柱前衍生-反相高效液相色谱法测定不同方法煮制的猪肉及其汤汁中的游离氨基酸

Determination of free amino acids in the pork and its broth cooked by different methods using reversed-phase high performance liquid chromatography coupled with pre-column derivatization

建立了反相高效液相色谱(RP-HPLC)分离检测用不同方法煮制的猪肉及其汤汁中17种游离氨基酸的方法。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,采用Nova-PakTM C18色谱柱分离,以AccQ.Tag Eluent A稀释液、乙腈和超纯水为流动相,梯度洗脱,检测波长为248 nm,在47 min内实现了17种氨基酸衍生物的基线分离。各氨基酸在1～100μmol/L(胱氨酸在0.5～50μmol/L)范围内呈现出良好的线性关系,相应的线性相关系数(r2)均大于0.99;17种氨基酸的检出限(以信噪比为3计)为0.29～0.96μmol/L;在汤汁中的加标回收率为86.5%～101.0%。该方法前处理过程简单,分离效果好,是检测肉及其汤汁中氨基酸的有效手段,可应用于肉制品的质量评定和工艺优化。

A reversed-phase high performance liquid chromatographic（RP-HPLC） method was developed for the determination of 17 amino acids in the pork and its broth cooked by different methods.6-Aminoquinolyl-N-hydroxyl-succinimidyl-carbamate（AQC） was used as pre-column derivatization reagent.In the RP-HPLC method,a Nova-PakTM C18 column was used with the dilution of AccQ·Tag Eluent A,acetonitrile and water as the mobile phases in a gradient elution mode.The 17 amino acids were baseline separated within 47 min with ultraviolet（UV） detection at 248 nm.Each amino acid showed good linearity in the range of 1-100 μmol/L except cystine（Cys2） in the range of 0.5-50 μmol/L with the correlation coefficient（r2） more than 0.99.The detection limits（S/N=3） of 17 amino acids were ranged from 0.29 to 0.96 μmol/L,and the spiked recoveries in a cooked broth sample were from 86.5% to 101.0%.The results showed that the proposed method can be applied to determine amino acids for meat quality assessment and process optimization with simple pretreatment and good separation.