HPLC法测定乙酰谷酰胺中异构体的含量

Determination of isomer in acetyl glutamine by HPLC

目的建立高效液相色谱法测定乙酰谷酰胺中异构体N-乙酰-D-谷氨酰胺的含量测定方法。方法采用高效液相色谱法,直链淀粉-三(S)-α-甲苯基氨基甲酸酯为填充剂(AS-H,4.6 mm×250 mm,5μm)手性色谱柱,检测波长210 nm,柱温30℃,流动相为正己烷(每1 000 mL加0.5 mL三氟乙酸和0.5 mL三乙胺)∶无水乙醇(每1 000 mL中加0.5 mL三氟乙酸和0.5 mL三乙胺)=90∶10。结果浓度在1.289 3~36.099 5μg·mL^(-1)范围内线性关系良好(r=0.998 2,n=5),平均回收率94.9%,RSD=1.6%(n=9),方法的检测限LOD为0.314 6μg·mL^(-1),定量限1.258 4μg·mL^(-1)。结论所用方法专属性强、结果准确、稳定性好,适用于乙酰谷酰胺中异构体N-乙酰-D-谷氨酰胺的检测。

Objective To establish an HPLC method for the determination of isomer N-acetyl-D-glutamine in acetyl glutamine.Methods HPLC was adopted,amylose-3（ S）-alpha-methylphenylcarbamate was used as filler（ AS-H,4.6 mm× 250 mm,5 μm） chiral chromatographic column.The detection wavelength was 210 nm,the column temperature was 30 ℃,the mobile phase was n-hexane（ 0.5 mL trifluoroacetic acid and 0.5 mL triethylamine per 1 000 mL） ： anhydrous ethanol（ 0.5 mL trifluoroacetic acid and 0.5 mL triethylamine per 1 000 mL） = 90∶ 10.Results The linear range of n-acetyl-dglutamine was 1.289 3- 36.099 5 μg·mL^（-1）（ r= 0.998 2,n= 5）.The average recovery was 94.9%,RSD= 1.6%（ n= 9）,and the detection limit of the method was 0.314 6 μg·mL^-1,and the quantitative limit was 1.258 4 μg·mL^-1. Conclusion The method was specific,accurate and stable,and can be used for the detection of the isomer N-acetyl-D-glutamine in acetyl glutamine.