Study On The Screening Of Anthocyanin-Rich Plants And The Mechanism Of Improving Atherosclerosiss

Anthocyanins are natural water-soluble pigments that are widely found in colored plants.Anthocyanins possess anti-oxidation,antiaging,anti-inflammatory and antibacterial activities as well as liver and eyesight protection.With the development of agriculture and biotechnology,more and more anthocyanin-rich plants have emerged,and the structure type,content distribution and biological activities of anthocyanins in these plants are not exactly the same.In addition,the incidence of cardiovascular and cerebrovascular diseases?CCD?is increasing,which is a serious threat to human health,and atherosclerosis?AS?is the common pathological basis for the induction of CCD.Therefore,finding natural production to effectively alleviate AS is essential for reducing the incidence of CCD.In this study,15 anthocyanin-rich plants were selected,and the extraction and purification process of anthocyanins was optimized.And then their anthocyanin content,antioxidant activity and stability of purified anthocyanins were determined and evaluated.The effect and mechanism of the purified anthocyanins from Lycium ruthenicum?LRPA?on AS were studied.The experimental results are as follows:1.The optimization of anthocyaninextraction and purification processTaking purple carrot as the representative,through the design of orthogonal test,the optimal extraction and purification process of anthocyanin was as follows:50%ethanol solution?pH=2?as solvent,material liquid ratio was 1:4,smashing and extracting for 1.5 min,filtered.And then the filter residue was ultrasonically extracted once for 40 min at 25°C with 50%ethanol solution?pH=2?,the material liquid ratio was 1:2.After filtration,the filtrates were combined and concentrated by flash evaporation at 40°C to 1/5 of the original volume to obtain a concentrate.The concentrate was enriched and purified by Diaion HP 2 MGL macroporous resin column chromatography.Firstly,the water-soluble impurities were removed by deionized water,and then changed to the eluent with a 60%ethanol solution?pH=3?at a flow rate of 0.6 mL/min.The dark eluate was collected and concentrated by flash evaporation and rotary evaporation at 40°C to dryness,and obtained total anthocyanin purified powder?anthocyanin purification?.2.Performance evaluation of purified anthocyanins from 15 anthocyanin-rich plantsPurified anthocyanins from 15 plants were prepared by the optimal extraction and purification process.The total anthocyanin content?TAC?and cyanidin-3-O-glucoside?C-3-G?were detected by pH difference method and HPLC,respectively.The anthocyanins content of berries were higher than those of vegetables and the TAC of LRPA was the highest in the 15 plants,but there were only 8 plants containing C-3-G.Stability in different pH,light,temperature conditions and antioxidant capacity with DPPH,FRAP,TRP assays showed that LRPA had stronger activity of scavenging free radical and antioxidation,and the aqueous solution of LRPA had better stability under acidic low temperature and dark conditions.Therefore,the Lycium ruthenicum was selected as the object of the next biological activity study.2.Effect of LRPA on improving atherosclerosisAt the beginning,the AS rat model was constructed by vitamin D₃?700,000IU/kg·BW,i.p.?and high-fat diet for 8 weeks,and vitamin D₃ was divided into four injections every other day.After successful modeling,these rats were randomly divided into 6 groups:normal group?NC;saline?,model group?AS;saline?,simvastatin-positive group?SIM;50mg/kg/d?,low-dose?Low;50 mg/kg/BW?,middle-dose?Mid;100mg/kg/d?,high-dose?Hig;200mg/kg/d?LRPA medication groups.After intragastric administration for 6 weeks,serum and histopathological analysis showed that the lipid and inflammatory factors levels in serum were significantly improved after treatment with simvastatin and LRPA.The intima foam cells,lipid and calcium salt deposition and intimal thickening in aorta and vacuolization in liver were significantly reduced.LRPA significantly improved atherosclerosis with a dose-dependent manner.3.Mechanism of LRPA improving atherosclerosisThe development of AS is closely related to the disorder of gut microbiota,and anthocyanins are mainly metabolized and absorbed in the intestines.16S rDNA sequencing of the cecal contents and the expression of related proteins in aorta and liver of AS rats were detected.LRPA treatment significantly improved small intestine villi integrity,preventing LPS from infiltrating blood;increased the abundance of Bacteroides,Bifidobacteria,Lactobacillus,Akkermansia,Lachnospiraceae_NK4A136_group and Roseburia;reducedtheabundanceofFirmicutes,Enterobacteriaceae andPrevotellaceae_NK3B31_group,thereby inhibiting the expression of arterial NF-kB and VCAM-1inflammatory pathway and the liver cholesterol synthesis protein SREBP-2,and up-regulating The expression of the liver cholesterol metabolism protein CYP7A1,plays a role in improving atherosclerosis.