测定生物样本中神经酰胺含量的HPLC-FLD法  

HPLC-FLD determination of ceramide in biological samples

作　　者：娄婷婷[1] 李英娴[1] 方步武[1] 刘文东[1] 董伟林[1] 张庆伟[1] 薛春兰[1] 

机构地区：[1]天津医科大学,天津300070

出　　处：《药物分析杂志》2012年第10期1807-1812,共6页Chinese Journal of Pharmaceutical Analysis

基　　金：国家自然科学基金项目(No.30772856)

摘　　要：目的:建立高效液相色谱-荧光检测器分析生物样本中的神经酰胺。方法:神经酰胺及预处理样本在1 mol·L-1氢氧化钾乙醇溶液中沸水浴反应2 h脱酰基,再与邻苯二甲醛衍生试剂反应60～90 s,采用Kromasil C18(250 mm×4.5 mm,5μm)色谱柱,流动相为甲醇-0.02 mol·L-1磷酸二氢钾溶液(90∶10),流速1.0 mL·min-1,荧光检测器的激发波长和吸收波长分别为340 nm和455 nm,进样量50μL。结果:神经酰胺浓度在0.05～6.4μg·mL-1范围内与衍生物峰面积呈良好的线性关系(r=0.9999),回收率为93.2%～107.5%,RSD<7.6%。结论:本方法具有快速、准确、灵敏度高、稳定性好等优点,适用于生物样品中微量神经酰胺的检测。Objective：To establish an HPLC method for determination of ceramide in biological samples.Methods：Ceramide was reacted with preprocessed samples in 1 mol·L-1 KOH ethanol solution to be deacetylated by boiling water bath for 2 h,then reacted with derivatizing reagent o-phthalaldehyde for 60-90 s,with a sample size of 50 μL.The analysis was performed with Kromasil C18column（250 mm×4.5 mm,5 μm）.The mobile phase was methanol-0.02 mol·L-1 potassium dihydrogen phosphate solution（90∶ 10）at a flow rate of 1.0 mL·min-1,and the excitation wavelength and emission wavelength were set at 340 nm and 455 nm respectively.Results：The linear range of ceramide was 0.05-6.4 μg·mL-1（r=0.9999）,and the average recovery was 93.2%-107.5%.Conclusion：The method has a series of advantages,as it is fast,precise,sensitive and reproducible.It is suitable for the trace ceramide detection in biological samples.

关 键 词：生物样本 神经酰胺 柱前衍生化 高效液相色谱法 荧光检测 微量测定 质谱分析 

分 类 号：R917[医药卫生—药物分析学]