柱前衍生化反相高效液相色谱法测定N-甘氨酰-L-谷氨酰胺一水合物含量

Determination of N - Glycyl - L - Glutamine Monohydrate by Precolumn Derivatization RP - HPLC

目的:用反相高效液相色谱法测定N-甘氨酰-L-谷氨酰胺一水合物含量。方法:以2,4-二硝基氟苯作柱前衍生化试剂,在碱性条件下制得2,4-二硝基苯基-N-甘氨酰-L-谷氨酰胺,上ODS色谱柱,置日本岛津SCL-10AVP高效液相色谱仪上进行分析。流动相为0.05mol·L^(-1)醋酸钠水溶液(用冰醋酸调pH6.4)-乙腈(80:20),检测波长为360nm,进样量20μL,流速1mL·min^(-1)。结果:N-甘氨酰-L-谷氨酰胺-水合物在16～80μg·mL^(-1)范围内线性良好,r=0.994 3,回归方程Y=1.2436×10^(-4)X,重现性RSD=0.31％(n=5)。结论:本法可用于测定N-甘氨酰-L-谷氨酸胺-水合物的含量。

Objective: To establish a RP - HPLC method to determine N - glycyl - L - glutamine monohydrate (GGM)with 2,4 -dinitrofluorobenzene(DNFB)as a precolumn derivative regent. Methods:2,4 - Dinitrophenyl -N - glycyl - L - glutamine monohydrate ( DNP - GGM ) was made from the mixture of DNFB with GGM on the alkaline condition at 80℃. Then the reaction solution was analyzed by Shimadzu SCL - 10AVP HPLC instrument on ODS column with a flow phase of acetonitrile - aqueous solution of 0 . 05 mol @@L-1 sodium acetate , pH 6 . 4 (20: 80). Detection wavelength was 360 nm. Flow rate was 1. 0 mL·min-1. Results:The derivative DNP - GGM could be separated from 2,4 - dinitrophenol which was the hydrolysate of DNFB on an ODS column within 10 minutes. There was a good linear correlation in the range of 16- 80μg·mL-1, r = 0. 994 3 , Y = 1. 243 6×10^(-4)X, RSD =0. 31%(n=5). Conclusion: This method was the best assay to determine GGM.