Joachim Frost

Lethal poisonings with AH-7921 in combination with other substances

AH-7921 is a synthetic μ-opioid agonist, approximately equipotent with morphine. We report the death of two young individuals after ingestion of AH-7921 in combination with other psychoactive drugs. In the first case a young man died shortly after ingesting Internet drugs. Toxicological analysis of post mortem peripheral blood revealed AH-7921 (0.43 mg/L), 2-FMA (0.0069 mg/L) and 3-MMC (0.0021 mg/L) as well as codeine (0.42 mg/L), codeine-6-glucuronide (0.77 mg/L) and acetaminophen (18.7 mg/L). The second case involved a young female found dead at home. The only positive finding at medicolegal autopsy was needle marks. Toxicological analysis revealed AH-7921 (0.33 mg/L), methoxetamine (MXE) (0.064 mg/L), etizolam (0.27 mg/L), phenazepam (1.33 mg/L), 7-aminonitrazepam (0.043 mg/L), diazepam (0.046 mg/L), nordiazepam (0.073 mg/L), and oxazepam (0.018 mg/L) in blood. In both cases intoxication with AH-7921 in combination with other psychoactive drugs was considered to be the cause of death.

Investigation of morphine and morphine glucuronide levels and cytochrome P450 isoenzyme 2D6 genotype in codeine-related deaths

Compared to morphine and morphine-6-glucuronide (M6G), codeine and its other major metabolites codeine-6-glucuronide and norcodeine have weak affinity to opioid μ-receptors. Analgesic effects of codeine are thus largely dependent on metabolic conversion to morphine by the polymorphic cytochrome P450 isoenzyme 2D6 (CYP2D6). How this relates to toxicity and post-mortem whole blood levels is not known. This paper presents a case series of codeine-related deaths where concentrations of morphine, M6G and morphine-3-glucuronide (M3G), as well as CYP2D6 genotype, are taken into account. Post-mortem toxicological specimens from a total of 1444 consecutive forensic autopsy cases in Central Norway were analyzed. Among these, 111 cases with detectable amounts of codeine in femoral blood were identified, of which 34 had femoral blood concentrations exceeding the TIAFT toxicity threshold of 0.3mg/L. Autopsy records of these 34 cases were retrieved and reviewed. In the 34 reviewed cases, there was a large variability in individual morphine to codeine concentration ratios (M/C ratios), and morphine levels could not be predicted from codeine concentrations, even when CYP2D6 genotype was known. 13 cases had codeine concentrations exceeding the TIAFT threshold for possibly lethal serum concentrations (1.6 mg/L). Among these, 8 individuals had morphine concentrations below the toxic threshold according to TIAFT (0.15 mg/L). In one case, morphine as well as M6G and M3G concentrations were below the limit of detection. A comprehensive investigation of codeine-related fatalities should, in addition to a detailed case history, include quantification of morphine and morphine metabolites. CYP2D6 genotyping may be of interest in cases with unexpectedly high or low M/C ratios.

A Validated Method for Simultaneous Determination of Codeine, Codeine-6-Glucuronide, Norcodeine, Morphine, Morphine-3-Glucuronide and Morphine-6-Glucuronide in Post-Mortem Blood, Vitreous Fluid, Muscle, Fat and Brain Tissue by LC–MS

The toxicodynamics and, to a lesser degree, toxicokinetics of the widely used opiate codeine remain a matter of controversy. To address this issue, analytical methods capable of providing reliable quantification of codeine metabolites alongside codeine concentrations are required. This article presents a validated method for simultaneous determination of codeine, codeine metabolites codeine-6-glucuronide (C6G), norcodeine and morphine, and morphine metabolites morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G) in post-mortem whole blood, vitreous fluid, muscle, fat and brain tissue by high-performance liquid chromatography mass spectrometry. Samples were prepared by solid-phase extraction. The validated ranges were 1.5-300 ng/mL for codeine, norcodeine and morphine, and 23-4,600 ng/mL for C6G, M3G and M6G, with exceptions for norcodeine in muscle (3-300 ng/mL), morphine in muscle, fat and brain (3-300 ng/mL) and M6G in fat (46-4,600 ng/mL). Within-run and between-run accuracy (88.1-114.1%) and precision (CV 0.6-12.7%), matrix effects (CV 0.3-13.5%) and recovery (57.8-94.1%) were validated at two concentration levels; 3 and 150 ng/mL for codeine, norcodeine and morphine, and 46 and 2,300 ng/mL for C6G, M3G and M6G. Freeze-thaw and long-term stability (6 months at -80°C) was assessed, showing no significant changes in analyte concentrations (-12 to +8%). The method was applied in two authentic forensic autopsy cases implicating codeine in both therapeutic and presumably lethal concentration levels.

Post-mortem levels and tissue distribution of codeine, codeine-6-glucuronide, norcodeine, morphine and morphine glucuronides in a series of codeine-related deaths

