Multiresidue screening of common psychotropic drugs in hair samples by ultra-high performance liquid chromatography-tandem mass spectrometry

Introduction In the forensic context, the determination of common psychotropic drugs at low concentration is increasingly requested in hair samples for the retrospective investigation of chronic drug abuse and dependence. To meet the high demand for drug analysis in hair samples, toxicology laboratories are forced to update their procedures for the identification of an increasing number of drugs but also to develop rapid, simpler and more sensitive screening techniques that achieve shorter sample preparation and analysis time, in order to increase the laboratory throughput. Our goal was to develop and validate a sensitive multiresidue screening method for hair samples using a dedicated UHPLC-MS/MS protocol. Methods Decontaminated hair samples were extracted with methanol overnight at +55°C. 50 µL of methanol was directly transferred into a vial for UHPLC-MS/MS analysis. A Shimadzu Nexera UHPLC-system interfaced to an Applied Biosystem 5500TM triple-quadrupole mass spectrometer was utilized in the ESI positive ion mode. LC separation was performed using a Waters® UHPLC column (BEH C-18, 100x2.1 mm i.d, 1.7 µm particle diameter) maintained at +35°C. A 7 min gradient elution with formic acid 5 mM and acetonitrile (90:10 to 0:100, v:v), followed by 1 min isocratic elution, at a flow rate of 0.4 mL/min was performed. Data were acquired at unit mass resolution in the selected-reaction monitoring (SRM) mode. Validation according to ISO 17025 and international guidelines is in progress. Preliminary data Morphine, codeine, 6-MAM, methadone, buprenorphine, amphetamine, methamphetamine, MDA, MDMA, MDEA, Δ9-THC, cocaine and benzoylecgonine were extracted from hair samples and detected after simple sample processing. To increase selectivity and detection capability, different SRM time-windows were created. Three SRM transitions were utilized for identifying and determining each analyte. To maximize the fragment ion signals while maintaining comparable precursor ion abundance, different collision energies were optimized for each compound. For each analyte, calibration was performed by internal standardization and the calibration parameters were obtained using the least squares regression method. Linearity range was observed from 0.04 to 1.0 ng/mg for Δ9-THC and from 0.2 to 5.0 ng/mg for all other analytes. Precision, accuracy and trueness, detection and quantification limits (LODs and LOQs), recovery, selectivity, specificity, carry-over and matrix effect phenomena will be evaluated. The advanced technology of UHPLC-MS/MS permits to obtain, with a single extraction step and without derivatization, rapid separation of many compounds belonging to different drug families and simultaneous monitoring of a high number of transitions with high detection capability, selectivity and resolution power. New screening strategies for analyzing hair samples are progressively developed and applied in the routine use of toxicology laboratories, with the final purpose of increasing sample-throughput and reduce the number of confirmatory analyses by GC-MS methods. Novel aspect Application of UHPLC-MS/MS instrumentation to hair analysis allows high sample-throughput, sensitivity and selectivity in forensic investigation, for new drug-screening strategies.