Sempio Cristina, Scheidweiler Karl B, Barnes Allan J, Huestis Marilyn A
Chemistry and Drug Metabolism Section, IRP, National Institute on Drug Abuse, National Institutes of Health, Baltimore, MD, 21224, USA.
University of Maryland School of Medicine, Baltimore, MD, 21224, USA.
Drug Test Anal. 2018 Mar;10(3):518-529. doi: 10.1002/dta.2230. Epub 2017 Aug 16.
Prolonged urinary cannabinoid excretion in chronic frequent cannabis users confounds identification of recent cannabis intake that may be important in treatment, workplace, clinical, and forensic testing programs. In addition, differentiation of synthetic Δ9-tetrahydrocannabinol (THC) intake from cannabis plant products might be an important interpretive issue. THC, 11-hydroxy-THC (11-OH-THC) and 11-nor-9-carboxy-THC (THCCOOH) urine concentrations were evaluated during previous controlled cannabis administration studies following tandem alkaline/E. coli β-glucuronidase hydrolysis. We optimized recombinant β-glucuronidase enzymatic urinary hydrolysis before simultaneous liquid chromatography tandem mass spectrometry (LC-MS/MS) quantification of THC, 11-OH-THC, THCCOOH, cannabidiol (CBD), cannabinol (CBN), cannabigerol (CBG), tetrahydrocannabivarin (THCV) and 11-nor-9-carboxy-THCV (THCVCOOH) in urine. Enzyme amount, incubation time and temperature, buffer molarity and pH were optimized using pooled urine samples collected during a National Institute on Drug Abuse, Institutional Review Board-approved clinical study. Optimized cannabinoid hydrolysis with recombinant β-glucuronidase was achieved with 2000 IU enzyme, 2 M pH 6.8 sodium phosphate buffer, and 0.2 mL urine at 37°C for 16 h. The LC-MS/MS quantification method for hydrolyzed urinary cannabinoids was validated per the Scientific Working Group on Toxicology guidelines. Linear ranges were 1-250 μg/L for THC and CBG, 2-250 μg/L for 11-OH-THC, CBD, CBN, THCV and THCVCOOH, and 1-500 μg/L for THCCOOH. Inter-batch analytical bias was 92.4-112.4%, imprecision 4.4-9.3% CV (n = 25), extraction efficiency 44.3-97.1% and matrix effect -29.6 to 1.8% (n = 10). The method was utilized to analyze urine specimens collected during our controlled smoked, vaporized, and edible cannabis administration study to improve interpretation of urine cannabinoid test results.
慢性频繁使用大麻者尿液中大麻素排泄时间延长，这使得在治疗、工作场所、临床和法医检测项目中对近期大麻摄入情况的识别变得复杂，而这种识别可能很重要。此外，区分合成Δ9-四氢大麻酚（THC）摄入与大麻植物产品摄入可能是一个重要的解释问题。在之前的对照大麻给药研究中，采用串联碱性/大肠杆菌β-葡萄糖醛酸酶水解后，对尿液中THC、11-羟基-THC（11-OH-THC）和11-去甲-9-羧基-THC（THCCOOH）的浓度进行了评估。在通过液相色谱串联质谱法（LC-MS/MS）同时定量尿液中THC、11-OH-THC、THCCOOH、大麻二酚（CBD）、大麻酚（CBN）、大麻萜酚（CBG）、四氢大麻酚（THCV）和11-去甲-9-羧基-四氢大麻酚（THCVCOOH）之前，我们优化了重组β-葡萄糖醛酸酶的酶促尿液水解。使用在国立药物滥用研究所机构审查委员会批准的临床研究期间收集的混合尿液样本，对酶量、孵育时间和温度、缓冲液摩尔浓度和pH进行了优化。使用2000 IU酶、2 M pH 6.8的磷酸钠缓冲液和0.2 mL尿液，在37°C下孵育16小时，实现了用重组β-葡萄糖醛酸酶对大麻素的优化水解。根据毒理学科学工作组的指南，对水解后尿液中大麻素的LC-MS/MS定量方法进行了验证。THC和CBG的线性范围为1-250μg/L，11-OH-THC、CBD、CBN、THCV和THCVCOOH的线性范围为2-250μg/L，THCCOOH的线性范围为1-500μg/L。批间分析偏差为92.4-112.4%，不精密度为4.4-9.3%CV（n = 25），提取效率为44.3-97.1%，基质效应为-29.6至1.8%（n = 10）。该方法用于分析在我们的对照吸烟、雾化和食用大麻给药研究期间收集的尿液样本，以改进对尿液大麻素检测结果的解释。
