Analysis Of Major Chemical Components In Guava Leaves And Investigation Of Its Treating Type 2 Diabetes On Amino Acids Profile

Guava leaves extracts have been reported to have hypoglycemic activity,which has been widely used in diabetes treatment in the folk.However,its chemical composition is complex,and its mechanism has not been clearly explained.Through the optimization of extraction condition and GC-MS analysis combined with molecular weight alignment,the comprehensive and rapid identification of the volatile components in guava leaves was established.To establish an HPLC method for the determination of ursolic acid and oleanolic acid in guava leaves,the enrichment conditions of the two components were optimized by the QuEChERS method.By analyzing the changes of plasma amino acids profiles of type 2 diabetic rats induced by guava leaves extracts,and the correlation analysis of pharmacodynamics results,the pathway and molecular mechanism of hypoglycemic activity of guava leaves extracts for type 2 diabetes were discussed,which provides experimental and theoretical basis for the diabetes prediction and the development of new drugs for the treatment of diabetes.This paper is mainly divided into the following aspects:1.Through GC-MS analysis,a comprehensive and rapid identification of the volatile components of guava leaves were established.In this paper,the volatile components extracted from guava leaves by steam distillation(SD)was investigated and comprehensively analyzed.To obtain more sensitive and accurate analysis,the sample preparation procedure for Gas Chromatography-Mass Spectrometry(GC-MS)analysis of the volatile compounds was investigated and then 27 compounds were identified in the samples obtained by SD method.The volatile components in guava leaves were made up largely of sesquiterpenoids,including β-cubebene,caryophyllene,aromadendrene,a-cadinene,and calamenene.The results indicated that the volatile components from the guava leaves extracted by SD technology was quite different,and the research provided comprehensive information for the volatile compounds in the guava leaves2.Through the investigation of QuEChERS,the active ingredients,ursolic acid and oleanolic acid,in the guava leaves were enriched,and the HPLC method for the determination of ursolic acid and oleanolic acid in guava leaves were established.Under the optimized conditions,the method showed good linearity over a range of 1-320 g mL-1,with correlation coefficients above 0.999.The limits of detection of UA and OA were 0.18 and 0.36 μg mL-1,respectively.The intraday and interday precision were below 1.95 and 2.55%,respectively.The accuracies of the UA and OA determinations ranged from 97.4 to 111.4%.The contents of UA and OA in the guava leaves samples were 2.50 and 0.73 mg g-1,respectively.Compared with last references using ultrasonic assist extraction,microwave assist extraction and solvent accelerated extraction,QuEChERS has the advantages of less experiment time,high extraction efficiency and less use of organic solvents,which were more green,convenient,economical and effective.3.To establish the chromatographic analysis method for the determination of 21 amino acids in rat plasma,the amino acids were derivatized with 4-fluoro-7-nitro-2,1,3-benzoxadiazole(NBD-F),combined with HPLC-UV for quantitative analysis.The linear correlation coefficients were more than 0.99,the intra-day precision(n = 5)was 0.88-6.19%,and the inter-day precision(n = 5)was 2.01-6.81%.The range of the limits of detection was 0.0177-0.2286 μmol/L;the range of the limits of quantitation was 0.0593-0.7619 pmol/L,and the recoveries rate were 90.23%-115.65%.The concentrations of amino acids were in the range of 34.40-486.09 μmol/L.The results showed that the analytical method is simple,low cost,high sensitivity and good separation effect.It can be used for the determination of the concentrations of amino acids in rat plasma.4.Three doses of guava leaves extracts and positive control drug glipizide were used to intervene in T2DM rats.The study found that guava leaves extracts can improve fasting blood glucose and lipid levels in T2DM rats and impaired glucose tolerance in T2DM rats.And found when the dose was 1200 mg kg-1,guava leaves extracts have the most significant effect on the improvement of hyperglycemia,blood lipid and glucose tolerance in T2DM rats.Through the analysis of amino acids profile ine the gavage administration of 10,20,28 days,the concentraction datas modeled by PCA and OPLS-DA to find the different metabolites,and related metabolic pathways were constructed through SIMCA-P 14.1 and MetaboAnalyst 3.0 database,for deeply analysis of the mechanism of hypoglycemic effect in guava leaves extracts.