Liquid chromatography-electrospray ionization mass spectrometry for the detection of lysergide and a major metabolite, 2-oxo-3-hydroxy-LSD, in urine and blood.

A method is presented for the quantitative measurement of lysergide (LSD) and its metabolite 2-oxo-3-hydroxy-LSD (O-H-LSD) in urine and blood. O-H-LSD has been reported to have urinary concentrations many times higher than LSD. Measuring its presence in urine would significantly extend the detection time for confirming LSD abuse. A single-step liquid-liquid extraction was performed on 5-mL urine samples prior to separation by gradient liquid chromatography (LC). Electrospray ionization was used to produce the positively charged ions of O-H-LSD, 2-oxo-3-hydroxy-LAMPA (O-H-LAMPA, internal standard), LSD, and iso-LSD. Varying the orifice voltage in the intermediate-pressure region of the source generated the fragmentation necessary to produce qualifying ions. Selected ion monitoring allowed detection limits of 400 pg/mL and 100 pg/mL for O-H-LSD and LSD, respectively. The method was linear for O-H-LSD from 400 to 8000 pg/mL and for LSD from 100 to 6000 pg/mL. LSD-positive samples (n = 9) analyzed by the liquid chromatography-mass spectrometry method were found to contain mean concentrations of 6378 pg/mL O-H-LSD (332-21371 pg/mL) and 844 pg/mL LSD (177-2456 pg/mL). O-H-LSD urinary concentrations were between 0.9 and 19.8 times higher than LSD (mean = 10.2). Whole-blood samples were also analyzed following additional sample cleanup. LSD was measured in the blood samples, but no O-H-LSD was detected. Enzymatic hydrolysis was carried out on LSD-positive samples (n = 6) to evaluate the existence of conjugated O-H-LSD. Beta-glucuronidase from Helix pomatia was incubated with urine samples at 37 degrees C, pH 5.2 for 24 h. At an enzymatic activity of approximately 4000 units per milliliter of urine, no significant (p = 0.05) difference was seen between hydrolyzed and nonhydrolyzed samples suggesting an absence of O-H-LSD-glucuronic acid conjugation.

Assessment of the ion-trap mass spectrometer for routine qualitative and quantitative analysis of drugs of abuse extracted from urine.

The ion-trap mass spectrometer (MS) has been available as a detector for gas chromatography (GC) for nearly two decades. However, it still occupies a minor role in forensic toxicology drug-testing laboratories. Quadrupole MS instruments make up the majority of GC detectors used in drug confirmation. This work addresses the use of these two MS detectors, comparing the ion ratio precision and quantitative accuracy for the analysis of different classes of abused drugs extracted from urine. Urine specimens were prepared at five concentrations each for amphetamine (AMP), methamphetamine (METH), benzoylecgonine (BZE), delta9-carboxy-tetrahydrocannabinol (delta9-THCCOOH), phencyclidine (PCP), morphine (MOR), codeine (COD), and 6-acetylmorphine (6-AM). Concentration ranges for AMP, METH, BZE, delta9-THCCOOH, PCP, MOR, COD, and 6-AM were 50-2500, 50-5000, 15-800, 1.5-65, 1-250, 500-32000, 250-21000, and 1.5-118 ng/mL, respectively. Sample extracts were injected into a GC-quadrupole MS operating in selected ion monitoring (SIM) mode and a GC-ion-trap MS operating in either selected ion storage (SIS) or full scan (FS) mode. Precision was assessed by the evaluation of five ion ratios for n = 15 injections at each concentration using a single-point calibration. Precision measurements for SIM ion ratios provided coefficients of variation (CV) between 2.6 and 9.8% for all drugs. By comparison, the SIS and FS data yielded CV ranges of 4.0-12.8% and 4.0-11.2%, respectively. The total ion ratio failure rates were 0.2% (SIM), 0.7% (SIS), and 1.2% (FS) for the eight drugs analyzed. Overall, the SIS mode produced stable, comparable mean ratios over the concentration ranges examined, but had greater variance within batch runs. Examination of postmortem and quality-control samples produced forensically accurate quantitation by SIS when compared to SIM. Furthermore, sensitivity of FS was equivalent to SIM for all compounds examined except for 6-AM.

Detection of lysergic acid diethylamide (LSD) in urine by gas chromatography-ion trap tandem mass spectrometry.

A confirmatory method for the detection and quantitation of lysergic acid diethylamide (LSD) is presented. The method employs gas chromatography-tandem mass spectrometry (GC-MS-MS) using an internal ionization ion trap detector for sensitive MS-MS-in-time measurements of LSD extracted from urine. Following a single-step solid-phase extraction of 5 mL of urine, underivatized LSD can be measured with limits of quantitation and detection of 80 and 20 pg/mL, respectively. Temperature-programmed on-column injections of urine extracts were linear over the concentration range 20-2000 pg/mL (r2 = 0.999). Intraday and interday coefficients of variation were < 6% and < 13%, respectively. This procedure has been applied to quality-control specimens and LSD-positive samples in this laboratory. Comparisons with alternate GC-MS methods and extraction procedures are discussed.