Study On The Hypolipidemic Activity And Its Mechanism Of Phenols From Canarium Album L.

With the development of economic and the continuous improvement of people's living standards in China,more and more patients with hyperlipemia increased and developed in a younger age.Hyperlipidemia harms enormously and is closely related with cardiovascular disease which endangers human life.Due to the influence of traditional Chinese medicine and food homology theory,the use of natural extracts as a prescription for prevention and treatment of hyperlipidemia has become a hot research topic in the academic field.Many publications have demonstrated that phenolic compounds obtained from plants have a variety of physiological functions,including blood lipid regulating,liver protecting and so on.Canarium album,a kind of medicine and edible fruit,possessed a wide variety of phenolic components.Therefore,the objective of this research was to optimize the extraction method of phenolic compounds from Canarium album,evaluate the effect of olive phenolic extract(TPCA)on blood lipids,and further explore its mechanism.This research played the certain roles of offering the theory support and practice guidance to promoting the economic value of Canarium album products which could be developed into the functional food for controlling hyperlipidemia.The main research contents and results of this research are as following:1.Different phenolic components contribitions in five cultivas of Canarium album.In order to study the effects of cultiva and maturity on the content of total phenols,flavonoids and ellagic acid,5 major cultivars from Fujian province were studied.The results showed that the contents of total phenols and flavonoids in all cultivars appeared a decreasing trend with the increasing of maturity,the content of ellagic acid reached the maximum at 80%maturity level and then decreased with ripening.The contents of total phenols,flavonoids and ellagic acid in all cultivars appeared a trend as:ChangYing>Zhaoan>ZiLaiYuan>HuiYuan>TangXiang.According to the results of research,the maximum contents of total phenols and flavonoids are 22.18mg/mL and 0.65mg/mL respectively in "ChangYing"cultivas at 70%maturity level,and the content of ellagic acid reached the highest value of 2.52 mg/mL at 80%maturity level.Based on the above results,"Changying" with the maturity of 70?80%level was selected as the cultivas for following extraction of phenolic compounds.2.The effect of olive phenolic extract(TPCA)on blood lipid and liver tissue in non-alcoholic fatty liver(NAFLD)rats.Taking the TPCA(the total phenolic content was 930 mg/mL)from"Changying" as testing material,the effects of blood lipid and liver tissue on NAFLD rats have been studied.The results showed that:compared with the high-fat diet fed rats,fed the TPCA with middle dose(50mg/kg.bw)and high dose(150mg/kg.Bw)can significantly reduce the serum TC?TG and LDL-C levels and enhance the HDL-C levels(p<0.01)in NAFLD rats;at the same time,significantly reduce TC,TG and MDA levels were also observed in liver tissue,the activity of GSH-Px was also increased significantly as well as inhibit the activity of aminotransferase(p<0.01).The liver pathological tissue section in NAFLD rats showed that:fed the TPCA with middle dose(50mg/kg.bw)and high dose(150mg/kg.Bw)can reduce fat vacuoles in the cytoplasm of liver tissue,reduce the infiltration of inflammatory cells,alleviate hepatic steatosis.It can conclude that TPCA can inhibit liver fat deposition and lipid peroxidation in NAFLD rats,resulting the alleviation of liver injury,have obvious effects of reducing blood lipid and liver protection.3.The mechanism of hypolipidemic activity of olive phenolic extract(TPCA)on NAFLD rats(1)The effect of TPCA on lipid accumulation in HepG2 cells The effect of TPCA on lipid metabolism was studied through the MTT test.The expression level of the lipid metabolism related targets were analyzed by real-time PCR assay and Western blot assay.The results showed that compared to normal cells,the lipid content in the model cells was significantly increased,the polyphenols and flavonoids from Canarium album can significantly inhibit the oleic acid-induced accumulation of lipids in HepG2 cells(P<0.001).The Real time PCR and western blotting results showed that that polyphenols or flavonoids extract decreased fatty acid synthase(FAS)and sterol regulatory element binding protein 1(SREBP-1)content,increased PPAR-a content in Mrna in a dose-dependant manner.Western blotting also showed that Canarium album extract could promote the phosphorylation of AMPKal.In summary,we demonstrated that Canarium album extract restrained lipid excessive accumulation induced by oleic acid by up-regulating the phosphorylation of AMPK,downregulating the expression level of FAS and SREBP-1,and upregulating the expression level of PPAR-a in hepatocarcinoma cells.(2)Effect of the phenols from Canarium album(TPCA)on intestinal floraThe high throughput sequencing technology was used to analyse the difference of intestinal flora from colon of mice after treated with the TPCA on NAFLD mice.The most dominant bacteria in the intestine of the mice in each group were Bacteroidetes and Firmicutes.The intestinal flora changes trend of low doses of TPCA fed mice were:with the increasing of dose,the abundance of Bacteroidetes and Firmicutes decreased first and then increased,but the abundance of Verrucomicrobia increased first and then decreased.Compared with healthy mice,the abundance of Bacteroidales_S24-7_group_norank in the intestine of NAFLD mice significantly decreased but significantly increased of Allobaculum.Further comparison reveals that Akkermansia accounted for 6%in the intestinal of healthy mice but almost non-existent in NAFLD mice,and fedding with TPCA can significantly increased Akkermansia to as high as 14%in the intestinal of NAFLD mice.I showed that TPCA can regulate the abundance of intestinal flora in NAFLD mice,especially to increase the quantity of Akkermansia bacteria and regulate the metabolism of high fat diet in vivo as well as thus play a role in reducing blood lipid.(3)The metabolism of the phenols of Canarium album(TPCA)in miceIn order for further exploring underlying mechanism of pathogenesis and liver protecting of TPCA on NAFLD mice,the metabonomics analytical platform of UPLC-MS,metabonomics analysis of serum and liver tissue were used to analysis the organs of the healthy mice,NAFLD and NAFLD fed with low,middle and high dose of TPCA.PCA and PLS-DA analysis showed that TPCA could upregulate the protein and gene expression of adiponectin receptor 2,PPAR-alpha,CPT1,ACOX1(P<0.01).Metabolic pathway analysis showed that TPCA could significantly interfere with the metabolism of purine,tyrosine and lipids,and enhance lipids catabolism.Those results show that TPCA can reduce the liver lipid deposition by enhancing liver lipid oxidation and catabolism,and play a role in the treatment of NAFLD.In summary,the results showed that TPCA have the activities of hyperlipidemia and liver protection,and their underlying mechanisms are as follows:1.TPCA restrained the lipid excessive accumulation by facilitating phosphorylation of AMPK,downregulating the expression level of FAS and SREBP-1,and upregulating PPAR-a expression;2.TPCA can regulate the abundance of intestinal flora,especially increased the abundance of Akkermansia;3.TPCA can upregulate the protein and its gene expressing of adiponectin receptor 2,PPAR-alpha,CPT1,AC0X1(P<0.01);4.TPCA could significantly interfere with the metabolism of purine,tyrosine and lipids,and enhance lipids catabolism.