体积排阻色谱法测定牛奶、乳粉、乳清粉中α-乳白蛋白的含量

Determination the Content ofα-lactalbumin in Milk,Milk Powder and Whey Powder by Size Exclusion Chromatography

建立了体积排阻色谱法检测牛奶、乳粉、乳清粉中α-乳白蛋白含量的分析方法。使样品中的α-乳白蛋白经过盐酸胍变性以及2-巯基乙醇还原后,形成展开的α-乳白蛋白的单体结构,调节pH至6.5~7.5,串联使用两根体积排阻色谱柱MAbPac SEC-1300A(7.8 mm×300 mm),盐酸胍为流动相,等梯度洗脱,用光电二极管阵列检测器在280 nm进行检测。本方法可以同时适用于牛奶、乳粉以及乳清粉中α-乳白蛋白的分离分析。α-乳白蛋白的线性范围为10~500 mg/L,线性相关系数均大于0.99。α-乳白蛋白的检出浓度为3.00μg/mL(S/N=3),定量浓度为10.0 g/mL(S/N=10)。牛奶、乳粉-乳白蛋白含量的分析方法。样1品中的α-乳白蛋白经过盐酸胍变性以及2-巯基乙醇还原后,形成展开的α-乳白蛋白的单体结构,调节pH至6.5~7.5,串联使用两根体积排阻色谱柱MAbPac SEC-1300A(7.8 mm×300 mm),盐酸胍为流动相,等梯度洗脱,用光电二极管阵列检测器在280 nm进行检测。本方法可以同时适用于牛奶、乳粉以及乳清粉中α-乳白蛋白的分离分析。α-乳白蛋白的线性范围为10~500 mg/L,线性相关系和乳清粉中α-乳白蛋白低中高3种浓度的加标平均回收率分别为90.9%~98.2%、100.0%~101.4%和95.1%~100.4%,6次重复性实验的相对标准偏差(relative standard deviation,RSD)均小于10%。该方法简便、准确、灵敏,适合测定牛奶、乳粉和乳清粉中α-乳白蛋白的含量。

To establish an analytical method for detectingα-lactalbumin content in milk,milk powder and whey powder.Guanidine Hydrochloride andβ-mercaptoethanol were used to degenerate and reduct the protein to form the monomer structure ofα-lactalbumin.Two connected volume exclusion chromatography columns MAbPac SEC-1300A(7.8 mm 300 mm)were used to seperate the sample with guanidine buffer as mobile phase and detectedɑ-lactalbumin by a photodiode array detector at 280 nm.The method can be applied to seperate and deteminate the content ofα-lactalbumin in milk,milk powder and whey powder.The linear range ofɑ-lactalbumin was 10-500 mg/L,linear correlation coefficient was greater than 0.99.The detection limit was 3.00μg/mL(S/N=3)and the quntification limit was 10.0μg/mL(S/N=10).The average recovery ofɑ-lactalbumin in milk,milk powde and whey powder was 90.9%-98.9%98.2%,100.0%-101.4%and 95.1%~101.4%,respectively.The relative standard deviation(RSD)of six replicates experiments was less than 10%.The method is simple,accurate,sensitive and suitable for determinating ofɑ-lactalbumin in milk,milk powder and whey powder.