| Hyperlipidemia is a very common diseases in our society.Diseases caused by hyperlipidemia such as atherosclerosis and cardiovascular diseases have been a serious threat to our health.At present, the treatment of hyperlipidemia is mainly based on drugs, although it has some effect, it also have inevitable some side effects inevitably. Therefore, the development of hypolipidemic drugs is particularly important.Bioactive peptides derived from protein degradation intermediates have many physiological functions. Large number of studies showed that the enzymatic hydrolysis products of various dietary proteins can effectively reduce the level of blood lipids in the body and the preparation of bioactive peptides from specific proteins has become a research hotspot.In this paper,using modern technology to screen, isolate, purificat and identify active peptides with lowering blood lipid activity from different fermentation time sour meat and studing its antioxidant activity in vitro and its effect on function factor of rat thoracic aorta endothelial cell with lab-made fermented sour meat as raw material to provide basic research data and development guidelines for the development of traditional fermented functional food sour meat and avoid adverse effects to health because of eating pork with too much saturated fat acid. The main results are as follows:1. By measuring the inhibition ability of crude peptides from different fermentation time sour meat to 3-hydroxy-3-methylglutaryl-coenzyme A reductase(HMGR),we found that all crude peptides from different fermentation time sour meat have no inhibitory activity to HMGR. Crude peptides from different fermentation time sour meat all have certain binding capacity to 3 kinds of bile acid salts,crude peptides from 20 days fermented meat with the highest binding capacity, the average binding capacity is 0.445±0.036 μmol/mg,slightly higher than 40 days fermented meat peptides and significantly higher than 0 and 60 days fermented meat peptides. This showed that crude peptides from fermented sour meat can achieve the effect of lowering cholesterol levels by combining with bile acid salts.2. Four kinds of resin were sifted by adsorption-desorption experiment, and the DA201-C resin was chose as the best one to purify crude peptides. Dynamic adsorption and desorption experiments were performed to determine the optimum process conditions of purifying sour meat crude peptides through DA201-C resin. The optimum process conditions were : concentration of samples 5 mg/m L, sample velocity 1.5 m L/min, elution velocity of 3 m L / min, elution volume 3.5 BV,elution solvent volume fraction of 75% ethanol.In these conditions,the binding capacity to sodium cholate, glycocholate sodium and sodium taurocholate of elution fractions compared to 20 days fermented crude peptides were respectively increased by 43.8%, 60.1%, 26.7%.The component separated through macroporous resin was divided into two components F1 and F2 by Sephadex G-25 gel chromatography. The molecular weight range of F1 was 1.6 k Da-5.1kDa and F2 was 265 Da-1.4 kDa.The binding capacity to bile acid salts of F1 showed a certain decline but the binding capacity of F2 increased greatly, the average binding capacity to the bile acid was increased from 0.616 μmol/mg to 0.767 μmol/mg, especially its binding capacity to sodium taurocholate, was about 1.75 times higher than before.Using RP-HPLC analyzing F2, 6 elution peaks were obtained. Amino acid composition analysis showed that the F2 component contains 17 kinds of amino acids, including 7 essential amino acids, 5 semi essential amino acids, 5 kinds of non essential amino acids.The highest content in F2 were histidine, phenylalanine and glutamic acid and they occupied 14.82%, 14.15%, 9.13% of the total amount of amino acids respectively, accounting for about 38.1% of the total amount of amino acids. Hydrophobic amino acid and aromatic amino acid accounted for about 46.58% and 34.45% of the total amount of amino acids respectively.Through comparing with related research we inferred that high content of hydrophobic and aromatic amino acids may be the reason F2 group own binding effect on bile acid salts and phenylalanine may be a key component.3. Studing the physicochemical properties of bile salts binding peptides,it showed that bile salts binding peptides owned good temperature stability. At 4℃,25℃,50℃ conditions,its binding ability didn’t have significant change,when at 75℃ and 100℃ conditions,bile salts binding ability decreased significantly, but still have a high binding capacity. Bile acid peptides were not sensitive to p H changes, it had high binding ability no matter in acid or alkali conditions. The binding ability of peptides had a certain degree of reduce when treated by digestive enzyme, but still had a high binding activity.4. Determination the antioxidant activity of bile salts binding peptides in vitro and studing its effects on functional factors in rat thoracic aorta endothelial cells, the results showed that F2 groups had good scavenging activity to hydroxyl radical, superoxide anion radical and DPPH radical,when at 5 mg/mL scavenge rate was 88.7%, 72.7%, 78.3%, respectively.The removing power to hydroxyl radical was the best,the semi scavenging concentration was IC50=2.04 mg/mL.Bile salts binding peptides had no obvious toxicity to endothelial cells and could promote the release of NO in vascular vascular endothelial cells, but the effect is not obvious. It also could promote the secretion of prostaglandin in vascular endothelial cells and its promotion effect was correlated with the dose of bile salts binding peptides in a certain concentration range.This showed that bile salts binding peptides could reduce the content of cholesterol through binding bile salts,on the other side, bile salts binding peptides could regulate blood lipid disorder and inhibit atherosclerosis by romoving free radicals.At the same time, bile salts binding peptides could promote the secretion of NO, PGI2 in vascular endothelial cells to prevent atherosclerosis and inhibit coronary heart disease. |
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