LC-MS/MS quantitation of non-psychotropic cannabinoid cannabidiol in aqueous humor



doi: 10.1016/j.jpba.2023.115324.


Online ahead of print.

Affiliations

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Alyssa S Aebersold et al.


J Pharm Biomed Anal.


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Abstract

Cannabidiol (CBD) is the most abundant non-psychotropic phytocannabinoid isolated from Cannabis sativa. To support preclinical studies of ocular pharmacology of CBD, a bioanalytical method was developed and validated for quantification of CBD in aqueous humor using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Aqueous humor samples were subjected to protein precipitation by acetonitrile, followed by chromatographic separation using reversed phase LC on a Raptor ARC-18 column with mobile phase A: 0.1 % (v/v) formic acid in water (B) 0.1 % formic acid in acetonitrile (B) as eluents. Detection was carried out with a triple quadrupole mass spectrometer with electrospray ionization operated in positive ion mode. Stable-isotope labeled CBD (CBD-d3) was used as internal standard. The total run time was 8 min. Quantification was accomplished within the validated concentration range of 0.5-500 ng/mL for CBD using a 5 µL sample. The lower limit of quantitation was 0.5 ng/mL. Inter- and intra-day precision is 4.737-7.620 % and 3.426-5.830 %, respectively. Inter- and intra-day accuracy ranged between 99.01 % and 100.2 % and 99.85-101.4 % respectively. The extraction recoveries were found to be 66.06 ± 5.146 %. The established method was successfully applied to investigate ocular pharmacokinetics of CBD in mice. Following intraperitoneal (i.p.) administration of 50 mg/kg CBD, its concentration reaches a Cmax of 71.55 ± 36.64 ng/mL in aqueous humor, with a Tmax of 2 h and a half-life of 1.046 h. The AUC was 183.4 ± 49.17 ng * h/mL. The development and validation of this LC-MS/MS method is an important step toward the goal of assessing the aqueous humor concentrations of CBD and correlating the concentrations of this phytocannabinoid with its ocular pharmacologic effects.


Keywords:

Aqueous humor; Cannabidiol; Liquid chromatography; Tandem mass spectrometry.

Conflict of interest statement

Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

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