Innovative development and validation of an HPLC/DAD method for the qualitative and quantitative determination of major cannabinoids in cannabis plant material

Abstract

GC is commonly used for the analysis of cannabis samples, e.g. in forensic chemistry. However, as this method is based on heating of the sample, acidic forms of cannabinoids are decarboxylated into their neutral counterparts. Conversely, HPLC permits the determination of the original composition of plant cannabinoids by direct analysis. Several HPLC methods have been described in the literature, but most of them failed to separate efficiently all the cannabinoids or were not validated according to general guidelines. By use of an innovative methodology for modelling chromatographic responses, a simple and accurate HPLC/DAD method was developed for the quantification of major neutral and acidic cannabinoids present in cannabis plant material: Delta9-tetrahydrocannabinol (THC), THC acid (THCA), cannabidiol (CBD), CBD acid (CBDA), cannabigerol (CBG), CBG acid (CBGA) and cannabinol (CBN). Delta8-Tetrahydrocannabinol (Delta8-THC) was determined qualitatively. Following the practice of design of experiments, predictive multilinear models were developed and used in order to find optimal chromatographic analytical conditions. The method was validated following an approach using accuracy profiles based on beta-expectation tolerance intervals for the total error measurement, and assessing the measurements uncertainty. This analytical method can be used for diverse applications, e.g. plant phenotype determination, evaluation of psychoactive potency and control of material quality.

Characterisation of cannabinoid composition in a diverse Cannabis sativa L. germplasm collection

“…Sample preparation and extraction followed those of De Backer et al (2009). Leaf material was dried at 35°C in a forced ventilation oven for 48 h. Seeds and stalks were separated and discarded manually and 1 g of dried leaf material was ground using a Mixer Mill MM 301 (Retsch GmbH) at 30 rotations/second for 30 s.…”

“…The pentyl-alkyl-cannabinoids THCA and cannabidiolic acid (CBDA) are the major constituents found in plants ( De Backer et al 2009;Swift et al 2013), although a series of propyl-alkyl-cannabinoid homologues [delta-9-tetrahydrocannabivarinic acid (THCVA) and cannabidivarinic acid (CBDVA)] also occur in plants from specific geographical regions (Baker et al 1980;Hillig and Mahlberg 2004). Pentyl-cannabinoids are formed from cannabigerolic acid (CBGA) (Taura et al 2007b), while propyl-cannabinoids are formed from cannabigerovarinic acid (CBGVA) (Shoyama et al 1984).…”

“…Gas chromatography is commonly used to characterise chemotype and relies on the thermal conversion of acidic to neutral cannabinoids, although this reaction can vary between laboratories and is only partial (Dussy et al 2005). Liquid chromatography methods developed more recently can detect both acidic and neutral cannabinoids, therefore providing a more precise characterisation of chemotype (De Backer et al 2009). Several DNA markers associated with the genes encoding THCA and/or CBDA synthase have been found beneficial in predicting chemotype during early stages of plant development (Kojoma et al 2006;Pacifico et al 2006;Rotherham and Harbison 2011;Staginnus et al 2014), with the most comprehensively studied in terms of genetic linkage and sample population screening being the dominant D589 (Staginnus et al 2014) and co-dominant B1080/B1192 (Pacifico et al 2006) DNA sequence characterised amplified region (SCAR) markers respectively.…”

A Reliable Method for the Separation and Detection of Synthetic Cannabinoids by Supercritical Fluid Chromatography with Mass Spectrometry, and Its Application to Plant Products

“…The analysis involves an immunoassay, GC-MS and LC-MS for the confirmation and quantification of the drugs. [9][10][11][12][13][14][15][16][17][18][19][20][21][22] The immunochemical method is generally used as a preliminary analysis because of its evident advantage in rapidity. However, the main drawback is the low selectivity of the target compound.…”

Contrast in coherent Raman scattering microscopy

“…In some strains of the Cannabis plant the local concentrations of THC and its precursor, ∆ 9 -tetrahydrocannabinolic acid (THCa), can reach up to 25% [113]. The presence and ratios of minority cannabinoids, such as cannabinol (CBN) and cannabidiol (CBD), can significantly alter the physiological and psychological effects of consuming the plant.…”