In recent years, the surge in use of tetrahydrocannabinol (THC) and cannabidiol (CBD) has increased the need for sensitive and specific analytical assays to measure the said compounds in patients, to establish dose–effect relationships and to gain knowledge of their pharmacokinetics and metabolism. We developed and validated an online extraction high-performance liquid chromatography-tandem mass spectrometry (HPLC–MS-MS) method for simultaneous quantification of 17 cannabinoids and metabolites including THC and its metabolites, CBD and its metabolites and other minor cannabinoids in human plasma. CBD-glucuronide (CBD-gluc) standard was produced in-house by isolation of CBD-gluc from urine of patients using pure CBD oil. For calibration standards and quality control samples, human plasma was spiked with cannabinoids at varying concentrations within the working range of the respective compound and 200 µL of the plasma was extracted using a simple one-step protein precipitation procedure. The extracts were analyzed using online trapping LC/LC–atmospheric pressure chemical ionization–MS-MS running in the positive multiple reaction monitoring mode. The lower limit of quantification ranged from 0.78 to 7.8 ng/mL, and the upper limits of quantification were between 100 and 2,000 ng/mL. Inter-day analytical accuracy and imprecision ranged from 90.4% to 111% and from 3.1% to 17.4%, respectively. The analysis of plasma samples collected during clinical studies showed that (3R-trans)-cannabidiol-7-oic acid (7-CBD-COOH) was the major human metabolite with 5960% (59.6-fold) of CBD followed by 7-hydroxy-CBD (177%), CBD-gluc (157%) and 6α-hydroxy-CBD (39.8%); 6β-hydroxy-CBD was not detected in any of the samples. In the present study, we developed and validated a robust LC–MS-MS assay for the simultaneous quantification of cannabinoids and their metabolites, which has been used to measure >5,000 samples in clinical studies. Moreover, we were able to quantify CBD-gluc and showed that 7-CBD-COOH, 7-hydroxy-CBD and CBD-gluc are the major CBD metabolites in human plasma.

中文翻译：

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通过 LC-MS-MS 同时定量人血浆中的 17 种大麻素

近年来，四氢大麻酚 (THC) 和大麻二酚 (CBD) 的使用激增增加了对敏感和特异性分析测定的需求，以测量患者体内的上述化合物、建立剂量-效应关系并了解其药代动力学和代谢。我们开发并验证了一种在线提取高效液相色谱-串联质谱 (HPLC-MS-MS) 方法，用于同时定量 17 种大麻素和代谢物，包括 THC 及其代谢物、CBD 及其代谢物和人血浆中的其他少量大麻素。CBD-葡萄糖苷酸 (CBD-gluc) 标准品是通过使用纯 CBD 油从患者尿液中分离 CBD-gluc 内部生产的。对于校准标准和质量控制样品，在相应化合物的工作范围内，人血浆中加入了不同浓度的大麻素，并使用简单的一步蛋白质沉淀程序提取了 200 µL 血浆。使用在线捕集 LC/LC-大气压化学电离-MS-MS 分析提取物，在正多反应监测模式下运行。定量下限为 0.78 至 7.8 ng/mL，定量上限为 100 至 2,000 ng/mL。日间分析准确度和不精密度分别为 90.4% 至 111% 和 3.1% 至 17.4%。对临床研究期间收集的血浆样本的分析表明，(3R-trans)-cannabidiol-7-oic acid (7-CBD-COOH) 是主要的人体代谢物，其中 CBD 占 5960%（59.6 倍），其次是 7-羟基-CBD (177%), CBD-gluc (157%) 和 6α-羟基-CBD (39.8%)；在任何样品中均未检测到 6β-羟基-CBD。在本研究中，我们开发并验证了一种稳健的 LC-MS-MS 测定法，用于同时定量大麻素及其代谢物，该测定法已用于在临床研究中测量超过 5,000 个样品。此外，我们能够量化 CBD-gluc 并表明 7-CBD-COOH、7-羟基-CBD 和 CBD-gluc 是人体血浆中的主要 CBD 代谢物。