In post-mortem (PM) investigations, forensic toxicologists attempt to identify legal or illegal substances present before death and determine how they contributed to the cause of death. A critical challenge is ensuring that PM sample concentrations accurately reflect those at the time of death, as postmortem redistribution (PMR) can alter these levels due to anatomical and physiological changes. The PMR phenomenon is called a ‘toxicological nightmare’. PMR significantly affects post-mortem drug concentrations, particularly for lipophilic drugs and those with a high volume of distribution. The emergence of new psychoactive substances (NPSs) has led to a growing recognition of their role as a significant public health concern, frequently associated with fatalities related to polydrug use. These substances are renowned for their ability to induce intoxication at low doses, which has led to the continuous updating of toxicological and forensic methods to improve detection and adopt new analytical standards. The comprehensive detection of NPS metabolites, some of which are still undiscovered, presents an additional analytical challenge, as do their metabolic pathways. This complicates their identification in fatal cases using standard analytical methods, potentially leading to an underestimation of their actual prevalence in toxicological results. Furthermore, the interpretation of analytical results is hindered by the absence of data on PM blood levels and the specific contributions of NPS to causes of death, exacerbated by the lack of knowledge of whether the PMR phenomenon influences them. This paper presents a comprehensive review of the literature on post-mortem cases involving various NPS, categorized according to classifications by the United Nations Office on Drugs and Crime (UNODC) and the European Union Drugs Agency (EUDA). These categories include cathinones, phenylethylamines, arylalkylamines, phencyclidine-type substances, phenmetrazines, piperazines, phenidates, aminoindanes, LSD-like NPSs, tryptamines, fentanyl analogs, designer benzodiazepines, synthetic cannabinoids, and nitazenes. This review covers not only postmortem blood levels but also the stability of the substances studied, the methods of analysis, and attempts to shed some light on the PMR phenomenon. This review used various key terms, such as PMR, NPS, and the names of previously categorized substances and drug analyses across multiple peer-reviewed journals and databases, including Scopus, Google Schoolar, Springer, PubMed, and Wiley Online Library. In addition, references from retrieved articles were examined to identify additional relevant research. Interpreting post-mortem toxicological results is complex and lacks definitive guidelines, requiring a nuanced understanding of its challenges and potential pitfalls. As a result, post-mortem toxicology is often regarded as an art. The primary aim of this review is to provide forensic toxicologists with a comprehensive framework to assist in the evaluation and interpretation of NPS analysis. This guide is intended to complement the existing knowledge and practices applied in forensic laboratories within the toxicological analysis of post-mortem cases.

Few studies have reported the prevalence or incidence about the consumption of new psychoactive substances (NPS). The hair analysis can be useful for this purpose. At the present, ketamine is the most consumed arylcyclohexylamine associated to young consumers and polyconsumption profiles. For this reason, ketamine consumer cases become very interesting to provide information on NPS prevalence. In this work, ten former cases of the National Institute of Toxicology and Forensic Science (INTCF) of Madrid Department (INTCFM), all of them belonging to defendants accused of crimes against public health and who had been found positive to ketamine, were reassessed. At the first toxicological analysis of those hair samples, a positive consume in ketamine had been determined by gas chromatography coupled to mass spectrometry (GC-MS). In this work, the same hair samples were reanalyzed by high-resolution mass spectrometry ( UHPLC-HRMS/MS) using an incubation methanolic extraction combined with a single, simpler, non-selective and direct sample pre-treatment. After corroborating the GC-MS results previously obtained for the same samples, the detection of additional NPS using this new methodology evidenced its benefits and opened the possibility to perform a NPS prevalence study. In brief, in those cases with a positive consumption in ketamine, a polyconsumption of other drugs and NPS was found, including different arylcyclohexylamines as deschloroketamine, 3-MeO-PCP and methoxetamine; and cathinones as methylmetcathinone and N-ethyl-pentylone.

The analysis of drugs of abuse in hair and other biological matrices of forensic interest requires great selectivity and sensitivity. This is done traditionally through target analysis, with one or more analytical methods, or with different and specific preanalytical phases, and complex procedures performed by the toxicological laboratories, and there is no exception with ketamine-like compounds, such as methoxetamine, a new psychoactive substance whose use has increased in the last decades, and continues to grow quickly year by year. More validated methods of analysis are needed to detect these substances in low concentrations selectively. Reanalyzing the samples of a former case of a polydrug consumer accused of a crime against public health in Spain, five metabolites of methoxetamine (normethoxetamine, O-desmethylmethoxetamine, dehydromethoxetamine, dihydronormethoxetamine and hydroxynormethoxetamine) were tentatively detected using a high-resolution technique, that is, liquid chromatography coupled to high-resolution mass spectrometry (LC–HR-MS-MS). The highest analytical selectivity of LC–HR-MS-MS method together a universal and simpler pretreatment stages has demonstrated to allow faster analysis and more sensitivity than the one performed traditionally at the INTCF laboratories, which was gas chromatography coupled to mass spectrometry.

The analysis of drugs of abuse in hair and other biological matrices of forensic interest requires great selectivity and sensitivity. This has been traditionally achieved through target analysis, using one or more analytical methods that include different preanalytical stages, and more complex procedures followed by toxicological laboratories. There is no exception with 2C-series drugs, such as 2C-B, a new psychoactive substance (NPS), which use has emerged and significantly increased, year by year, in the last decades. Continuously new analytical methods are required to selectively detect and identify these new marketed substances at very low concentrations. In this case report, one former case of a polydrug consumer (charged of a crime against public health in Spain) was reanalyzed in hair matrix. In this reanalysis, 2C-B has been positively detected and identified using liquid chromatography coupled to high-resolution mass spectrometry (UHPLC-HRMS/MS). The most selective analytical UHPLC-HRMS/MS method alongside a universal and simpler pretreatment methodology has opened up more possibilities for the detection of substances of different chemical structure and optimization of different HRMS/MS detection approaches allowing the identification of 2-CB in the hair of a real forensic case.