高效液相色谱法检测发酵液中四氢嘧啶及羟基四氢嘧啶  

Determination of ectoine and hydroxyectoine in fermentation broth by high performance liquid chromatography

作　　者：刘紫寒 郭秋爽 周超 孙杨 李华[1,2] 刘宇鹏 Zihan Liu;Qiushuang Guo;Chao Zhou;Yang Sun;Hua Li;Yupeng Liu(Institute of Microbial Engineering,School of Life Sciences,Henan University,Kaifeng,Henan 475004,China;Engineering Research Center for Applied Microbiology of Henan Province,Kaifeng,Henan 475004,China)

机构地区：[1]河南大学生命科学学院微生物工程研究所,河南开封475004 [2]河南省应用微生物工程研究中心,河南开封475004

出　　处：《日用化学工业（中英文）》2022年第11期1236-1240,共5页China Surfactant Detergent & Cosmetics

基　　金：河南省自然科学基金资助项目(202300410055);河南省重点研发与推广专项(No.192102110179)。

摘　　要：为实现发酵液样品中四氢嘧啶和羟基四氢嘧啶的分离检测,使用高效液相色谱(HPLC)技术建立一种测定中度嗜盐菌发酵液中四氢嘧啶及羟基四氢嘧啶含量的方法。使用乙醇抽提法提取样品,采用Vertex^(TM) NH_(2)(4.6×250 mm,5μm)液相色谱柱对中度嗜盐菌发酵液中四氢嘧啶及四氢嘧啶进行了分析,选用紫外(UV)检测器,对比不同检测条件。通过色谱条件优化,选定了70%乙腈作为流动相,柱温30℃,检测波长210 nm,流速0.8 mL/min。发酵液样品中四氢嘧啶和羟基四氢嘧啶在各自范围内线性关系良好,两种样品的平均回收率在99.2%~102%,精密度偏差在1%~2%范围内,检出限为1.5μg/mL,定量限为5μg/mL,分离度为1.70。该方法检测范围大,具有较好的重复性、分离度及稳定性,适用于发酵液中四氢嘧啶及羟基四氢嘧啶的分离检测。A method was developed for the determination of ectoine and hydroxyectoine in fermentation broth samples of moderately saline bacteria using high performance liquid chromatography(HPLC).Samples were extracted using ethanol extraction,and ectoine and hydroxyectoine standards were dissolved and diluted in ultrapure water.Vertex^(TM) NH_(2)(4.6×250 mm,5μm)liquid chromatographic column was used to analyse ectoine and hydroxyectoine in the fermentation broth of moderately halophilic bacteria.Waters 1515 high performance liquid chromatograph with ultraviolet(UV)detector was selected to compare the different detection conditions.The chromatographic conditions were optimized by using acetonitrile(70%)as the mobile phase with a column temperature of 30℃,a detection wavelength of 210 nm and a flow rate of 0.8 mL/min.The linearity of ectoine and hydroxyectoine in the fermentation broth samples is good in their respective ranges.The average recoveries of the two samples are in the range of 99.2%to 102%,the precision deviations are in the range of 1%to 2%,the limit of detection is 1.5μg/mL,the limit of quantification is 5μg/mL and the separation degree is 1.70.The method is suitable for the separation and determination of ectoine and hydroxyectoine in fermentation broth and for the quantitative analysis of ectoine and hydroxyectoine under laboratory conditions.

关 键 词：中度嗜盐菌 高效液相色谱 四氢嘧啶 羟基四氢嘧啶 

分 类 号：TQ658[化学工程—精细化工]