In order to determine whether individuals were compliant with the marijuana abstinence instructions, urine samples were collected pre- (session 1) and post-5-day abstinence period (session 2). Collected samples were frozen immediately and shipped on ice to the Center for Human Toxicology at the University of Utah Health Sciences Center for quantitative THC metabolite analysis. A gas chromatograph-mass spectrometer using electron impact ionization (GC/MS EI) and selective ion monitoring (SIM) was used to quantitate levels of 11-nor-9-carboxy–Δ9-tetrahydrocannabinol (THC–COOH), the primary metabolite of marijuana. Briefly, a four-point standard curve (0, 7.5, 15 and 30 ng/ml) was generated for THC–COOH in drug-free urine. The curve was run in duplicate, quality control samples in triplicate and samples singly. Prior to extraction, 30 ng of internal standard (d3-?9 THC–COOH) was added to all samples, standards and controls. Quantitation was achieved by calculating the peak-area ratios of derivatized THC–COOH/internal standard. Observing a reduction in THC–COOH urine levels between the pre- and post-abstinence period was used to confirm abstinence from marijuana use during the 5-day abstinence period.
