DETERMINATION OF PHOSPHATIDYLETANOL IN DRIED BLOOD SPOT BY UHPLC-MS/MS: EVALUATION OF ALCOHOL CONSUMPTION IN CHEMICAL DEPENDENTS

We developed and validated an UHPLC-MS/MS method for the quantification of phosphatidylethanol (PEth) in dried blood spots (DBS). Sample preparation was a liquid extraction and chromatographic separation was performed in an Acquity C8 column. The mobile phases were 4 mM ammonium acetate in water (A) and acetonitrile (B) at 24:76 (v/v). The total analytical run time was 6 minutes, with retention time of 3.36 min for PEth and 4.13 min for phosphatidylpropanol. The method was linear from 10 to 3000 ng mL-1, specific, with, precise and accurate. The analyte was stable in DBS stored from -20 to 45 °C for 21 days. Matrix effect was compensated with the internal standard (-5.21% to + 6.09%). The method was applied in the evaluation of alcohol consumption in DBS from 25 chemical dependents. The PEth concentrations ranged from 14.5 to 2380.8 ng mL-1, with significant correlation with the self-report alcohol consumption AUDIT scores (r=0.41).

本研究建立并验证了一种用于干血斑（DBS）中磷脂酰乙醇胺（PEth）定量分析的超高效液相色谱-串联质谱法（UHPLC-MS/MS）。样品前处理采用液萃取法，色谱分离于Acquity C8色谱柱上完成。流动相分别为含4 mM乙酸铵的水溶液（流动相A）与乙腈（流动相B），二者体积比为24:76。总分析运行时长为6分钟，磷脂酰乙醇胺的保留时间为3.36 min，磷脂酰丙醇的保留时间为4.13 min。该方法线性范围为10~3000 ng·mL⁻¹，专属性良好，精密度与准确度俱佳。分析物在-20℃至45℃条件下储存21天的干血斑中保持稳定。基质效应通过内标进行补偿，补偿区间为-5.21%至+6.09%。本方法应用于25名酒精依赖者的干血斑样本的酒精摄入评估。样本中磷脂酰乙醇胺的浓度范围为14.5~2380.8 ng·mL⁻¹，与自我报告的酒精摄入量及酒精使用障碍识别测试（AUDIT）评分呈显著相关（r=0.41）。