柱前衍生化高效液相色谱法测定人血浆中花生四烯酸乙醇胺浓度

目的：建立血浆中花生四烯酸乙醇胺（Anandamide）的柱前衍生化高效液相色谱（HPLC）测定方法。方法：取血浆样品1mL碳酸氢钠碱化,无水甲苯提取,衍生化试剂DBD-COCL衍生,进样HPLC测定,采用外标法。色谱柱为Agilent C18（250×4.6mm,5μm）,流动相为50mmol.L^-1醋酸钠水溶液-乙腈,采用梯度洗脱,流速1.0ml.min^-1,激发波长为336nm,发射波长为560nm。结果：血浆中Anandamide线性关系良好,日内日间RSD〈10%,范围1-30ng/ml,血浆中花生四烯酸乙醇胺最低检测限为0.50-0.85ng（S/N=3）。结果：该方法样品处理过程简便,重复性好,符合生物样品测定要求,并适用于临床血样中anandamide浓度的测定。

Determination of Anandamide in human plasma using reversed-phase high performance Liquid Chromatography fluorescence detection with pre-column derivatization

Objective：A liquid-liquid extraction-based reversed-phase HPLC method with fuorometric detection was validated and applied for the analysis of anandamide in human plasma.Method：After alkalization with NaHCO3,plasma were extracted by toluene and analyzed using a high performance liquid chromatography equipped with a flurescence detector.The mobile phase consisted of two eluents,solvent A（acetonitrile） and solvent B（sodium acetate,pH 6.5）.The analyte was separated using gradient at a flow rate of 1 mL/min,spectrophotometric detection at 560nm with an excitation wavelength of 336nm. Results：The assay exhibited a good liner relationship with limits of detection ranging from 0.50 to 0.85ng（S/N=3）.The validated quantification range was 1-30ng·ml^-1.the intraday precision and interday precision were both less than 10%.Conclusion：The method is suitable to determine the concentration of Anandamide in human plasma.