Mahlke Nina Sophia, Ziesenitz Victoria, Mikus Gerd, Skopp Gisela
Institute of Legal Medicine and Traffic Medicine, University Hospital, Voss-Str. 2, 69115, Heidelberg, Germany.
Int J Legal Med. 2014 Sep;128(5):771-8. doi: 10.1007/s00414-014-1040-y. Epub 2014 Jul 6.
A rapid and sensitive liquid chromatography/tandem mass spectrometric (LC-MS/MS) method for simultaneous quantification of fentanyl (F), norfentanyl (NF), despropionylfentanyl (DPF), and hydroxynorfentanyl (OHNF) in human plasma and urine specimens has been developed and validated according to international guidelines. Analytes were extracted from 250-μL plasma or urine by liquid-liquid extraction. OHNF in urine affords a second extraction step and analysis with a different column. Calibration curves in plasma were linear from 0.05-10 ng/mL for F, 0.07-0.5 ng/mL for NF, 0.02-1.0 ng/ml for DPF, and 0.67-3.0 ng/mL for OHNF; in urine, from 0.09-10.0, 0.17-50, 0.08-1.0, and 1.0-5.0 ng/mL for F, NF, DPF, and OHNF, respectively. Analytical bias and intra- and inter-assay imprecision were within ± 15 % of target, except for OHNF in plasma and DPF in urine at the respective lower quality control level. All analytes were stable in processed samples when stored for 24 h at room temperature. Recoveries and process efficiencies were above 82.9 and 75.1 % for all analytes in plasma and urine. The low level of DPF in plasma indicated with a matrix effect of 71.3 % moderate ion suppression, all other analytes in plasma and urine showed no matrix effects. The lower limit of quantification (LOQ) in plasma was 0.05, 0.07, 0.02 and 0.67 ng/mL for F, NF, DPF, and OHNF, respectively. In urine, the LOQ of F, NF, DPF, and OHNF were 0.09, 0.17, 0.08, and 1.28 ng/mL, respectively. This assay has been applied to human specimens collected during a clinical drug-drug interaction study.
已根据国际指南开发并验证了一种快速灵敏的液相色谱/串联质谱（LC-MS/MS）方法，用于同时定量测定人血浆和尿液样本中的芬太尼（F）、去甲芬太尼（NF）、去丙酰芬太尼（DPF）和羟基去甲芬太尼（OHNF）。通过液液萃取从250μL血浆或尿液中提取分析物。尿液中的OHNF需要进行第二步萃取，并使用不同的色谱柱进行分析。血浆中的校准曲线在以下范围内呈线性：F为0.05 - 10 ng/mL，NF为0.07 - 0.5 ng/mL，DPF为0.02 - 1.0 ng/mL，OHNF为0.67 - 3.0 ng/mL；尿液中的校准曲线分别为：F为0.09 - 10.0 ng/mL，NF为0.17 - 50 ng/mL，DPF为0.08 - 1.0 ng/mL，OHNF为1.0 - 5.0 ng/mL。除了在各自较低质量控制水平下血浆中的OHNF和尿液中的DPF外，分析偏差以及批内和批间不精密度均在目标值的±15%以内。所有分析物在处理后的样本中于室温下保存24小时时均稳定。血浆和尿液中所有分析物的回收率和处理效率分别高于82.9%和75.1%。血浆中DPF含量较低，表明存在71.3%的中等离子抑制基质效应，血浆和尿液中的所有其他分析物均未显示基质效应。血浆中F、NF、DPF和OHNF的定量下限（LOQ）分别为0.05、0.07、0.02和0.67 ng/mL。在尿液中，F、NF、DPF和OHNF的LOQ分别为0.09、0.17、0.08和1.28 ng/mL。该测定法已应用于一项临床药物相互作用研究期间收集的人体样本。
