Thomas Z. Bosy

Regional distribution of methamphetamine in autopsied brain of chronic human methamphetamine users

We measured levels of methamphetamine and those of its metabolite amphetamine in 15 autopsied brain regions of 14 human methamphetamine users. Only slight regional differences were observed in drug concentrations among the brain areas. Although, some redistribution of the drugs probably occurred postmortem, these data suggest that methamphetamine might not be preferentially retained in dopamine-rich brain areas but is heterogenously distributed in brain of chronic human users of the drug. The possible pharmacological actions of methamphetamine in both dopamine-rich and poor brain areas of chronic drug users need to be considered.

Regional Distribution of Cocaine in Postmortem Brain of Chronic Human Cocaine Users

We measured concentrations of cocaine and its major metabolites (benzoylecgonine, ecgonine methylester, norcocaine, and cocaethylene) in 15 autopsied brain regions of 14 human chronic cocaine users. Only slight differences were observed in concentrations of cocaine and its metabolites amongst the examined brain areas. Although it is likely that some postmortem redistribution of the drug must have occurred, our data are consistent with the possibility that behaviorally relevant doses of cocaine are widely distributed throughout the brain of humans who use the drug on a chronic basis. Consideration should therefore be given to the possible pharmacological and toxicological actions of cocaine in both striatal and extra-striatal brain areas in human users of the drug.

A Fatality Involving AH-7921

A case is presented of a 19-year-old white male who was found dead in bed by a friend. While no anatomic cause of death was observed at autopsy, toxicological analysis of his blood identified AH-7921, a synthetic opioid. AH-7921 was isolated by liquid-liquid extraction into n-butyl chloride from alkalinized samples. Extracts were analyzed and quantified by gas chromatography mass spectrometry in selected ion monitoring mode. The heart blood had an AH-7921 concentration of 3.9 mg/L and the peripheral blood concentration was 9.1 mg/L. In addition to the blood, all submitted postmortem specimens including urine, liver, kidney, spleen, heart, lung, brain, bile and stomach content were quantified. The following concentrations of AH-7921 were reported: 6.0 mg/L in urine, 26 mg/kg in liver, 7.2 mg/kg in kidney, 8.0 mg/kg in spleen, 5.1 mg/kg in heart, 21 mg/kg in lung, 7.7 mg/kg in brain, 17 mg/L in bile and 120 mg/125 mL in the stomach content. The medical examiner reported that the cause of death was opioid intoxication and the manner of death was accident.

Analysis of Parent Synthetic Cannabinoids in Blood and Urinary Metabolites by Liquid Chromatography Tandem Mass Spectrometry

Synthetic cannabinoids emerged on the designer drug market in recent years due to their ability to produce cannabis-like effects without the risk of detection by traditional drug testing techniques such as immunoassay and gas chromatography-mass spectrometry. As government agencies work to schedule existing synthetic cannabinoids, new, unregulated and structurally diverse compounds continue to be developed and sold. Synthetic cannabinoids undergo extensive metabolic conversion. Consequently, both blood and urine specimens may play an important role in the forensic analysis of synthetic cannabinoids. It has been observed that structurally similar synthetic cannabinoids follow common metabolic pathways, which often produce metabolites with similar metabolic transformations. Presented are two validated quantitative methods for extracting and identifying 15 parent synthetic cannabinoids in blood, 17 synthetic cannabinoid metabolites in urine and the qualitative identification of 2 additional parent compounds. The linear range for most synthetic cannabinoid compounds monitored was 0.1-10 ng/mL with the limit of detection between 0.01 and 0.5 ng/mL. Selectivity, specificity, accuracy, precision, recovery and matrix effect were also examined and determined to be acceptable for each compound. The validated methods were used to analyze a compilation of synthetic cannabinoid investigative cases where both blood and urine specimens were submitted. The study suggests a strong correlation between the metabolites detected in urine and the parent compounds found in blood.

Detection of 25C-NBOMe in Three Related Cases

An accidental death associated with the use of the designer drug, 2-(4-chloro-2,5-dimethoxyphenyl)-N-(2-methoxybenzyl)ethanamine (25C-NBOMe), is reported. A 23-year-old Caucasian male experienced severe respiratory distress and died after being subdued by military law enforcement. At autopsy, remarkable findings upon internal examination included mild to moderate coronary atherosclerosis, biventricular dilation, mild right ventricular hypertrophy and bilateral pulmonary edema and congestion. The decedents blood contained no drugs, ethanol or other volatile compounds. Pseudoephedrine, nicotine and cotinine were detected in his urine. A LC-QTOF designer drug screen, employing a basic solid-phase extraction, was used to isolate 25C-NBOMe, 25C-NBOH and 2C-C from both blood and urine specimens. Quantitative analysis was performed by LC-MS-MS operating in multiple reaction monitoring mode. 25C-NBOMe and 2C-C were present in the blood (2.07 and 0.12 ng/mL) and in the urine (27.43 ng/mL and 0.38 ng/mL), respectively. 25C-NBOMe concentrations were determined by standard addition in the brain (19.10 ng/g), spleen (27.13 ng/g), lung (25.21 ng/g), liver (15.20 ng/g), kidney (25.06 ng/g) and gastric contents (30.24 µg total in 100 mL submitted). On the basis of decedent case history, autopsy and toxicological findings, the medical examiner ruled the cause of death as 25C-NBOMe toxicity temporally associated with excited delirium and forcible restraint. The manner of death was ruled accidental.

