Purpose: Drug detection: to present a method for detecting and analyzing enantiomers of MDMA and metabolites in blood and urine. Design: Samples drawn from a volunteer enrolled in a randomized, placebo-controlled, double-blind study of the effects of 100 mg MDMA given alone or with 0.8 mg/kg ethanol. However, only samples from the MDMA-only condition were used in this study. Design of analysis was a non-experimental comparison of samples collected over time using achiral and chiral methods (detecting different enantiomers).

Subjects: One MDMA-experienced male volunteer enrolled in the study described in the report by Hernandez-Lopez and colleagues (2002). This study examined male volunteers (ages 19-36, weight 59-81 kg, height 167-183 cm) recruited via Òword of mouth.Ó Criteria for Inclusion - Lack of major psychiatric or medical illness as assessed through interview and physical examination, routine laboratory tests, urinalysis and ECG. Lack of substance abuse (except for nicotine dependence), and extensive CYP2D6 metabolizer as assessed via urinary dextromethorphan/dextorphan ratio. Lack of adverse psychiatric or medical reactions to ecstasy/MDMA. All participants (including this one) were required to have had taken ecstasy at least five times over a lifetime and to have experience with acute ethanol intoxication.

Measures: Gas chromatography-mass spectrometry (GC-MS) analysis for detecting MDMA, 4-hydroxy-3-methoxymethamphetamine (HMMA), 3,4-methylenedioxyamphetamine (MDA) and 4-hydroxy-3-methyoxyamphetamine (HMA). The authors specify a GC-MS method for detecting racemate and each enantiomer. MDMA and metabolites detected and quantified in blood samples collected at 0, 15, 30, 45, 60, 75, and 90 min and at 2, 3, 4, 6, 8, and 10 h post-MDMA. urine samples apparently collected at least from 1-2 h post-MDMA and for an unspecified period of time. (Specific information on urine collection not provided either in the this paper or in the previous publication (Hernandez-Lopez et al. 2002.) Both chiral and achiral analyses were performed on 1 plasma sample taken 90 min post-MDMA and in one urine sample collected 1-2 h post-MDMA.

Analyses: The newly developed method of chiral detection was compared with the achiral method. The method was tested for detection limit, quantification limit and error rate. R/S ratios were calculated for enantiomers of MDMA and for 2 of 3 metabolites (HMMA and MDA) in blood and urine.

Results: Both R-(-) and S-(+) enantiomers of MDMA and HMMA were detected in blood and urine, but the researchers were unable to establish the identities of the R-(-) and S-(+) enantiomers of HMA or MDA; the different stereoisomers are referred to as (1)-MDA or HMA and (2)-MDA or HMA. (The method is described in detail and involves two derivatization steps). Only low levels of the MDMA metabolite HMA were detected in urine, and none were detected in blood. Serum and urine levels of each substances are listed from blood taken at 90 min post-drug and urine at 1-2 h post-drug and compared on chiral and achiral analysis. Quantification of blood levels Ð Achiral analysis detected 98.7 mcg/L ± MDMA (92.1 mcg/L summed enantiomer determinations. Chiral analysis detected 53.1 mcg/L R-(-)-MDMA and 39 mcg/L S-(+)-MDMA, with a R/S ratio of 1.36. Achiral analysis detected 3.1 mcg/L ± MDA, and 2.1 mcg/L summed enantiomer determinations. Chiral analysis detected 0.6 mcg/L (1)-MDA and 1.4 mcg/L (2)-MDA. (R and S designations not given because they could not be determined). The R/S ratio for MDA was 0.43. Achiral analysis detected 173 mcg/L ± HMMA, 169.7 mcg/L summed enantiomer determinants. Chiral analysis detected 93.6 mcg/L R-(-)-HMMA and 86.1 mcg/L S-(+)-HMMA, with an R/S ratio of 0.97. Neither achiral nor chiral analysis detected HMA (or its enantiomers) in blood. Quantification of Urinary Samples - Achiral analysis detected 449.8 mcg/L ± MDMA (466.9 mcg/L summed enantiomer determinations. Chiral analysis detected 265.3 mcg/L R-(-)-MDMA and 201.6 mcg/L S-(+)-MDMA, with a R/S ratio of 1.32. Achiral analysis detected 7.8 mcg/L ± MDA, and 7.9 mcg/L summed enantiomer determinations. Chiral analysis detected 2.2 mcg/L (1)-MDA and 5.7 mcg/L (2)-MDA. (R and S designations not given because they could not be determined). The R/S ratio for MDA was 0.39. Achiral analysis detected 326.7 mcg/L ± HMMA, 324.2 mcg/L summed enantiomer determinants. Chiral analysis detected 154.5 mcg/L R-(-)-HMMA and 169.7 mcg/L S-(+)-HMMA, with an R/S ratio of 0.91. Achiral analysis apparently did not detect ± HMA, though it detected summed enantiomer determinants of 2.7 mcg/L. Chiral analysis was only able to detect (2)-HMA, at 2.7 mcg/L. Hence, no R/S ratio was calculated.

Overall Effects: The authors described a method for detecting the enantiomers of MDMA and its metabolites HMMA, MDA and HMA in blood and urine. Through this method, they detected and compared levels of R-(-) and S-(+) enantiomers in blood and urine collected from a male volunteer given 100 mg MDMA. As also noted in a previous publication (Pizarro et al. 2002), differences arise in the R/S ratios of HMMA and MDA. Specifically, the R/S ratios of MDMA and MDA are opposite each other in both blood and urine, with R-(-)-MDMA more abundant than S-(+)-MDMA and S-(+)-MDA more abundant than R-(-)-MDA. However, the R/S ratio for HMMA in blood and urine is close to 1 (0.97 in blood and 0.91 in urine), meaning that levels of R-(-)-HMMA and S-(+)-HMMA are equally abundant. As seen in previous studies (De la Torre et al. 2000; Fallon et al. 2000; Pizarro et al. 2002), HMMA is detected in greater quantities than MDA. Previous studies also indicate that the R/S ratio for HMMA does not change much over time.

Adverse Effects: None reported in this paper; study examined one volunteer, with data appearing in other publications.

Comments: This paper is one in a series of papers published by the same research team (e.g. Navarro et al. 2001; Segura et al. 2001; Picchini et al. 2002; Pizarro et al. 2002) reporting on drug detection and drug deposition in human volunteers. Results of the chiral GC-MS analysis suggest that the metabolism of MDMA (or intermediate metabolite) to HMMA is not stereoselective, as would be expected if metabolism occurred through CYP2D6. Hence this method also supports findings in a previous paper (Pizarro et al. 2002) and hypothesis in a review paper (Kraemer and Maurer 2002) for the involvement of enzymes other than CYP2D6 in the human metabolism of MDMA. So far, all experimental studies of MDMA enantiomer deposition in humans have employed only male participants. While it seems unlikely that gender differences exist in drug deposition, future research should examine MDMA metabolism in females. This paper is not intended as a study across subjects, and only features a sample of one. However, the results compare well with findings from a sample of 6 (Pizarro et al. 2002).