An investigation of the stability of free and glucuronidated 11-nor-delta9-tetrahydrocannabinol-9-carboxylic acid in authentic urine samples.
Preanalytical stability of a drug and its major metabolites is an important consideration in pharmacokinetic studies or whenever the analyte pattern is used to estimate drug habits. Firstly, the stability of free and glucuronidated 11-nor-delta9-tetrahydrocannabinol-9-carboxylic acid (THCCOOH, THCCOOglu) in authentic urine samples was investigated. Random urine samples of cannabis users (n = 38) were stored at -20, 4, and 20 degrees C up to 15 days and up to 5 days at 40 degrees C, and alterations of the analyte pattern during storage were followed by liquid chromatography-tandem mass spectrometry. Secondly, the influence of pH (range 5.0-8.0) on the stability of the analytes was studied using spiked urine to elucidate the results obtained from authentic samples. In authentic urine samples, the initial pH ranged from 5.1 to 8.8. The glucuronide was found to be highly labile at a storage temperature of 4 degrees C and above. Initially, 18 urine samples tested positive for THCCOOH. After 2 days storage at 20 degrees C, THCCOOH was detectable in a further 4 samples, and 7 more samples tested positive for THCCOOH (5-81 ng/mL) after 15 days. Depending on time and temperature, the glucuronide concentration decreased, resulting in an increase of THCCOOH concentration. However, a loss in mean total THCCOOH concentration was found, which was significantly higher in deteriorated samples than in samples without signs of deterioration after 15 days of storage at 20 degrees C. In the drug-free urine sample separately spiked with THCCOOglu or THCCOOH, the investigations on the stability of the target analytes at various pH values revealed that THCCOOH was stable at pH 5.0. At higher pH values, its concentration slightly decreased with time, and about 69% of the initial THCCOOH concentration was still present at pH 8.0 on day 5. THCCOOglu concentrations rapidly decreased with increasing pH value. For example, only 72% of the initial THCCOOglu concentration could be detected at pH 5.0 on day 1. Degradation of the glucuronide resulted in formation of THCCOOH, which was observed even at pH 5.0. In light of the present findings, advanced forensic interpretations based on the presence of THCCOOH or the pattern of THCCOOH and THCCOOglu in stored urine samples seems questionable.
Institute of Forensic Medicine, Ruprecht-Karls-University, Voss-Str. 2, 69115 Heidelberg, Germany. gisela_skopp@med.uni-heidelberg.de
Gisela
G
Skopp
Lucia
L
Pötsch
D016477:Artifacts; D003951:Diagnostic Errors; D013759:Dronabinol / Q000031:analogs & derivatives* / Q000652:urine*; D004355:Drug Stability; D005554:Forensic Medicine / Q000379:methods*; D020719:Glucuronides / Q000652:urine*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D013058:Mass Spectrometry; D015813:Substance Abuse Detection / Q000379:methods*; D019966:Substance-Related Disorders / Q000652:urine*
D020719:Glucuronides; C016780:11-nor-delta(9)-tetrahydrocannabinol-9-carboxylic acid; D013759:Dronabinol