一种连续分离纯化酪蛋白磷酸肽的新工艺

A Novel Process of Continuous Separation and Purification of Casein Phosphopeptides

为建立一条制备酪蛋白磷酸肽(CPPs)的新工艺路线。采用酶解与超滤耦联技术初步分离制备酪蛋白生物活性肽,然后通过树脂层析将酪蛋白生物活性肽中的酪蛋白磷酸肽和非磷酸肽分离。选用碱性蛋白酶Alcalase在切向流酶膜反应器中(膜截留分子质量10kD)连续酶解3h、酶解温度50℃、pH8.5。重点研究大孔弱碱性阴离子交换树脂D303对酪蛋白生物活性肽的分离纯化工艺,并采用单因素试验优化CPPs的分离纯化工艺参数,确定D303树脂的较佳动态吸附解吸条件:上样流速1BV/h、上样量100mL、洗脱温度45℃、洗脱酸浓度0.15mol/L HCl,获得磷酸肽N/P比为7.21,P洗脱得率为93.11%。

A novel continuous preparation process of casein phosphopeptides（CPPs）was developed using enzymatic membrane reactor and anion-exchange chromatography.Alcalase was used to continuously hydrolyze 10%casein at pH 8.5 and 50℃for 3 h in a tangential flow filter（TFF）enzymatic membrane reactor（EMR）.The separation and purification of the resulting CPPs using anion-exchange resin D303 was investigated and the technological parameters were optimized by single factor design based on the recovery rate of phosphate and the molar ratio of nitrogen to phosphate.The results showed that the appropriate adorption/desorption conditions of D303 were as follows：loading permeate flow rate of 1 BV/h,sample loading quantity of 100 mL,elution temperature of 45℃,and hydrochloric acid concentration for elution of 0.15 mol/L,and the recovery rate of phosphate from the permeate was 93.11%and the molar ratio of nitrogen to phosphate was 7.21 under these conditions.