Quantitative analysis of drugs in hair by UHPLC high resolution mass spectrometry

Abstract

Liquid chromatographic methods coupled to high resolution mass spectrometry are increasingly used to identify compounds in various matrices including hair but there are few recommendations regarding the parameters and their criteria to identify a compound. In this study we present a method for the identification and quantification of a range of drugs and discuss the parameters used to identify a compound with high resolution mass spectrometry.Drugs were extracted from hair by incubation in a buffer:solvent mixture at 37°C during 18h. Analysis was performed on a chromatographic system comprised of an Agilent 6550 QTOF coupled to a 1290 Infinity UHPLC system. High resolution accurate mass data were acquired in the All Ions mode and exported into Mass Hunter Quantitative software for quantitation and identification using qualifier fragment ions.Validation included selectivity, matrix effects, calibration range, within day and between day precision and accuracy. The analytes were 7-amino-flunitrazepam, 7-amino-clonazepam, 7-amino-nitrazepam, acetylmorphine, alimemazine, alprazolam, amphetamine, benzoylecgonine, buprenorphine, diazepam, ethylmorphine, fentanyl, hydroxyzine, ketobemidone, codeine, cocaine, MDMA, methadone, methamphetamine, morphine, oxycodone, promethazine, propiomazine, propoxyphene, tramadol, zaleplone, zolpidem, and zopiclone. As proof of concept, hair from 29 authentic post mortem cases were analysed.The calibration range was established between 0.05ng/mg to 5.0ng/mg for all analytes except fentanyl (0.02–2.0), buprenorphine (0.04–2.0), and ketobemidone (0.05–4.0) as well as for alimemazine, amphetamine, cocaine, methadone, and promethazine (0.10–5.0). For all analytes, the accuracy of the fortified pooled hair matrix was 84–108% at the low level and 89–106% at the high level. The within series precisions were between 1.4 and 6.7% and the between series precisions were between 1.4 and 10.1%.From the 29 autopsy cases, 121 positive findings were encountered from 23 of the analytes in concentrations similar to those previously published.We conclude that the developed method proved precise and accurate and that it had sufficient performance for the purpose of detecting regular use of drugs or treatment with prescription drugs. To identify a compound we recommend the use of ion ratios as a complement to instrument software “matching scores”.