Determination of 11-nor-delta9-tetrahydrocannabinol-9-carboxylic acid in urine using high-performance liquid chromatography and electrospray ionization mass spectrometry.

High-performance liquid chromatography with electrospray ionization mass spectrometry was used to determine 11-nor-delta9-tetrahydrocannabinol-9-carboxylic acid (THC-COOH) in urine. After basic hydrolysis of conjugates, the compound was extracted using SPEC-PLUS-3ML-C18 solid-phase extraction columns. A deuterium labelled internal standard (d3-THC-COOH) was added prior to hydrolysis. Separation was performed on a reversed-phase Zorbax Eclipse XDB-C8 analytical column (150x3.0 mm I.D.) using a gradient program from 60 to 80% acetonitrile (4 mM formic acid) at a flow-rate of 0.5 ml/min. The compounds were detected by single ion monitoring of m/z 345 and m/z 348 for the protonated molecules [THC-COOH+H]+ and [d3-THC-COOH+H]+, respectively. The precision and accuracy were tested on spiked urine samples in the range 2.5-125 ng/ml. The mean recovery was 95% (n = 58), coefficients of variations were 2.2-4.3% and the limit of detection 2 ng/ml. Diagnostic qualifying ions of THC-COOH (m/z 327 and m/z 299) and d3-THC-COOH (m/z 330) were generated using up-front collision-induced dissociation. The relative ion intensities in clinical samples (n = 21) were within +/-20% deviation compared with standards. Using this tolerance and the presence of the ions m/z 327 and m/z 299 at the correct retention times as the acceptance criteria for identification of THC-COOH positive samples, the limit of detection was 15 ng/ml. The LC-MS method complies with the current recommendations on drugs of abuse testing, in which mass spectrometric detection is emphasized.