Cannabidiol, ∆9-tetrahydrocannabinol, and metabolites in human blood by volumetric absorptive microsampling and LC-MS/MS following controlled administration in epilepsy patients

Federica Pigliasco; Sara Malaca; Alfredo Fabrizio Lo Faro; Anastasio Tini; Giuliana Cangemi; Alessia Cafaro; Sebastiano Barco; Antonella Riva; Angelica Pisati; Elisabetta Amadori; Pasquale Striano; Adriano Tagliabracci; Marilyn Ann Huestis; Francesco Paolo Busardò

doi:10.3389/fphar.2022.1038754

Cannabidiol, ∆⁹-tetrahydrocannabinol, and metabolites in human blood by volumetric absorptive microsampling and LC-MS/MS following controlled administration in epilepsy patients

Front Pharmacol. 2022 Oct 24:13:1038754. doi: 10.3389/fphar.2022.1038754. eCollection 2022.

Authors

Federica Pigliasco¹, Sara Malaca², Alfredo Fabrizio Lo Faro², Anastasio Tini², Giuliana Cangemi³, Alessia Cafaro^{3

4}, Sebastiano Barco³, Antonella Riva¹, Angelica Pisati¹, Elisabetta Amadori¹, Pasquale Striano^{1

5}, Adriano Tagliabracci², Marilyn Ann Huestis⁶, Francesco Paolo Busardò²

Affiliations

¹ Department of Neurosciences, Rehabilitation, Ophthalmology, Genetics, Maternal and Child Health (DINOGMI), University of Genoa, Genoa, Italy.
² Department of Excellence-Biomedical Sciences and Public Health, Università Politecnica delle Marche, Ancona, Italy.
³ Chromatography and Mass Spectrometry Section, Central Laboratory of Analyses, IRCCS Istituto Giannina Gaslini, Genoa, Italy.
⁴ Department of Internal Medicine, Pharmacology & Toxicology Unit, University of Genoa, Genoa, Italy.
⁵ Paediatric Neurology and Muscular Disease Unit, IRCCS Istituto Giannina Gaslini, Genoa, Italy.
⁶ Institute of Emerging Health Professions, Thomas Jefferson University, Philadelphia, PA, United States.

Abstract

Cannabidiol (CBD) exhibits anti-inflammatory, anxiolytic, antiseizure, and neuroprotective proprieties without addictive or psychotropic side effects, as opposed to Δ⁹-tetrahydrocannabinol (THC). While recreational cannabis contains higher THC and lower CBD concentrations, medical cannabis contains THC and CBD in different ratios, along with minor phytocannabinoids, terpenes, flavonoids and other chemicals. A volumetric absorptive microsampling (VAMS) method combined with ultra-high-performance liquid chromatography coupled with mass spectrometry in tandem for quantification of CBD, THC and their respective metabolites: cannabidiol-7-oic acid (7-COOH-CBD); 7-hydroxy-cannabidiol (7-OH-CBD); 6-alpha-hydroxy-cannabidiol (6-α-OH-CBD); and 6-beta-hydroxycannabidiol (6-β-OH-CBD); 11- Hydroxy-Δ⁹-tetrahydrocannabinol (11-OH-THC) and 11-Nor-9-carboxy-Δ⁹-tetrahydrocannabinol (THCCOOH). After overnight enzymatic glucuronide hydrolysis at 37°C, samples underwent acidic along with basic liquid-liquid extraction with hexane: ethyl acetate (9:1, v/v). Chromatographic separation was carried out on a C18 column, with the mass spectrometer operated in multiple reaction monitoring mode and negative electrospray ionization. Seven patients with intractable epilepsy were dosed with various CBD-containing formulations and blood collected just before their daily morning administration. The method was validated following international guidelines in toxicology. Linear ranges were (ng/ml) 0.5-25 THC, 11-OH-THC, THCCOOH, 6-α-OH-CBD and 6-β-OH-CBD; 10-500 CBD and 7-OH-CBD; and 20-5000 7-COOH-CBD. 7-COOH-CBD was present in the highest concentrations, followed by 7-OH-CBD and CBD. This analytical method is useful for investigating CBD, THC and their major metabolites in epilepsy patients treated with CBD preparations employing a minimally invasive microsampling technique requiring only 30 µL blood.

Keywords: CBD metabolites; UHPLC-MS/MS; cannabinoids; medical cannabis; serum.