Two-dimensional gas chromatography/electron-impact mass spectrometry with cryofocusing for simultaneous quantification of MDMA, MDA, HMMA, HMA, and MDEA in human plasma.
BACKGROUND
3,4-Methylenedioxymethamphetamine (MDMA, or Ecstasy) is a popular recreational drug. Analysis of MDMA and metabolites in human plasma, particularly in pharmacokinetic studies, requires low limits of quantification. Two-dimensional GC/MS with cryofocusing is a chromatographic technique recognized for its increased selectivity and resolution.


METHODS
This method simultaneously quantifies 3,4-methylenedioxyethylamphetamine (MDEA), MDMA, and its metabolites, 3,4-methylenedioxyamphetamine (MDA), 4-hydroxy-3-methoxymethamphetamine (HMMA), and 4-hydroxy-3-methoxyamphetamine (HMA) in human plasma. With hydrochloric acid, we hydrolyzed 1 mL plasma, fortified with internal standard. Analytes were subjected to solid-phase extraction, derivatized with heptafluorobutyric acid anhydride, and quantified using cryofocused 2-dimensional GC/MS operated in electron-impact selected ion-monitoring mode.


RESULTS
Limits of quantification were 1.0 microg/L for MDA and 2.5 microg/L for MDEA, MDMA, HMMA, and HMA. Calibration curves were linear to 100 microg/L for MDA and HMA and to 400 microg/L for MDEA, MDMA, and HMMA, with r(2) > 0.997. At 3 concentrations spanning the linear dynamic range of the assay, mean overall extraction efficiencies from plasma were > or =85% for all compounds of interest. Recoveries were 85.6% to 107.2% of target, and intra- and interassay imprecision (CV) was <8.5% for all drugs at 3 concentrations within the range of the assay. None of the 66 exogenous compounds tested interfered with analyte quantification.


CONCLUSIONS
This GC/MS assay provides low limits of quantification for simultaneous determination of MDEA, MDMA, and metabolites MDA, HMMA, and HMA in human plasma. The 2D chromatographic system should be suitable for application to other analytes and to other complex matrices.
Kolbrich
Erin A
EA
Chemistry and Drug Metabolism, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, Baltimore, MD, USA.
Lowe
Ross H
RH
Huestis
Marilyn A
MA
D015104:3,4-Methylenedioxyamphetamine / Q000031:analogs & derivatives / Q000097:blood*; D002138:Calibration; D000697:Central Nervous System Stimulants / Q000097:blood*; D004298:Dopamine / Q000031:analogs & derivatives* / Q000097:blood; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods; D006801:Humans; D006868:Hydrolysis; D008694:Methamphetamine / Q000031:analogs & derivatives* / Q000097:blood; D018817:N-Methyl-3,4-methylenedioxyamphetamine / Q000097:blood*; D015203:Reproducibility of Results; D015813:Substance Abuse Detection / Q000379:methods*
D000697:Central Nervous System Stimulants; C079646:4-hydroxy-3-methoxymethamphetamine; C031105:3-O-methyl-alpha-methyldopamine; D008694:Methamphetamine; D015104:3,4-Methylenedioxyamphetamine; D018817:N-Methyl-3,4-methylenedioxyamphetamine; C051800:3,4-methylenedioxyethamphetamine; D004298:Dopamine