Journal of chromatography. A

Rapid quantitative chiral amphetamines liquid chromatography-tandem mass spectrometry: method in plasma and oral fluid with a cost-effective chiral derivatizing reagent.

PMID 25065924


Methamphetamine is a widely abused psychostimulant containing a chiral center. Consumption of over-the-counter and prescription medications may yield positive amphetamines results, but chiral separation of l- and d-methamphetamine and its metabolite amphetamine can help determine whether the source was licit or illicit. We present the first LC-MS/MS method with precolumn derivatization for methamphetamine and amphetamine chiral resolution in plasma and oral fluid collected with the Oral-Eze(®) and Quantisal™ devices. To 0.5mL plasma, 0.75mL Oral-Eze, or 1mL Quantisal specimen racemic d11-methamphetamine and amphetamine internal standards were added, followed by protein precipitation. Samples were centrifuged and supernatants loaded onto pre-conditioned Phenomenex(®) Strata™-XC Polymeric Strong Cation solid phase extraction columns. After washing, analytes were eluted with 5% ammonium hydroxide in methanol. The eluate was evaporated to dryness and reconstituted in water. Derivatization was performed with 1-fluoro-2,4-dinitrophenyl-5-l-alanineamide (Marfey's reagent) and heating at 45°C for 1h. Derivatized enantiomer separations were performed under isocratic conditions (methanol:water, 60:40) with a Phenomenex(®) Kinetex(®) 2.6μm C18 column. Analytes were identified and quantified by two MRM transitions and their ratio on a 3200 QTrap (AB Sciex) mass spectrometer in ESI negative mode. In all three matrices, the method was linear for all enantiomers from 1 to 500μg/L, with imprecision and accuracy of ≤11.3% and 85.3-108%, respectively. Extraction efficiencies ranged from 67.4 to 117% and matrix effects from -17.0 to 468%, with variation always ≤19.1%. Authentic plasma and OF specimens were collected from an IRB-approved study that included controlled Vicks(®) VapoInhaler™ administration. The present method is sensitive, selective, economic and rapid (separations accomplished in <10min), and improves methamphetamine result interpretation.