Chromatographic interferences potentially inflating the levels of Δ9-THC in Cannabis Sativa plant samples and possible solutions

Researchers in the Chemical Sciences Division (CSD) at the National Institute of Standards and Technology (NIST) have been evaluating existing and developing new analytical methods for the differentiation of hemp and marijuana since the passage of the Agriculture Improvement Act of 2018. This legislation defined hemp as a Cannabis sativa plant containing 0.3 % or less Δ⁹-tetrahydrocannabinol (Δ⁹-THC) and removed hemp from the United States Drug Enforcement Agency controlled substances list. The United States Department of Agriculture later clarified that this threshold must be representative of the total Δ⁹-THC, which includes the Δ⁹-THC acidic precursor Δ⁹-tetrahydrocannabinolic acid (Δ⁹-THCA). As a result, the burden of making these distinctions fell to forensic and Cannabis testing laboratories. NIST CSD has previously demonstrated accurate and precise analytical measurements for Δ⁹-THC, Δ⁹-THCA, and nine other related cannabinoids in well-characterized samples from interlaboratory studies at NIST by liquid chromatography with photodiode array detection (LC-PDA) following a methanolic extraction. This publication expands this method for the first time to include 16 commercial hemp samples and 20 seized Cannabis samples, simulating the types of samples typically analyzed by forensic and Cannabis testing laboratories. The results presented here highlight chromatographic interferences from CBNA and synthetic Δ⁸-THC by-products for Δ⁹-THC that can inflate its mass fraction levels and lead to the misidentification of Cannabis plant samples as marijuana when they are actually hemp. Data collected for 7448 plant samples in 2023 by a Cannabis testing laboratory were used to demonstrate the prevalence of these chromatographic interferences for the first time in a large sample population. Examples are included to demonstrate the resolution of chromatographic interferences through modification of the chromatographic method, selective detection methods, and peak deconvolution.