Determination of amphetamine enantiomers in urine by conductive vial electromembrane extraction and ultra‐high performance supercritical fluid chromatography tandem mass spectrometry

Abstract

Separation and quantification of amphetamine enantiomers are commonly used to distinguish between consumption of prescription amphetamine (mostly S-amphetamine) and illicit forms of the drug (racemate). In this study, electromembrane extraction with prototype conductive vials was combined with ultra-high performance supercritical fluid chromatography (UHPSFC-MS/MS) to quantify R-and Samphetamine in urine. Amphetamine was extracted from 100 μL urine, diluted with 25 μL internal standard solution and 175 μL 130 mM formic acid, across a supported liquid membrane (SLM) consisting of 9 μL of a 1:1(w/w) mixture of 2-nitrophenyloctyl ether (NPOE) and bis(2-ethylhexyl)phosphite (DEHPi) into an acceptor phase containing 300 μL 130 mM formic acid. The extraction was facilitated by the application of 30 V for 15 min. Enantiomeric separation was achieved using UHPSFC-MS/MS with a chiral stationary phase. The calibration range was 50-10,000 ng/mL for each enantiomer. The between-assay CV was ≤5%, within-assay CV ≤ 1.5%, and bias within ±2%. Recoveries were 83%-90% (CV ≤ 6%), and internal standard corrected matrix effects were 99-105 (CV ≤ 2%). The matrix effects ranged from 96% to 98% (CV ≤ 8%) when not corrected by the internal standard. The EME method was compared with a chiral routine method that employed liquid-liquid extraction (LLE) for sample preparation. Assay results were in agreement with the routine method, and the mean deviation between methods was 3%, ranging from À21% to 31%. Finally, sample preparation greenness was assessed using the AGREEprep tool, which resulted in a greenness score of 0.54 for conductive vial EME, opposed to 0.47 for semi-automated 96-well LLE.

Generic Conditions for Electromembrane Extraction of Acids with Low to Moderate Hydrophilicity in Human Plasma

“…First, the stability and performance of 1-octanol with plasma samples was attempted improved by mixing it 1:1 (weight ratio) with 2-nitrophenyl octyl ether (NPOE), based on previous reports [21,22,34]. However, this did not improve the stability.…”

“…In 2022, the first commercial EME equipment was however launched, based on using conductive vials to house the sample and acceptor solution [19]. Since then, this vial-based format has been applied for EME in therapeutic drug monitoring (TDM) [12,20] and analysis of drugs-ofabuse [21,22].…”