柱前衍生-高效液相色谱法检测魔芋飞粉中的神经酰胺  

Determination of Ceramide in Konjac Flying Powder by Precolumn Derivation-High Performance Liquid Chromatography

作　　者：张新月 李玖江 李玥[3] 陈尚卫 朱松 ZHANG Xinyue;LI Jiujiang;LI Yue;CHEN Shangwei;ZHU Song(State Key Laboratory of Food Science and Resources,Jiangnan University,Wuxi 214122 Jiangsu;Sichuan Hanchen Biologic Technology Co.Ltd,Leshan 620036 Sichuan;School of Food Science and Technology,Jiangnan University,Wuxi 214122)

机构地区：[1]江南大学食品科学与资源挖掘全国重点实验室,江苏无锡214122 [2]四川晗晨生物科技有限公司,四川眉山620036 [3]江南大学食品学院,江苏无锡214122

出　　处：《分析科学学报》2024年第3期309-314,共6页Journal of Analytical Science

基　　金：国家自然科学基金项目(No.32172197)。

摘　　要：建立了一种柱前衍生-高效液相色谱测定魔芋飞粉中神经酰胺的方法。将神经酰胺与氢氧化钾乙醇溶液在高温条件下进行脱酰基反应,再与9-氯甲酸芴甲酯(FMOC-Cl)衍生化试剂反应,高效液相色谱分离后,采用紫外或荧光检测器检测。考察并确定了脱酰基以及衍生化反应的最佳条件,在110℃条件下用0.5 mol/L氢氧化钾乙醇溶液对神经酰胺标品或样品处理1.5 h,然后用10 mg/mL的FOMC-Cl对其进行在线衍生。采用Waters Symmetry C18柱(250 mm×4.6 mm, 5μm)分离,甲醇/水/磷酸(体积比10/90/0.05)-甲醇为流动相进行梯度洗脱,荧光检测器的激发波长和吸收波长分别为266 nm和305 nm;紫外检测器波长为262 nm。在神经酰胺浓度为20~500μg/mL范围内方法的线性关系良好,其中紫外检测器检测的相关系数(R^(2))为0.9978,荧光检测器检测的R2为0.9978,回收率为97.3%~103.6%,相对标准偏差<1.64%。所建立方法采用紫外和荧光检测器测定目标物的检测限分别为0.48μg/mL和0.0082μg/mL,定量限分别为1.6μg/mL和0.027μg/mL。An analytical method for the determination of ceramide in konjac flying powder by precolumn derivatization-high performance liquid chromatography(HPLC)was developed.Ceramide was deacylated with KOH ethanol solution at high temperature for a period of time,and then reacted with 9-fluorenylmethyl chloroformate(FMOC-Cl).The optimum conditions for deacylation and derivatization were determined as follows:Ceramide was treated with 0.5 mol/L KOH ethanol solution at 110℃for 1.5 h;then derived online by 10 mg/mL FOMC-Cl.The separation was performed with Waters Symmetry C18 column(250 mm×4.6 mm,5μm).Water/methanol/phosphoric acid(90/10/0.05,v/v/v)-methanol was used as mobile phase for gradient elution.The linear range of ceramide was 20-500μg/ml(UV R^(2)=0.9978;FLD R^(2)=0.9978),and the average recovery was 97.3%-103.6%with relative standard deviations(RSD)less than 1.64%.The limits of detection(LOD)and quantitation(LOQ)were 0.48μg/mL(UV)and 0.0082μg/mL(FLD),and 1.6μg/mL(UV)and 0.027μg/mL(FLD).

关 键 词：神经酰胺 脱酰基 在线衍生 高效液相色谱 9-氯甲酸芴甲酯 

分 类 号：O657.72[理学—分析化学]