Qualitative and quantitative analysis of THC, 11-hydroxy-THC and 11-nor-9-carboxy-THC in whole blood by ultra-performance liquid chromatography/tandem mass spectrometry

A qualitative and quantitative analytical method was developed for the simultaneous determination of δ⁹-tetrahydrocannabinol (THC), 11-hydroxy-A9-tetrahydrocannabinol (11-OH-THC) and l1-nor-9-carboxy- δ⁹-tetrahydrocannabinol (THC-COOH) in whole blood. The samples were prepared by solid-phase extraction followed by ultra-performance liquid chromatography/tandem mass spectrometry (UPLC/MS/MS) analysis using positive ion electrospray ionization and multiple reaction monitoring. The chromatographic separation was performed with an Acquity UPLC^® HSS T3 (50 × 2.1 mm i.d., 1.8 μm) reversed-phase column using a methanol/2 mM ammonium formate (formic acid 0.1%) gradient in a total run time of 9.5 min. MS/MS detection was achieved with two precursor-product ion transitions per substance. The method was fully validated, including selectivity and capacity of identification, according to the identification criteria (two transitions per substance, signal-tonoise ratio, relative retention time and ion ratio) without the presence of interferences, limit of detection (0.2 μg/L for THC and 0.5 μg/L for 11-OH-THC and THC-COOH), limit of quantitation (0.5 μg/L for all cannabinoids), recovery (53-115%), carryover, matrix effect (34-43%), linearity (0.5-100 μg/L), intra-assay precision (CV < 10% for the relative peak area ratios and <0.1% for the relative retention time), inter-assay accuracy (mean relative error <10%) and precision (CV <11%). The method has already been successfully used in proficiency tests and subsequently applied to authentic samples in routine forensic analysis.