Enzymatic hydrolysis pre-column derivatisation UPLC method for the detection of asparagine and glutamine in FSMP
FAN Zhiying, WANG Qi, WU Yichun, LUO Min, XIAO Weimin, BAO Yuechao, LAN Xiong, CHEN Jiaping, YANG Guowu, YANG Yan
1. Baotou Medical College, Inner Mongolia University of Science and Technology, Baotou 014040, China;
2. Shenzhen Academy of Metrology & Quality Inspection, Shenzhen 518109, China;
3. School of Public Health (Shenzhen), Sun Yat-sen University, Shenzhen 518107, China
Abstract: A high performance liquid chromatography method was established for the detection of asparagine (Asn) and glutamine (Gln) in special medical purpose food (FSMP) using enzyme hydrolysis-dansyl chloride pre-column derivatization. After the sample was enzymatically hydrolyzed by streptomyces griseus protease (Pronase E), the supernatant was collected and subjected to pre-column derivatization with Dns-Cl. The derivatized product was applied to an Agilent Zorbax SB-C18 chromatography column (250 mm × 4.6 mm，5 μm).The separation was performed using a 10 mM solution of disodium hydrogen phosphate (pH 6.5) and methanol as the mobile phase for gradient elution and quantification using an external standard method. The results indicate that the two amino acids are present at a concentration of 0.500-50.0μg/mL. The linearity is good within the concentration range of g/mL (R≥0.999), the recovery rate is 95.3%-105% and the RSD is 0.95%-3.22%. The method was validated for enzymatic digestion efficiency and measured content using two protein standards, with good accuracy (δ<10%).The method fills the gap in detection methods for Asn and Gln in FSMP, can be used to detect Asn and Gln content in FSMP, and is currently being used to develop national food safety standards.
Keywords: asparagine;glutamine;food for special medical purposes;dansulfonyl chloride;streptomyces griseus protease
2023, 49(11):127-131  收稿日期: 2023-04-29;收到修改稿日期: 2023-06-25
基金项目: 国家重点研发计划科研项目(2021YFF0701900，2021YFF0701903）
作者简介: 范志颖(1996-),女,内蒙古自治区通辽市人,研究生,专业方向为公共卫生。
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