高效液相色谱-串联质谱法测定人血清中文拉法辛和O-去甲基文拉法辛浓度的不确定度评价

目的：评价高效液相串联质谱（Liquid chromatography tandem mass spectrometry，LC-MS/MS）法测定人血清中文拉法辛和O-去甲基文拉法辛浓度的不确定度。方法：对可能会引入不确定度的步骤进行分析，包括测量重复性、样品称量、溶液配制、样品处理、仪器允差、基质效应、提取回收、标准曲线拟合等，评估扩展不确定度。结果：文拉法辛低（12 ng·mL^-1）、中（120 ng·mL^-1）、高（300 ng·mL^-1）浓度质控样品的拓展不确定度分别为：U_L=0.847 ng·mL^-1，U_M=7.518 ng·mL^-1，U_H=20.776 ng·mL^-1；O-去甲基文拉法辛低（60 ng·mL^-1）、中（600 ng·mL^-1）、高（1 500 ng·mL^-1）浓度质控样品的拓展不确定度分别为：U_L=11.666 ng·mL^-1，U_M=91.479 ng·mL^-1，U_H=254.523 ng·mL^-1（P=95%，k=2）。结论：LC-MS/MS法测定人血清中文拉法辛和O-去甲基文拉法辛浓度的不确定度主要来源于标准曲线拟合、基质效应和提取回收过程。选择合适的同位素氘代内标浓度能够有效降低标准曲线拟合过程引入的不确定度。

Uncertainty evaluationin the determination of venlafaxine and O-desmethylvenlafaxine in human serum by HPLC-MS/MS

OBJECTIVE To evaluate the uncertainty of high performance liquid chromatography tandem mass spectrometry (LC-MS/MS) method for the determination of Chinese venlafaxine and O-desmethylvenlafaxine in human serum.METHODS Possible factors with regard to the uncertainty were analyzed, including weighing of standard substances, preparation of standard solutions, preparation of serum samples, tolerance of instruments, repeatability, matrix effect, extraction recovery and standard curve fitting, to assess the expanded uncertainty. RESULTS The expanded uncertainty of venlafaxine were 0.847 ng·mL^-1, 7.518 ng·mL^-1 and 20.775 ng·mL^-1at low (12 ng·mL^-1), middle (120 ng·mL^-1) and high (300 ng·mL^-1) concentrations, respectively; for O-desmethylvenlafaxine, the expanded uncertainty were 11.666 ng·mL^-1,91.479 ng·mL^-1,254.523ng·mL^-1 at low (60 ng·mL^-1), middle (600 ng·mL^-1) and high (1 500 ng·mL^-1) concentrations respectively (P=95%, k=2).CONCLUSION The uncertainty of LC-MS/MS determination of Chinese venlafaxine and O-desmethylvenlafaxine in human serum was mainly related to calibration curve fitting, matrix effect and extraction recovery. The uncertainty introduced by the standard curve fitting process can be effectively reduced by selecting the appropriate concentration of the isotope deuterated internal standard.