Quantitative analysis of Δ8- and Δ9-tetrahydrocannabinol metabolites and isomers: a rapid assay in urine by LC-MS/MS

An increasing number of cannabis-related products have become available and entered the market, particularly those containing cannabidiol (CBD) and Delta 8-tetrahydrocannabinol (Delta 8-THC). Analytical methods for cannabinoids in urine have been described extensively in the literature. However, methods providing good resolution for distinguishing interferences from THC positional isomers are needed. The aim of this project was to develop and validate a liquid chromatography with tandem mass spectrometry (LC-MS/MS) method to quantitate a broad panel of cannabinoids in authentic urine specimens. The method was optimized to quantitate Delta 8-THC and Delta 9-THC, 11-OH-Delta 8-THC and 11-OH-Delta 9-THC, Delta 8-THC-COOH and Delta 9-THC-COOH, CBD, 7-COOH-CBD, CBG, and CBN, and validated with the guidance of the American Academy of Forensic Sciences Standards Board (ASB) Standard 036. The validated assay was then used to evaluate urine samples collected over various time points from female patients (N = 69) enrolled in a study assessing prevalence of marijuana/CBD use during pregnancy from November 2022 to May 2024. Delta 8- and Delta 9- isomers were chromatographically resolved and successfully separated. For all analytes, the lower limit of quantitation (LLOQ) was determined to be 10 ng/mL, and the upper limit of quantitation (ULOQ) was 1000 ng/mL. In the authentic samples, the most frequently detected analyte was Delta 9-THC-COOH, with a median concentration of 278 ng/mL (n = 38). Delta 9-THC and 11-OH-Delta 9-THC were detected with a median concentration of 42.4 ng/mL (n = 5) and 65.7 ng/mL (n = 34), respectively. Delta 8-THC-COOH was detected in n = 3 specimens, with a median concentration of 25.5 ng/mL. The study provided a rapid assay for the analysis of cannabinoids in urine.