Determination of 11 Cannabinoids in Biomass and Extracts of Different Varieties of Cannabis Using High-Performance Liquid Chromatography

Abstract

An HPLC single-laboratory validation was performed for the detection and quantification of the 11 major cannabinoids in most cannabis varieties, namely, cannabidiolic acid (CBDA), cannabigerolic acid (CBGA), cannabigerol (CBG), cannabidiol (CBD), tetrahydrocannabivarin (THCV), cannabinol (CBN), Δ9-trans-tetrahydrocannabinol (Δ9-THC), Δ8-trans-tetrahydrocannabinol (Δ8-THC), cannabicyclol (CBL), cannabichromene (CBC), and Δ9-tetrahydrocannabinolic acid-A (THCAA). The analysis was carried out on the biomass and extracts of these varieties. Methanol-chloroform (9:1, v/v) was used for extraction, 4-androstene-3,17-dione was used as the internal standard, and separation was achieved in 22.2 min on a C18 column using a two- step gradient elution. The method was validated for the 11 cannabinoids. The concentration-response relationship of the method indicated a linear relationship between the concentration and peak area with r2 values of >0.99 for all 11 cannabinoids. Method accuracy was determined through a spike study, and recovery ranged from 89.7 to 105.5% with an RSD of 0.19 to 6.32% for CBDA, CBD, THCV, CBN, Δ9-THC, CBL, CBC, and THCAA; recovery was 84.7, 84.2, and 67.7% for the minor constituents, CBGA, CBG, and Δ8-THC, respectively, with an RSD of 2.58 to 4.96%. The validated method is simple, sensitive, and reproducible and is therefore suitable for the detection and quantification of these cannabinoids in different types of cannabis plant materials.

A review on the recent advances in HPLC, UHPLC and UPLC analyses of naturally occurring cannabinoids (2010–2019)

“…Various HPLC and UPLC methods that have been reported for the analysis of cannabinoids in C. sativa L. plant samples and Cannabis consumer products, e.g. hashish, marijuana and cannabis oils, since the year 2010, are summarised in Tables and , and appraised in the following subsections.…”

Chromatographic characterisation of 11 phytocannabinoids: Quantitative and fit‐to‐purpose performance as a function of extra‐column variance

“…For example, many published methods have the notable absence of Δ 8 ‐THC, which is likely avoided due to the chromatographic difficulties associated with separating it from Δ 9 ‐THC. Achieving resolution of these compounds requires long run times (17–36 min) that impact throughput . Further complicating these analyses are the presence of acidic and neutral cannabinoids that are integral to the biological activity of cannabis, but generally have different requirements to achieve adequate chromatographic resolution.…”

“…extracts, edibles, blood, hair, urine), including methods for potency testing. Most of these methods can be generally divided into those using chromatographic separation with optical [7][8][9][10][11][12] or mass spectrometry (MS) detection. 11,[13][14][15][16] Despite the abundance of analytical methods, challenges persist.…”

“…Achieving resolution of these compounds requires long run times (17-36 min) that impact throughput. 7,8,14 Further complicating these analyses are the presence of acidic and neutral cannabinoids that are integral to the FIGURE 1 Structures of the phytocannabinoids used in the current study and some of their related derivatives present in lower abundance in cannabis. Known pathways for formation are shown, with those products showing Δ, [O] generally formed through combustion.…”

Leaner and greener analysis of cannabinoids

“…The solvent composition, mass to solvent ratio, extraction technique and time vary considerably between methods. Separations of cannabinoids use different mobile phases, columns and gradients, and given the number of minor cannabinoids present in authentic materials, there is a possibility for co-elution of peaks and inaccurate quantitative results [11, 12]. Rigorous validation procedures are necessary to ensure that the results of any analytical method are reliable.…”