This article presents levels and tissue distribution of codeine, codeine-6-glucuronide (C6G), norcodeine, morphine and the morphine metabolites morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G) in post-mortem blood (peripheral and heart blood), vitreous fluid, muscle, fat and brain tissue in a series of 23 codeine-related fatalities. CYP2D6 genotype is also determined and taken into account. Quantification of codeine, C6G, norcodeine, morphine, M3G and M6G was performed with a validated solid phase extraction LC-MS method. The series comprise 19 deaths (83%) attributed to mixed drug intoxication, 4 deaths (17%) attributed to other causes of death, and no cases of unambiguous monointoxication with codeine. The typical peripheral blood concentration pattern in individual cases was C6G≫codeine≫norcodeine>morphine, and M3G>M6G>morphine. In matrices other than blood, the concentration pattern was similar, although in a less systematic fashion. Measured concentrations were generally lower in matrices other than blood, especially in brain and fat, and in particular for the glucuronides (C6G, M3G and M6G) and, to some extent, morphine. In brain tissue, the presumed active moieties morphine and M6G were both below the LLOQ (0.0080mg/L and 0.058mg/L, respectively) in a majority of cases. In general, there was a large variability in both measured concentrations and calculated blood/tissue concentration ratios. There was also a large variability in calculated ratios of morphine to codeine, C6G to codeine and norcodeine to codeine in all matrices, and CYP2D6 genotype was not a reliable predictor of these ratios. The different blood/tissue concentration ratios showed no systematic relationship with the post-mortem interval. No coherent degradation or formation patterns for codeine, morphine, M3G and M6G were observed upon reanalysis in peripheral blood after storage.

Enantiomeric separation and quantification of R/S-amphetamine in urine by ultra-high performance supercritical fluid chromatography tandem mass spectrometry

To distinguish between legal and illegal consumption of amphetamine reliable analytical methods for chiral separation of the R- and S-enantiomers of amphetamine in biological specimens are required. In this regard, supercritical fluid chromatography (SFC) has several potential advantages over liquid chromatography, including rapid separation of enantiomers due to low viscosity and high diffusivity of supercritical carbon dioxide, the main component in the SFC mobile phase. A method for enantiomeric separation and quantification of R- and S-amphetamine in urine was developed and validated using ultra-high performance supercritical fluid chromatography-tandem mass spectrometry (UHPSFC-MS/MS). Sample preparation prior to UHPSFC-MS/MS analysis was a semi-automatic solid phase extraction method. The UHPSFC-MS/MS method used a Chiralpak AD-3 column with a mobile phase consisting of CO2 and 0.2% cyclohexylamine in 2-propanol. The injection volume was 2 μL and run-time was 6 min. MS/MS detection was performed with positive electrospray ionization and two multiple reaction monitoring transitions (m/z 136.1 > 119.0 and m/z 136.1 > 91.0). The calibration range was 50-10,000 ng/mL for each enantiomer. The between-assay relative standard deviations were in the range of 3.7-7.6%. Recovery was 92-93% and matrix effects ranged from 100 to 104% corrected with internal standard. After development and validation, the method has been successfully implemented in routine use at our laboratory for both separation and quantification of R/S-amphetamine, and has proved to be a reliable and useful tool for distinguishing intake of R- and S-amphetamine in authentic patient samples.

Differences in combinations and concentrations of drugs of abuse in fatal intoxication and driving under the influence cases

BACKGROUND In toxicology, international classification systems focus on single intoxicants as the cause of death. It is, however, well known that very few drug related deaths are caused by a single substance and that information concerning the drug concentrations as well as the combinations of drugs are essential in order to ascertain the cause of death. The aim of the study was to assess whether those prone to fatal intoxications differ significantly from chronic drug users - in terms of demographics and drug exposure patterns. MATERIAL AND METHODS Fatal psychoactive drug intoxications in Norway during 2012, where a forensic autopsy including toxicological analysis were performed, were included. Analytical findings in blood were compared with concentrations in blood from apprehended drivers under the influence of drugs and ethanol (DUID) during the same time period. The opioid and benzodiazepine concentrations were assessed as morphine and diazepam equivalents, respectively, in order to compare concentrations across the different groups. RESULTS A total of 194 autopsy cases and 4811 DUID cases were included. Opioids were detected in around 90% of the drug intoxication cases, but in only 16% of the DUID cases. The number of substances detected in fatal intoxications was 4.9 compared to 2.6 in the DUID cases. The total opioid concentrations were significantly higher in the fatal intoxication cases compared to DUID cases (229ng/mL versus 56.9ng/mL morphine equivalents, respectively). Benzodiazepines were detected in 90% of the fatal cases. Only one fatal opioid mono-intoxication was found; a case with a very high methadone concentration (1238ng/mL). DISCUSSION Mono-intoxication with heroin was not seen in any of the fatal intoxications in Norway, and single drug intoxications were rare (1.5%). Fatal intoxications were caused by a combination of drugs with significantly more substances as well as higher total drug concentrations among the fatal cases compared to the DUID cases. The combination of opioids and benzodiazepines seemed to represent an increased risk of death. CONCLUSION The total load of drugs influence the degree of intoxication and the total concentration level must be considered, including the total number of substances. Our findings imply that international statistics regarding an opioid being the main intoxicant should have a shift in focus towards combinations of drugs (especially opioids and benzodiazepines) as a major risk factor for fatal drug overdoses.

Death by self-inflicted asphyxia with helium - First case reports from Norway and review of the literature

ABSTRACT An increasing number of asphyxia suicides by inhalation of inert gases have been reported from different parts of the world over the last decade. So far this phenomenon has not been described in our country. This article presents the first two case reports from Norway of presumed suicide by asphyxiation due to helium inhalation from a closed plastic bag over the head. In both cases a forensic autopsy, which included comprehensive toxicological analysis, was requested and performed. In the two cases death was attributed to asphyxia due to helium inhalation, and suffocation due to a plastic bag over the head and aspiration of gastric contents, respectively. Toxicological analysis revealed no findings contributing to the deaths. The absence of toxicological and autopsy findings to determine the cause of death in such cases may represent challenges of clinical and forensic significance. In contrast to the promotion of this method by euthanasia interest groups for the terminally ill reported suicides by helium asphyxiation primarily involve relatively young individuals suffering from psychiatric and/or substance use disorders, and not from terminal illness.