This paper presents a method for the determination of xylazine in whole blood using solid-phase extraction and gas chromatography-mass spectrometry. This technique required only 0.5 mL of sample, and protriptyline was used as internal standard (IS). Limits of detection and quantitation (LCQ) were 2 and 10 ng/mL, respectively. The method was found to be linear between the LOQ and 3.50 µg/mL, with correlation coefficients higher than 0.9922. Precision (intra- and interday) and accuracy were in conformity with the criteria normally accepted in bioanalytical method validation. The analyte was stable in the matrix for at least 18 h at room temperature and for at least three freeze/thaw cycles. Mean recovery, calculated at three concentration levels, was 87%. To the best of our knowledge, this is the first time that solid-phase extraction is used as sample preparation technique for the determination of this compound in biological media. Because of its simplicity and speed when compared to other extraction techniques, the herein described method can be successfully applied in the diagnosis of intoxications by xylazine.

Cannabis is the most consumed illicit drug worldwide, and its legal status is a source of concern. This study proposes a rapid procedure for the simultaneous quantification of Δ⁹-tetrahydrocannabinol (THC), 11-hydroxy-Δ⁹-tetrahydrocannabinol (11-OH-THC), 11-nor-9-carboxy-Δ⁹-tetrahydrocannabinol (THC-COOH), cannabidiol (CBD), and cannabinol (CBN) in urine samples. Microextraction by packed sorbent (MEPS) was used to pre-concentrate the analytes, which were detected by gas chromatography–mass spectrometry. The procedure was previously optimized, and the final conditions were: conditioning with 50 µL methanol and 50 µL of water, sample load with two draw–eject cycles, and washing with 310 µL of 0.1% formic acid in water with 5% isopropanol; the elution was made with 35 µL of 0.1% ammonium hydroxide in methanol. This fast extraction procedure allowed quantification in the ranges of 1–400 ng/mL for THC and CBD, 5–400 ng/mL for CBN and 11-OH-THC, and 10–400 ng/mL for THC-COOH with coefficients of determination higher than 0.99. The limits of quantification and detection were between 1 and 10 ng/mL using 0.25 mL of sample. The extraction efficiencies varied between 26 and 85%. This analytical method is the first allowing the for determination of cannabinoids in urine samples using MEPS, a fast, simple, and low-cost alternative to conventional techniques.

New Psychoactive Substances are currently a serious and growing problem affecting public health worldwide. By 2022, 1184 of these substances had been identified over a period of 16 years. Within these, α-pyrrolidinohexanophenone (α-PHP) and α-pyrrolidinoisohexanophenone (α-PiHP) have emerged, two synthetic cathinones from the pyrovalerone derivates subgroup that are positional isomers of each other. Alpha-PHP appeared on the Japanese illicit drug market in 2014 and, two years later, α-PiHP was identified for the first time in China. They were placed in schedule II on the list of Psychotropic Substances under International Control in 2020 and in March 2023, respectively. Both cathinones have no therapeutic potential for medical use and therefore are abused for recreational habits, which can lead to fatalities. The most frequent adverse effects reported are cardiac, psychiatric, and neurologic, and fatal intoxications have already been described. In Portugal, their consumption and consequent seizures are more prevalent on the archipelagos, which has been aggravating the health situation. In conclusion, these types of substances are a challenge for forensic toxicology since they are easily synthesized, modified, and placed on the market. Therefore, more studies to develop analytical methods to detect them and more comprehensive legislation should be applied. Thus, this review aimed to address the legislative, physicochemical, toxicological, and analytical aspects of both substances.

New Psychoactive Substances (NPS) still represent an issue of great concern worldwide despite efforts made by national and international control systems to limit the spread of these substances. Alpha-pyrrolidinohexanophenone (α-PHP) is a fairly recent synthetic cathinone (the second largest group of monitored substances in Europe) with only a few published studies on the substance. Though there is a low incidence of NPS consumption in Portugal, a recent increase in apprehensions and detection in biological matrices of the substance was verified. An analytical methodology was developed and validated for determining and quantitating α-PHP in blood. Solid phase extraction was employed for sample preparation (500 μL), further analysed by GC-MS-EI in single ion monitoring mode, with cocaine-d3 as the internal standard. Method validation followed the guidelines of the American National Standards Institute/AAFS Standards Board (ANSI/ASB Standard 036). The procedure was linear between 10 and 1000 ng/mL, with determination coefficients (r²) higher than 0.999. Carryover was not observed. A limit of detection of 5ng/mL and a limit of quantitation of 10 ng/mL were achieved. Intraday and intermediate precision and bias assessment showed satisfactory results (CV <17.7%; bias <11.6%), and extraction efficiency ranged from 98.5% to 103.3%. The stability of the substance was considered acceptable for at least 6h at room temperature, 48h in the autosampler, and 21 days after five freeze/thaw cycles. The developed methodology was applied to fifteen real samples from the Laboratory of Chemistry and Forensic Toxicology, Centre Branch, of the National Institute of Legal Medicine and Forensic Sciences, Portugal, with drug concentrations ranging from 15 ng/mL to 227 ng/mL. Available information for each case is also detailed in the present article.

Opioids are the drugs most commonly detected in overdose deaths and the second most consumed worldwide. An analytical methodology has been optimized and fully validated for the determination of codeine, morphine, 6-acetylmorphine, 6-acetylcodeine, oxycodone, oxymorphone and fentanyl in whole blood and pericardial fluid. The internal standards used were codeine-d₃, morphine-d₃, 6-acetylmorphine-d₃ and fentanyl-d₅. Before solid phase extraction, volumes of 250 μL of blood and pericardial fluid were subjected to a protein precipitation (with 750 μL of ice-cold acetonitrile) and a microwave induced oximation was performed using a solution of 1% aqueous hydroxylamine hydrochloride in PBS (1:2, v/v). Finally, the dried extracts were further derivatized with a solution of MSTFA+5% TMCS under microwave irradiation. The chromatographic analysis was carried out using gas chromatography–mass spectrometry operating in electron impact and selected ion monitoring mode. For all analytes the method was linear between 5 and 1000 ng/mL with determination coefficients (r²) higher than 0.99. Depending on the analyte and matrix the limit of detection varies between 3 and 4 ng/mL. Intra- and intermediate precision (<20%) and bias (±20%) were acceptable for all analytes in both matrices. The stability of the substances in the studied matrices was guaranteed, at least, 24 h in the autosampler, 4 h at room temperature and 30 days after three freeze/thaw cycles. This methodology was applied to real samples from the Laboratory of Chemistry and Forensic Toxicology, Centre Branch, of the National Institute of Legal Medicine and Forensic Sciences, Portugal.