Quantitative determination and validation of 17 cannabinoids in cannabis and hemp using liquid chromatography-tandem mass spectrometry

Abstract

The increase in production of cannabis for medical and recreational purposes in recent years has led to a corresponding increase in laboratories performing cannabinoid analysis of cannabis and hemp. We have developed and validated a quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) method that is simple, reliable, specific, and accurate for the analysis of 17 cannabinoids in cannabis and hemp. Liquid-solid sample extraction coupled with dilution into a calibration range from 10 to 10,000 ng/mL and LC-MS/MS analysis provides quantification of samples ranging from 0.002 to 200 mg/g (0.0002 to 20.0%) in matrix. Linearity of calibration curves in methanol was demonstrated with regression r² ≥ 0.99. Within-batch precision (0.5 to 6.5%) and accuracy (91.4 to 108.0%) and between-batch precision (0.9 to 5.1%) and accuracy (91.5 to 107.5%) were demonstrated for quality control (QC) samples in methanol. Within-batch precision (0.2 to 3.6%) and accuracy (85.4 to 111.6%) and between-batch precision (1.4 to 6.1 %) and accuracy (90.2 to 110.3%) were also evaluated with a candidate cannabis certified reference material (CRM). Repeatability (1.5 to 12.4% RSD) and intermediate precision (2.2 to 12.8% RSD) were demonstrated via analysis of seven cannabis samples with HorRat values ranging from 0.3 to 3.1. The method provides enhanced detection limits coupled with a large quantitative range for 17 cannabinoids in plant material. It is suitable for a wide range of applications including routine analysis for delta-9-tetrahydrocannabinol (Δ⁹-THC), delta-9-tetrahydrocannabinolic acid (Δ⁹-THCA), cannabidiol (CBD), cannabidiolic acid (CBDA), and cannabinol (CBN) as well as more advanced interrogation of samples for both major and minor cannabinoids.

Keywords: Cannabinoids; Cannabis; Hemp; Liquid chromatography-mass spectrometry; Validation.

Fig. 1

Chemical structures of the 17 cannabinoids targeted in the reported method

Fig. 2

LC-MS/MS chromatogram of 17 cannabinoids in a calibration standard at a concentration of 1000 ng/mL for each cannabinoid

Fig. 3

LC-MS/MS chromatogram of 17 cannabinoids in a cannabis sample extracted with methanol:water and diluted 1/100 with methanol. The responses of minor cannabinoids CBDV, CBDVA, THCV, and THCVA have been enhanced 5- to 20-fold while the responses of CBDA and Δ⁹-THCA have been reduced 5-fold to allow visualization of all peaks in the chromatogram

Fig. 4

LC-MS/MS chromatogram of 17 cannabinoids in a hemp sample extracted with methanol:water and diluted 1/10 with methanol. The responses of CBD and Δ⁹-THC have been reduced 2-fold while the responses of CBDA and Δ⁹-THCA have been reduced 10-fold to allow visualization of all peaks in the chromatogram

Fig. 5

Cumulative percent recovery of a subset of cannabinoids after 1 to 5 serial extractions. Two serial extractions provide recovery ≥ 98.75% while four serial extractions provide recovery ≥ 99.97%