建立了一种基于反相高效液相色谱(RP-HPLC)来分离并定量测定脱脂乳、加热脱脂乳及脱脂乳粉中未变性的乳清蛋白的分析方法.采用Venusil ASB-C8色谱柱；流速0.8 mL/min;检测波长215 nm;柱温30℃;流动相A为体积分数0.1％的三氟乙酸水溶液;流动相B为体积分数0.1％的三氟乙酸乙腈溶液;洗脱方式:流动相B开始在38％下等度洗脱2 min,然后在5 min内上升到43％,最后在43％下等度洗脱13 min.此方法在20 min内将4种乳清蛋白组分分离且各蛋白线性关系良好,适合定性和定量检测未变性的乳清蛋白组分.%Four main proteins (α-La,β-LgA,β—LgB, BSA) with the undenatured whey protein in skim milk,heat treatment skim milk and skim milk powder were isolated and determined by RP-HPLC. It was achieved by using a Venusil ASB-C8 column(250 mmx4.6 mm,rn5 μm,300A) at a flow-rate of 0.8 mL/min, column temperature 30 ℃ and monitored with 215 nm. A gradient of solvent A 0.1% (v/v) TFA dissolved in water and B consisting of 0.1% (v/v) TFA in acetonitrile was applied. The gradient started with 37% of solvent B keeping for 2min and was increased to 43% within 5min and then kept at 43% for 13 min. This method successfully separated these four proteins within twenty minutes.With good linear correlations about the four proteins , it was used to qualitative and quantitative analysis the composition of the undenatured whey proteins.
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