RESUMO
In Portugal, and worldwide, the abuse of psychoactive substances is increasing, with a significant incidence of deaths related to their consumption. Opiates are one of the most prevalent group of substances in that context, and they are responsible for a significant impact of the mentioned harms. Therefore, it becomes necessary to equip labs with faster and effective methods to identify and quantify these substances. This work describes the development and validation of a novel analytical method for the simultaneous determination of morphine, codeine and 6-monoacetylmorphine in blood samples by gas chromatography-tandem mass spectrometry (GC-MS/MS), using microextraction by packed sorbent (MEPS) for sample preparation. Before the MEPS procedure, a precipitation step with acetonitrile was performed. The MEPS parameters were optimized using the fractional factorial planning (2k-1) and surface response methodology. The final optimized conditions were: number of strokes (20), amount of formic acid in the washing solution (3.36%), number of washes of the sorbent (1), amount of ammonium hydroxide in the elution solution (2.36%) and number of elution cycles (11). After the extraction procedure, the analytes were derivatized with MSTFA with 5% TMS. Using a sample volume of 250⯵L, the method was validated according to internationally accepted standards. The method proved to be linear in the range of 5-1000â¯ng/mL with coefficients of determination greater than 0.99 for all analytes. Intra-and inter-day precision and accuracy were in accordance with the above-mentioned criteria, presenting coefficients of variation ≤15% and relative errors within a range of ± 15% of the theoretical concentration. The absolute recoveries ranged from 6 to 23%. The validated method was applied to the analysis of real samples, being an advantageous tool for the detection of those substances in blood. This is the first time that GCMS/MS with MEPS was used for the determination of these compounds in biological fluids.