发酵酸肉胆酸盐结合肽的分离纯化及初步鉴定

Purification and preliminary identification of bile acid salt binding peptide from fermented sour meat

以传统发酵酸肉为原料,制备胆酸盐结合肽并进行初步鉴定。结果表明:磷酸盐缓冲液溶解法提取发酵20 d酸肉粗肽含量较高且胆酸盐结合能力较好。通过不同极性大孔树脂筛选,发现DA201-C对酸肉粗肽样品的吸附和解吸效果最好,纯化酸肉粗肽最佳工艺条件为上样质量浓度5 mg/m L、上样流速1.5 m L/min、洗脱流速3 m L/min、洗脱体积3.5 BV、洗脱剂体积分数75%乙醇(V/V)。进一步采用葡聚糖凝胶层析分离得到F1和F2两组分,F2组分胆酸盐结合能力较好,是由分子质量范围在265 Da^1.4 k Da的混合小肽组成,F2有高含量的疏水性和芳香族氨基酸。这可能是F2组分具有胆酸盐结合效果的原因,苯丙氨酸可能是其具有胆酸盐结合活性的关键组分。

The cholate binding peptides was prepared and preliminarily identified by using traditional fermented sour meat as raw material. Results showed that the extraction rate of phosphate buffer solution was the highest, the content of crude polypeptide was higher and the binding ability of cholate was better after fermentation for 20 d. It was found that the effect of DA201-C on the adsorption and desorption of acidic crude peptide was the best through screening of different polar macroporous resins. The optimal conditions for the purification of acidic crude peptide with DA201-C macroporous resin were as follows：sample concentration of 5 mg/mL, sample flow rate of 1.5 mL/min, elution flow rate of 3 mL/min, elution volume of 3.5 BV, elution volume fraction of 75% ethanol （V/V）. Two components F1 and F2 were obtained through further separation using dextran gel chromatography, wherein the binding capacity of F2 group was better, F2 group was composed of small peptides with molecular weights ranged from 265 Da to 1.4 kDa and had a high content of hydrophobic and aromatic amino acids. This may be the reason for the binding of F2, and Phenylalanine may be a key component for its binding activity.