GC/MS analysis of amphetamine in urine with amphetamine-d5(side chain) as an internal standard

Abstract

This study examined the accessibility of using amphetamine-d5 (side chain) as an internal standard in quantitative analysis of the trichloroacetyl derivative of urinary amphetamine by gas chromatography / mass spectrometry (GC/MS). In GC/MS analysis , fragments with minimal interference between the assay protocol and the internal standard were selected for quantitation. In the full-scan mode, no interference from TCA-amphetamine-d5, the trichloroacetyl derivative of amphetamine-d5, was observed for the fragments of trichloroacetylamphetamine (TCA-amphetamine) with m/z 190, 188, 118 and 91. On the other hand, no interference from TCA-amphetamine was indicated in fragments of m/z 196 and 123 of TCA-amphetamine-d5. In the selective ion monitoring mode (SIM), the interference between m/z 190, 188, 118 and 91 of TCA-amphetamine and m/z 196, 194 and 123 of TCA-amphetamine-d5 was minimal. The fragments of m/z 190 from TCA-amphetamine and m/z 194 from TCA-amphetamine-d5 were thus selected for quantitation of urinary amphetamine by SIM. Calibration graph derived from the ratio of relative intensity of m/z 190 to m/z 194 versus the concentration of urinary amphetamine was linear over the concentration range of 31.25 - 2000 ng/mL (n=7, SD=0.0357, r=0.9987). Amphetamine-d5 was demonstrated to be a suitable internal standard in quantitative analysis of the trichloroacetyl derived urinary amphetamine by SIM analysis with GC/MS.