Development of a simple and sensitive HPLC–UV method for the simultaneous determination of cannabidiol and 9-tetrahydrocannabinol in rat plasma

Abstract

There has been increased interest in the medical use of cannabinoids in recent years, particularly in the predominant natural cannabinoids, cannabidiol (CBD) and 9-tetrahydrocannabinol (THC). The aim of the current study was to develop a sensitive and reliable method for the quantification of CBD and THC in rat plasma. A combination of protein precipitation using cold acetonitrile and liquid–liquid extraction using n hexane was utilised to extract CBD and THC from rat plasma. Samples were then evaporated and reconstituted in acetonitrile and 30 L was injected into an HPLC system. Separation was achieved using an ACE C18-PFP 150 mm × 4.6 mm, 3 m column at 55 ◦C with isocratic elution using a mobile phase consisting of acetonitrile–water (62:38, v/v) at 1 mL/min for 20 min. Both cannabinoids, as well as the internal standard (4,4-dichlorodiphenyltrichloroethane, DDT) were detected at 220 nm. Our new method showed linearity in the range of 10–10,000 ng/mL and a lower limit of quantifica tion (LLOQ) of 10 ng/mL for both cannabinoids, which is comparable to previously reported LC–MS/MS methods. Inter- and intra-day precision and accuracy were below 15% RSD and RE, respectively. To demonstrate the suitability ofthe method for in vivo studies in rats,the assay was applied to a preliminary pharmacokinetic study following IV bolus administration of 5 mg/kg CBD or THC. In conclusion, a simple, sensitive, and cost-efficient HPLC–UV method for the simultaneous determina tion of CBD and THC has been successfully developed, validated and applied to a pharmacokinetic study in rats.