Study On The Stability And Mechanism Of Assisted Anti-obesity Effect On High-fat Diet Mice Of Blueberry Polyphenols

Blueberry is belonging to family Ericaceae and genus Vaccinium,its fruit is blue and round,and is one of the five fruits recommended by the World's Food and Agriculture Organization?FAO?.Researches have shown that,blueberry fruits have many beneficial to human bodies and can prevent chronic diseases.Chronic diseases mostly related to the production of free radicals in human bodies,and blueberry has a strong antioxidant activity,and the antioxidant activity of blueberry could be derived from polyphenols.Obesity can increase the incidence of cardiovascular disease,type 2 diabetes,cancer,degenerative arthritis,and other chronic diseases.Therefore,the molecular mechanism of anti-obesity of blueberry polyphenols was studied to provide a theoretical basic for the development of natural functional food and weight loss drug.The extraction and purification technology of polyphenols from blueberry was optimized,and the stability of blueberry polyphenols was studied,the antioxidant activity of blueberry polyphenols was analyzed by chemical method and cell experiment,the changes of composition and antioxidant activity of blueberry polyphenols during in vitro gastro-intestinal digestion were compared,animal experiment was used to investigate the anti-obesity molecular mechanism and effect of blueberry polyphenols on physiological index,lipid metabolism,adipose tissue morphology,and gut microbiota.Through the research,the conclusions can be summarized as follows:Polyphenols was extracted from blueberry by ultrasonic technology.Based on the single factor experiment results,RSM?Response Surface Methodology?was used to optimize the technological parameter.Adjustment through practical feasibility,the optimum extraction conditions were as follows:material-to-liquid ratio of 1:26,extracting temperature of 40°C,ethanol concentration of 45%and extraction time of 120 min.Under these optimal conditions,the yield of polyphenols was 5.27 mg GAE/g.To purify the blueberry polyphenols,the macroporous resin XAD-7 was selected as the most suitable from among six candidates?D1400,S-8,D101,X-5,XAD-7,and AB-8?.The isothermal adsorption results indicated that the process was best explained by the Langmuir isotherm model,and polyphenols was adsorbed by XAD-7 through single molecular layer,and the adsorption was an exothermic process.The best purification parameters,optimized by adsorption and desorption tests,were determined as:blueberry polyphenols concentration of 2.0 mg/g;pH 2.0;ethanol desorption solution concentration of 70%?v/v?;flow rates for feeding and elution of 1 and 2 bed volumes?BV?/h,respectively;and a water wash volume of 3 BV.Under the optimum conditions,the purity of the blueberry polyphenols was improved from 46.3 to 86.48%.The antioxidant activities of the purified polyphenols were investigated.The results showed that the scavenging capacities of the blueberry polyphenols toward O^2-,ABTS and DPPH radicals were 15.51±1.25,9.32±0.76,and 7.10±0.58 mmol VCE/g,respectively.pH value,temperature,light,preservative,sugar and oxidant have significant effect on stability of blueberry polyphenols,pH value,temperature,light,preservative,sugar,oxidant and reductant have significant effect on antioxidant activity of blueberry polyphenols.Therefore,blueberry polyphenols should be preserved under low temperature and dark condition,and preservative,oxidant and reductant is not suitable to be added to the product.The thermal degradation of blueberry polyphenols satisfied the first–order kinetic equation and Arrhenius model;the degradation of blueberry polyphenols increased with the increase of temperature,and the active energy was 48.99 KJ/mol.The enthalpy was positive indicates that the thermal degradation of blueberry polyphenols was an endothermic process;the Gibbs free energy was positive indicates that the thermal degradation of blueberry polyphenols was not a spontaneous reaction;the entropy change was negative indicates that the transient state freedom was less than that of reactant.The results of polyphenols,anthocyanin,and flavonoid content and antioxidant activity before and after in vitro-simulated gastro-intestinal digestion suggested that,polyphenols,anthocyanin,and flavonoid were relatively stable under a gastric environment?the recoveries were 98.45%,93.90%and 102.77%,respectively?,polyphenols and flavonoid content were significantly increased?the recoveries were 141.67%and 161.41%,respectively?,anthocyanins was unstable under an intestinal environment?the recovery was 3.38%?,only a few phenolic compounds entered into the dialysis membrane?the recoveries of polyphenols,anthocyanin,and flavonoid were 13.93%,1.95%and 15.68%,respectively?,indicated that the bioaccessibility of polyphenol,anthocyanin,and flavonoid was very low.Furthermore,compared to the undigested extracts,the number of individual phenolics of extracts after in vitro digestion reduced 10.In vitro-simulated gastro-intestinal digestion significantly decreased the antioxidant activity of blueberry polyphenols,total antioxidant capacity?T-AOC?value,ABTS value and cellular antioxidant activity?CAA?value of blueberry polyphenols entered into the dialysis membrane were 8%,17%and 15%of undigested extracts,respectively.Changes of phenolic compound antioxidant activity during digestion correlated to polyphenols,flavonoid,and caffeic acid concentrations.Mice fed high-fat diet?HFD?exhibited increased body weight and adipose tissues weight,disordered lipid metabolism,induced severe microvesicular steatosis in hepatocytes and larger adipose cell sizes.In contrast,blueberry polyphenols inhibited body weight gain,returned lipid metabolism to normal,improve the pathological changes of hepatocytes and reduce the size of adipocytes of mice.Furthermore,blueberry polyphenols changed the composition of the gut microbiota and modulated specific bacteria in mice,such as Proteobacteria,Deferribacteres,Actinobacteria,Bifidobacterium Desulfovibrio,Adlercreutzia,Helicobacter,Flexispira,Adlercreutzia and Prevotella.The present results supplement the insight into underlying anti-obesity mechanisms of blueberry polyphenols,which provide a theoretical basis for its use as functional food to prevent obesity and alternative drugs for adjuvant treatment of obesity.