found that the model group identified 29differential metabolites,mainly phospholipids,amino acids,bile acids,energy metabolism related substances and other small molecule compounds.Similarly,the middle dose group had the most number of callback compounds,and there were a tendency to call back for 7 differential metabolites,involving phospholipid metabolism,amino acid metabolism,bile acid metabolism,and energy metabolism pathway.?3?Serum and liver tissue were analyzed using an optimized and validated lipidomics study.Serum lipidomics studies found that 65 lipid differential metabolites were identified in the model group compared with the blank group,mainly triglycerides,lecithin,sphingomyelin,hemolytic dimethylphosphatidylethanolamine,platelet activating factor,and double is a potential biomarker for phosphatidylethanolamine.Observing the regulation of these lipid differential metabolites by Taxifolin,it was found that the middle dose group had the most number of callback compounds and had a callback effect on 19 lipid differential metabolites.Glycerol phospholipid metabolism,sphingolipid metabolism,glycerol lipid metabolism pathway.Liver tissue lipidomics studies found that 71 lipid differential metabolites were identified in the model group compared with the blank group,The main biomarkers are triglyceride,lecithin,lysophosphatidylcholine,lysophosphatidylcholine,lysophosphatidylcholine,glycerol diester,dimethylphosphatidylethanolamine,platelet activating factor,phosphatidylserine serine,phosphatidylethanolamine,monogalactose diacylglycerol and phosphatidyl methanol.The number of callback compounds in medium dose group was the largest,and there was a tendency of callback to 35 lipid differential metabolites,involving glycerol phospholipid metabolism and glycerol metabolic pathway.Conclusion:?1?Taxifolin has protective effect on subacute hepatic injury induced by CCl4,and the protective effect increases with the increase of dosage in a certain range.Excessive dosage will aggravate the hepatic injury.?2?In the metabolomics study of serum and liver tissue,amino acids,bile acids,fatty acids,terpenoids,vitamins,energy metabolism related substances and other small molecule compounds were found as possible potential biomarkers.Mechanism may be related to regulation of amino acid metabolism,bile acid metabolism,fatty acid metabolism,sputum metabolism,vitamin metabolism disorders.?3?In serum and liver tissue lipidomics studies,triglycerides,lecithin,sphingomyelin,hemolytic dimethyl phosphatidylethanolamine,lysolecithin,lysophosphatidylethanolamine,diglyceride,bismethylphosphatidylethanolamine,Platelet activating factor,phosphatidylserine,phosphatidylethanolamine,monogalactosyl diacylglycerol,phosphatidyl methanol are possible lipid differential metabolites.The hepatoprotective mechanism of Taxifolin may regulate glycerophospholipid metabolism,sphingolipid metabolism,glycerol Related to ester metabolism disorders. |
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