Screening Of Hypolipidemic Components From The Vinegar Powder And The Regulation Of Lipid Metabolism

In recent years, with the change of lifestyle and dietary habits and other factors, the incidence of hyperlipidemia showed a significantly increasing trend. Hyperlipidemia is a disorder of lipid metabolism, and considered to be associated with atherosclerosis, coronary heart disease, myocardial infarction, cerebral paralysis and other cardiovascular and cerebrovascular diseases which had become a serious threat to human health. Vinegar, as a folk medicine since ancient times in China, has the effect of weight loss and lipid-lowering which was confirmed by many studies. Vinegar powder (VP) as a new member in the vinegar families was easy to transport. The original nutrition functional properties of vinegar were retained, and the original irritation was reduced. However, there is still little research on it.In this thesis, VP was prepared by spray drying from the traditional vinegar and hypolipidemic effects of the product were evaluated by animal experiments. Screening methods of hypolipidemic composition were established, and lipid-lowering active ingredients were isolated and purified from VP. The characteristic components of VP were determined and we aim to investigate the hypolipidemic material basis.The conditions of VP prepared were determined, and the characteristic components determined too. It laid the foundation for hypolipidemic product development. Animal experiments showed that the VP significantly decreased the serum total cholesterol (TC) (22.2%, P<0.01) and triglycerides (TG) (24.4%, P<0.01), increased high-density lipoprotein cholesterol (HDL-C) (22.0%, P<0.05) and HDL-C/TC (48.3%, P<0.01) in rats, while weight gain suppression was not significant (P>0.05).Screening methods of hypolipidemic composition for VP were established in vitro, including micellar bile method, based on high-throughput micro colorimetric lipase inhibitor screening method, cholesterol efflux screening model based on the Hep-G2 and RKO cells, liver X receptor activator of Cell Screening Model and a stable cell line selection. These four screening methods as a screening platform to establish foundation for development of lipid-lowering drugs and products, can be use to look for hypolipidemic components from different sources, multiple targets of herbs and other plants, natural products, microbial fermentation products by high-throughput screening.Two dipeptides from water-soluble components of VP were first isolated and purified, and identified as 3-isobutyl-hexahydropyrrolo [1,2-a] pyrazine-1,4-dione and 3-benzyl-hexahydropyrrolo [1,2-a] pyrazine-1,4-dione. The small molecular substances were extracted and purified, by modern chromatography (silica gel column chromatography, LH-20, Sephadex G-15, HPLC-ODS) and recrystallization, and identified by spectroscopic (MS, ESI, NMR) nine components, namely,2-methyl-3-hydroxy-γ-pyrone, TMP,2-hydroxy-3-phenylpropionate, hydroxyphenyl propionate, syringic acid, butyl 2-(2-acetamidopropanamido) acetate,5-acetyl pyrimidine-2,4,6 (1H,3H,5H)-trione, L-cycloserine, piperazine.A rapid and selective HPLC method was developed for the simultaneous analysis of acetoin (ACT) and tetramethylpyrazine (TMP). This method permits the simultaneous determination of ACT and TMP in fermentative foods with detection limits of 5.625μg/mL and 0.033μg/mL, respectively. The recovery was 96.03% for ACT and 92.06% for TMP. Correlation coefficients were greater than 0.9999 for the two compounds. The linearity ranges for ACT and TMP were in the range of 0.02-20 mg/L and 0.12-80μg/mL, respectively. The proposed method could be used for routine quality control of foods and beverages. The detection method is simple and rapid, sample preparation is simple, scientific and effective. The contents of ACT and TMP in 36 samples of Chinese black vinegars were quantified using the preceding HPLC method. The contents of them were fluctuating in the larger context. However, current data suggest that the content of TMP in vinegars is positively correlated with the content of ACT. In addition, another rapid and selective HPLC method was developed for dipeptide of 3-benzyl-hexahydropyrrolo [1,2-a] pyrazine-1,4-dione and found to be accurate, precise, and linear across the analytical range. The proposed method established foundation for further research on 3-benzyl-hexahydropyrrolo [1,2-a] pyrazine-1,4-dione in vinegar and products thereof.The water-soluble components of VP were-rich in polyphenols, flavonoids and other antioxidants, which had good free radical scavenging ability. Cell experiments confirmed that components of VP could remove cells ROS, decreased MDA levels of cells. That means VP could suppress lipid per-oxidation and then protect cells. VP inhibited the enzyme activity of lipase in a dose-dependent manner, and it is strongest inhibition of the components of molecular weight more than lOkDa. The purified components inhibited micelle formation of bile in vitro in different degrees, means that VP could inhibit the absorption of cholesterol.Cell experimental results showed that TMP and dipeptide had antioxidant capacity, which could significantly clear ROS and MDA and conspicuously increased the activity of SOD and CAT. Its antioxidant mechanism may be increased cell in the activity of SOD and CAT by suppressing lipid peroxidation. Furthermore, they could significantly improve intracellular cholesterol efflut, and significantly activate LXR receptors. Therefore, TMP and dipeptide showed prominently potential hypolipidemic function.