Development and Validation of an HPLC-UV/PDA Method for the Determination of Cannflavins in Different Cannabis sativa Chemovars.

Abstract

Cannabis sativa (C. sativa) is a psychoactive plant that has been used for millennia for medicinal, recreational, and industrial purposes. The main constituents of cannabis are the cannabinoids, with other constituents including terpenes and flavonoids that contribute to its bioactivity. Among the flavonoid class, there is a subclass, specific to cannabis, namely the cannflavins (A, B, and C), which are biologically active. This study is directed to the analysis of these constituents in various cannabis chemovars. In this study, an HPLC-PDA method was validated and applied to determine the content of cannflavins, namely, cannflavin A (CF-A), cannflavin B (CF-B), and cannflavin C (CF-C), in six different cannabis chemovars. The HPLC separation was achieved using a Luna^® C18 (150 × 4.6 mm × 3 μm) with isocratic elution using a mobile phase consisting of acetonitrile and water (65:35, v/v), both containing 0.1% formic acid at a flow rate of 1 mL/min, with the detector set at 342.4 nm. The method was validated according to the ICH guidelines and exhibited a linear relationship in the 5-500 ppm range with R² > 0.99. The method showed good recovery, ranging from 82% to 98%. The intra-day and inter-day relative standard deviations (% RSDs) were ≤5.29%. Consequently, the method was applied for the determination of all these cannflavins in the different cannabis chemovars. CF-A was the most abundant cannflavin in the examined samples (15.2-478.38 ppm). The method was shown to be simple, accurate, and selective.