柱前衍生化RP-HPLC法测定L-叔亮氨酸含量  

Determination of L-tert-leucine by reversed-phase high performance liquid chromatography with pre-column derivatization

作　　者：陈亭亭[1] 黄金[1] 陈蔚青[2] 王普[1] 

机构地区：[1]浙江工业大学药学院,浙江杭州310014 [2]浙江树人大学生物与环境工程学院,浙江杭州310015

出　　处：《浙江工业大学学报》2016年第4期431-434,共4页Journal of Zhejiang University of Technology

基　　金：浙江省公益性技术应用研究计划项目(2014C33274)

摘　　要：以2，3，4，6-四-O-乙酰基-β-D-吡喃葡萄糖异硫氰酸酯（GITC）为手性衍生化试剂，建立一种反相高效液相色谱测定L-叔亮氨酸的方法．以GITC为衍生化试剂对L-叔亮氨酸进行衍生化，优化衍’生化试剂用量和衍’生化时间等反应条件．使用岛津C18色谱柱（25Omm×4．6mm，5μm）分离衍生化产物，流动相为V（甲醇）：V（磷酸盐）-58：42的缓冲溶液（pH=2．8），流速为O．6mL／min，检测波长为254nm，柱温3O℃，进样量1O肛L，结果表明叔亮氨酸两个对映异构体的衍生化产物分离良好，在O．2～1．Ommo1／L范围内呈现良好的线性关系（r=O．9996）．该方法准确可靠，重现性好，适用于L一叔亮氨酸的含量测定．A pre-column derivatization-high performance liquid chromatographic(HPLC)method was established for the determination of L-tert-leucine(L-Tle)with 2,3,4,6-tetra-O-acetyl-β-Dglucopyranosyl isothiocyanate(GITC)as a chiral derivatization reagent.L-Tle was derivatized with GITC and the conditions of derivatization such as the quantity of GITC and the derivatized time were optimized.The resulting samples were analyzed by HPLC using an InterSustain C18column(250×4.6 mm,5μm).The mobile phase was a 58∶42 mixture of methanol and phosphate buffer(pH=2.8).The flow rate was 0.6mL/min,and the detection wavelength was254 nm.The column temperature was 30 ℃ and injection volume was 10μL.The obtained diastereomers of L-Tle were well separated,and had good linear relationship in the concentration range of 0.2~1.0mmol/L(r=0.999 6).This method is accurate,reliable and available for the determination of L-Tle.

关 键 词：柱前衍生化 反相高效液相色谱法 L-叔免氣酸 

分 类 号：Q939.97[生物学—微生物学]