Development and Validation of a Multi-drug Analysis for Hair

A single sample preparation and instrument method was developed for the analysis of 54 analytes in hair. Analytes in the panel included drug of abuse, synthetic opioids, and ethyl glucuronide. Hair is an alternative matrix used in forensic toxicology due to its longer detection window. The matrix is also more stable, does not require refrigerated storage, and is less invasive to collect compared to blood. The large panel saves the laboratory time and money when multiple drugs can be identified in a single confirmation. Additionally, when a limited amount of sample is available a single analysis helps save sample for other necessary analyses. 

Hair samples were decontaminated by washing with 2 x 10 mL of acetone and 2 x 10 mL of hexane. Then the hair was dried at room temperature and pulverized. 

Hair samples were extracted using UCT’s Clean Screen^® EtG solid phase extraction column. 30 mg of pulverized hair was added to 50 µL of internal standard and 1.5 mL of 0.1% formic acid in water. SPE columns were conditioned with 3 mL of 1% formic acid in methanol and 3 mL of 1% formic acid in water. After loading samples, columns were washed with 1 mL of methanol. Analytes were eluted with 2 mL of 1% formic acid in methanol and 2 mL of 1% ammonium hydroxide in methanol. Elutions were evaporated and reconstituted in 100 µL initial LC mobile phase composition. Samples were analyzed on an Agilent 1920 Infinity II LC system with a Sciex API 6500 QTrap. 

A single sample preparation and instrument method was validated for the simultaneous analysis of 31 drugs of abuse (DoA), 22 synthetic opioids, and EtG in human hair. Within-run precision, between-run precision, and accuracy across three concentrations fulfilled standard toxicology criteria. The recovery and matrix effects of most compounds were acceptable without sacrificing the panel’s limit of detection and lower limit of quantitation. Furthermore, this method was applied to 20 proficiency test samples and yielded acceptable z-score values. 

Citation: D. Vandelli, F. Palazzoli, P. Verri, V. Castagnetti, C. Profeta, A. Borghi, R. Cecchi, A novel method for the simultaneous determination of drugs of abuse, ethyl glucuronide and synthetic opioids in human hair through a single digestion, purification and analysis in LC-MS/MS, Journal of Chromatography B, Volume 1246, 2024, 124269, ISSN 1570-0232, https://doi.org/10.1016/j.jchromb.2024.124269.