超高效液相色谱分析酵母发酵乳清中的游离氨基酸

Analysis of free amino acids in whey fermentated by yeast using UPLC

建立一种简单、快速、稳定性好的超高效液相色谱法测定酵母菌发酵乳清粉后游离氨基酸的变化。样品经6-氨基喹啉基-N-羟基琥珀酰亚氨基甲酸酯（AQC）柱前衍生后,以AccQ·TagEluentA稀释液、乙腈和超纯水为流动相,梯度洗脱,在10min内实现了17种游离氨基酸的完全分离,在10~500μmol/L范围内呈良好的线性关系,最低检出限在0.0012~0.0024μmol/mL,加标回收率在84.5%~101.8%。结果表明：在乳清粉中检测到12种氨基酸,含量较高的是谷氨酸（Glu）、半胱氨酸（Cys）,经酿酒酵母单独发酵乳清后丝氨酸（Ser）和半胱氨酸（Cys）含量显著上升,其余氨基酸含量均降低,乳酸克鲁维酵母存在的情况下游离氨基酸的含量均降低,这为研究酵母菌发酵乳清粉后游离氨基酸的变化提供理论支持。

A simple rapid and more stable method by ultra performance liquid chromatography was developed for the determination of 17 free amino acids in the whey fermentated by yeast. 6-AminoquinolyI-N-hydroxyl-succinimidyl-carbamate （AQC） was used as pre-column derivatization reagent. The dilution of AccQoTag Eluent A, acetonitrile and water were used as the mobile phases in a gradient elution mode. The 17 free amino acids were absolutely separated within 10 min. Each amino acid showed good linearity in the range of 10-500 μmol/L except cystine in the range of 5-250 lamol/L. The detection limits （S/N=3） of 17 amino acids were ranged from 1.2 μmol/L to 2.4μmol/L and the spiked recoveries in a sample were from 84.5% to 101.8%. The results showed that there are 12 kinds of amino acids detected in the whey powder, glutamic acid （glu） and cysteine （Cys） is higher. After fermented bySaccharomyces cerevisiae alone, the serine and cysteine content increased significantly, the rest are decreased. When adding Kluyveromyces lactis, the free amino acids in whey are all decreased. This method can be applied to determine free amino acids in whey fermentation broth by yeast.