Chemistry and Drug Metabolism, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, 251 Bayview Boulevard, Suite 200, Room 05A721, Baltimore, MD 21224, USA.
J Chromatogr A. 2013 Jul 5;1297:123-30. doi: 10.1016/j.chroma.2013.04.071. Epub 2013 Apr 30.
Δ(9)-Tetrahydrocannabinol (THC) is the primary target in oral fluid (OF) for detecting cannabis intake. However, additional biomarkers are needed to solve interpretation issues, such as the possibility of passive inhalation by identifying 11-nor-9-carboxy-THC (THCCOOH), and determining recent cannabis smoking by identifying cannabidiol (CBD) and/or cannabinol (CBN). We developed and comprehensively validated a microflow liquid chromatography (LC)-high resolution mass spectrometry method for simultaneous quantification of THC, THCCOOH, CBD and CBN in OF collected with the Oral-Eze(®) and Quantisal™ devices. One milliliter OF-buffer solution (0.25mL OF and 0.5mL of Oral-Eze buffer, 1:3 dilution, or 0.75mL Quantisal buffer, 1:4 dilution) had proteins precipitated, and the supernatant subjected to CEREX™ Polycrom™ THC solid-phase extraction (SPE). Microflow LC reverse-phase separation was achieved with a gradient mobile phase of 10mM ammonium acetate pH 6 and acetonitrile over 10min. We employed a Q Exactive high resolution mass spectrometer, with compounds identified and quantified by targeted-MSMS experiments. The assay was linear 0.5-50ng/mL for THC, CBD and CBN, and 15-500pg/mL for THCCOOH. Intra- and inter-day and total imprecision were <10.8%CV and bias 86.5-104.9%. Extraction efficiency was 52.4-109.2%, process efficiency 12.2-88.9% and matrix effect ranged from -86 to -6.9%. All analytes were stable for 24h at 5°C on the autosampler. The method was applied to authentic OF specimens collected with Quantisal and Oral-Eze devices. This method provides a rapid simultaneous quantification of THCCOOH and THC, CBD, CBN, with good selectivity and sensitivity, providing the opportunity to improve interpretation of cannabinoid OF results by eliminating the possibility of passive inhalation and providing markers of recent cannabis smoking.
Δ(9)-四氢大麻酚（THC）是检测大麻摄入的口腔液（OF）中的主要目标物。然而，需要额外的生物标志物来解决解释问题，例如通过鉴定 11-去甲-9-羧基-THC（THCCOOH）来确定被动吸入的可能性，并通过鉴定大麻二酚（CBD）和/或大麻酚（CBN）来确定最近吸食大麻的情况。我们开发并全面验证了一种微流液相色谱（LC）-高分辨质谱法，用于同时定量检测 OF 中收集的 THC、THCCOOH、CBD 和 CBN，这些 OF 是使用 Oral-Eze(®) 和 Quantisal™ 设备收集的。将 1 毫升 OF-缓冲溶液（0.25 毫升 OF 和 0.5 毫升 Oral-Eze 缓冲液，1:3 稀释，或 0.75 毫升 Quantisal 缓冲液，1:4 稀释）中的蛋白质沉淀，然后将上清液进行 CEREX™ Polycrom™ THC 固相萃取（SPE）。微流 LC 反相分离是通过梯度移动相 10mM 乙酸铵 pH6 和乙腈在 10min 内实现的。我们采用了 Q Exactive 高分辨质谱仪，通过靶向-MSMS 实验对化合物进行鉴定和定量。该测定法在 0.5-50ng/mL 范围内对 THC、CBD 和 CBN 呈线性，在 15-500pg/mL 范围内对 THCCOOH 呈线性。日内和日间以及总精密度均<10.8%CV，偏差为 86.5-104.9%。提取效率为 52.4-109.2%，过程效率为 12.2-88.9%，基质效应范围为-86 至-6.9%。所有分析物在 5°C 下在自动进样器上 24h 内稳定。该方法应用于使用 Quantisal 和 Oral-Eze 设备收集的真实 OF 标本。该方法提供了一种快速同时定量检测 THCCOOH 和 THC、CBD、CBN 的方法，具有良好的选择性和灵敏度，通过消除被动吸入的可能性并提供最近吸食大麻的标志物，为改善大麻素 OF 结果的解释提供了机会。