Ultrafast Screening of Synthetic Cannabinoids and Synthetic Cathinones in Urine by RapidFire-Tandem Mass Spectrometry

Screening for emerging drugs of abuse, specifically synthetic cathinones and synthetic cannabinoids, is difficult for high-throughput laboratories as immunoassay kits are often unavailable. Consequently, most laboratories employ liquid chromatography-tandem mass spectrometry (LC-MS-MS) screening, which can be complex and time consuming as these techniques may require involved sample preparation and lengthy analysis times. The increasing demand for novel psychoactive substance testing necessitates alternative screening methods that are sensitive, fast and versatile. The RapidFire tandem mass spectrometry system (RF-MS-MS) provides a rapid and highly specific screen for these emerging drugs of abuse with minimal sample preparation and an instrumental analysis time of <14 s per sample. Presented here are two RF-MS-MS screening methods used to analyze 28 emerging drugs of abuse, 14 synthetic cannabinoids and 14 synthetic cathinones, in urine with run times of 9 and 12.6 s, respectively. Sample preparation and hydrolysis were performed in a 96-well plate with one multiple reaction monitoring transition used for the identification of each compound. Eighteen thousand urine specimens were screened by liquid-liquid extraction followed by LC-MS-MS analysis, and the results were compared with those obtained using the RF-MS-MS screening method. The analytical data illustrate the advantages of the RF-MS-MS methods.

Evaluation of Abalone β-Glucuronidase Substitution in Current Urine Hydrolysis Procedures

This study examined the potential of abalone β-glucuronidase as a viable and cost effective alternative to current hydrolysis procedures using acid, Helix pomatia β-glucuronidase and Escherichia coli β-glucuronidase. Abalone β-glucuronidase successfully hydrolyzed oxazepam-glucuronide and lorazepam-glucuronide within 5% of the spiked control concentration. Benzodiazepines present in authentic urine specimens were within 20% of the concentrations obtained with the current hydrolysis procedure using H. pomatia β-glucuronidase. JWH 018 N-(5-hydroxypentyl) β-d-glucuronide was hydrolyzed within 10% of the control concentration. Authentic urine specimens showed improved glucuronide cleavage using abalone β-glucuronidase with up to an 85% increase of drug concentration, compared with the results obtained using E. coli β-glucuronidase. The JWH 018 and JWH 073 carboxylic acid metabolites also showed increased drug concentrations of up to 24%. Abalone β-glucuronidase was able to completely hydrolyze a morphine-3-glucuronide control, but only 82% of total morphine was hydrolyzed in authentic urine specimens compared with acid hydrolysis results. Hydrolysis of codeine and hydromorphone varied between specimens, suggesting that abalone β-glucuronidase may not be as efficient in hydrolyzing the glucuronide linkages in opioid compounds compared with acid hydrolysis. Abalone β-glucuronidase demonstrates effectiveness as a low cost option for enzyme hydrolysis of benzodiazepines and synthetic cannabinoids.

Case Report of Methylone, Oxymorphone and Ethanol in a Fatality Case with Tissue Distribution

It is reasonable to expect the presence of multiple drugs to present a complicated picture of toxicity. We report a fatal case involving a young man who purchased illicit drugs and knowingly consumed them. After consuming these drugs and going to sleep in his friends car, he was found unresponsive the next morning with no signs of physical violence. Drugs found in the peripheral blood at autopsy were oxymorphone, methylone and ethanol at concentrations of 0.106, 0.50 and 130 mg/dL, respectively. The levels of oxymorphone and methylone in peripheral blood were comparable to those observed in other reported fatalities. Cocaine and benzoylecgonine were detected in the urine but not in the blood. Measureable concentrations were also observed for oxymorphone and methylone in urine, liver, kidney and bile. The physical findings at autopsy included pulmonary edema. This is the only reported fatal case involving this combination of drugs encountered in our laboratory.

Multidrug Toxicity Involving Sumatriptan

A multidrug fatality involving sumatriptan is reported. Sumatriptan is a tryptamine derivative that acts at 5-HT(1B/1D) receptors and is used for the treatment of migraines. The decedent was a 21-year-old white female found dead in bed by her spouse. No signs of physical trauma were observed and a large number of prescription medications were discovered at the scene. Toxicological analysis of the central blood revealed sumatriptan at a concentration of 1.03 mg/L. Following therapeutic dosing guidelines, sumatriptan concentrations do not exceed 0.095 mg/L. Sumatriptan was isolated by solid-phase extraction and analyzed using liquid chromatography-tandem mass spectrometry in multiple reaction monitoring mode. A tissue distribution study was completed with the following concentrations measured: 0.61 mg/L in femoral blood, 0.56 mg/L in iliac blood, 5.01 mg/L in urine, 0.51 mg/kg in liver, 3.66 mg/kg in kidney, 0.09 mg/kg in heart, 0.32 mg/kg in spleen, 0.01 mg/kg in brain, 15.99 mg/kg in lung and 78.54 mg/45 mL in the stomach contents. Carisoprodol, meprobamate, fluoxetine, doxylamine, orphenadrine, dextromethorphan and hydroxyzine were also present in the blood at the following concentrations: 3.35, 2.36, 0.63, 0.19, 0.06, 0.55 and 0.16 mg/L. The medical examiner ruled the cause of death as acute mixed drug toxicity and the manner of death as accident.