高效液相色谱法测定婴幼儿配方奶粉中酪蛋白磷酸肽含量

Determination of casein phosphopeptides in infant formula milk powder by high performance liquid chromatography

目的建立高效液相色谱法(highperformanceliquidchromatography,HPLC)测定婴幼儿配方奶粉中酪蛋白磷酸肽(casein phosphopeptide, CPP)的含量的方法。方法对样品静置温度、静置时间、沉淀分离方式进行优化,用高效液相色谱法对婴幼儿配方奶粉中CPP的含量进行测定。结果最优的前处理条件为样品溶解后调整pH至4.6,于4℃条件下静置1 h,过滤后上清液经0.22μm滤膜过滤,以Ultimate AQ-C_(18)色谱柱(250mm×4.6mm,5μm)分离。经钼蓝比色法鉴定,保留时间为8.387min的组分为CPP。该方法在10～1000mg/L范围内线性关系良好,相关系数为0.9993,定量限为4.40 mg/L,婴幼儿奶粉中添加量为500、1000、5000 mg/kg时, CPP的回收率为96.63%%～101.42%。结论该方法样品前处理简单,稳定性和可靠性良好,检测结果符合定性定量要求,可用于婴幼儿配方奶粉中CPP含量的测定。

Objective To establish a method of determination of casein phosphopeptides(CPP) in infant formula milk powder by high performance liquid chromatography(HPLC). Methods The sample standing temperature, standing time and precipitation separation method were optimized, and the content of CPP in infant formula was determined by HPLC. Result The optimal pretreatment conditions were as follows: the sample was dissolved and adjusted to pH 4.6, and allowed to stand at 4 ℃ for 1 h. After filtration, the supernatant was filtered through a 0.22 μm filter and separated by an Ultimate AQ-C18 column(250 mm×4.6 mm, 5 μm). It was identified by molybdenum blue colorimetric method, and the component with retention time of 8.387 min was CPP. The method had a good linearity in the range of 10-1000 mg/L, the correlation coefficient was 0.9993, and the limit of quantification was 4.40 mg/L. When the dosages of infant milk powder were 500, 1000, and 5000 mg/kg, the recovery rates of CPP were 96.63%%-101.42%. Conclusion Themethod is simple, stable and reliable, and the results complied with the qualitative and quantitative requirements, which can be used to detect CPP content in infant milk powder.