benzoylecgonine and Drug-Overdose

In this case report, we present an evaluation of the distribution of postmortem concentrations of butyr-fentanyl in a fatality attributed principally to the drug. A man who had a history of intravenous drug abuse was found unresponsive on the bathroom floor of his home. Drug paraphernalia was located on the bathroom counter. Toxicology testing, which initially screened positive for fentanyl by enzyme-linked immunosorbent assay, subsequently confirmed butyr-fentanyl, which was then quantitated by gas chromatography-mass spectrometry-specific ion monitoring (GC-MS SIM) analysis following liquid-liquid extraction. The butyr-fentanyl peripheral blood concentration was quantitated at 58 ng/mL compared with the central blood concentration of 97 ng/mL. The liver concentration was 320 ng/g, the vitreous was 40 ng/mL, the urine was 670 ng/mL and the gastric contained 170 mg. Acetyl-fentanyl was also detected in all biological specimens tested. Peripheral blood concentration was quantitated at 38 ng/mL compared with the central blood concentration of 32 ng/mL. The liver concentration was 110 ng/g, the vitreous was 38 ng/mL, the urine was 540 ng/mL and the gastric contained <70 mg. The only other drug detected was a relatively low concentration of benzoylecgonine. The cause of death was certified as acute butyr-fentanyl, acetyl-fentanyl and cocaine intoxication, and the manner of death was certified as accident.

Topics: Adult; Analgesics, Opioid; Cocaine; Drug Overdose; Enzyme-Linked Immunosorbent Assay; Fatal Outcome; Fentanyl; Forensic Toxicology; Gas Chromatography-Mass Spectrometry; Humans; Liquid-Liquid Extraction; Male; Opioid-Related Disorders; Substance Abuse, Intravenous

Cocaine is one of the most addictive drugs without a U.S. Food and Drug Administration (FDA)-approved medication. Enzyme therapy using an efficient cocaine-metabolizing enzyme is recognized as the most promising approach to cocaine overdose treatment. The actual enzyme, known as RBP-8000, under current clinical development for cocaine overdose treatment is our previously designed T172R/G173Q mutant of bacterial cocaine esterase (CocE). The T172R/G173Q mutant is effective in hydrolyzing cocaine but inactive against benzoylecgonine (a major, biologically active metabolite of cocaine). Unlike cocaine itself, benzoylecgonine has an unusually stable zwitterion structure resistant to further hydrolysis in the body and environment. In fact, benzoylecgonine can last in the body for a very long time (a few days) and, thus, is responsible for the long-term toxicity of cocaine and a commonly used marker for drug addiction diagnosis in pre-employment drug tests. Because CocE and its mutants are all active against cocaine and inactive against benzoylecgonine, one might simply assume that other enzymes that are active against cocaine are also inactive against benzoylecgonine. Here, through combined computational modeling and experimental studies, we demonstrate for the first time that human butyrylcholinesterase (BChE) is actually active against benzoylecgonine, and that a rationally designed BChE mutant can not only more efficiently accelerate cocaine hydrolysis but also significantly hydrolyze benzoylecgonine in vitro and in vivo. This sets the stage for advanced studies to design more efficient mutant enzymes valuable for the development of an ideal cocaine overdose enzyme therapy and for benzoylecgonine detoxification in the environment.

Topics: Animals; Butyrylcholinesterase; Cocaine; Cocaine-Related Disorders; Drug Overdose; Humans; Inactivation, Metabolic; Rats

The presence of cocaine (COC) in fluids or tissues does not prove that death was due to drug consumption and the interpretation of postmortem concentrations is more complex than attempts at making such correlations in the living. The purpose of this study was to investigate the distribution of cocaine and its metabolite benzoylecgonine in brain and compare with whole blood and vitreous humour. The distribution in three brain structures (prefrontal cortex, basal ganglia and cerebellum) was homogeneous. There is a strong correlation for cocaine concentrations between vitreous humour and brain, vitreous humour and whole blood, and whole blood and brain in overdose cases. In addition, the comparison of COC/benzoylecgonine (BE) ratios in different experimental specimens proved to be more appropriate for evaluating cocaine-related death than individual drug values. These findings suggest that the comparison of cocaine levels in different compartments is essential to assess the cause of death.

Topics: Adolescent; Adult; Analysis of Variance; Basal Ganglia; Cerebellum; Cocaine; Drug Overdose; Female; Forensic Toxicology; Humans; Male; Middle Aged; Narcotics; Postmortem Changes; Prefrontal Cortex; Vitreous Body; Young Adult

In recent years there has been an increase interest in cocaine-related death reflecting the rising trend in cocaine use in Europe. Nevertheless it is still now very difficult to attribute a death to cocaine. We can affirm that cocaine can be responsible for the cause of death only when there is a reasonably complete understanding of the circumstances or facts surrounding the death. Isolated blood cocaine levels are not enough to assess lethality, and should be always considered and evaluated in relation to concentrations of cocaine and benzoylecgonine concentrations in body tissue compartments, especially in brain and blood. We have reanalyzed all of our cocaine-related cases from 1990 to 2005, applying the methodology used by Spielher and Reed over 30 years ago. Our aim was to try to validate this model and verify its applicability and effectiveness after 20 years.

Topics: Brain; Brain Chemistry; Cocaine; Dopamine Uptake Inhibitors; Drug Overdose; Forensic Toxicology; Humans

Fatalities associated with fentanyl hydrochloride are increasingly seen in Massachusetts. Between September 2005 and November 2006, 5009 medicolegal investigations associated 107 deaths with licit or illicit fentanyl use, along with a co-detection of an opiate/opioid or cocaine/benzoylecognine, or both. Deaths associated with illicit fentanyl use occur in younger people (39.4 vs. 61.5 years) with higher fentanyl (17.1 ng/mL vs. 4.4 ng/mL) and lower morphine (76.9 ng/mL vs. 284.2 ng/mL) postmortem blood concentrations, and more frequent cocaine co-intoxication (65% vs. 3%), than deaths associated with illicit fentanyl use. A wide range of postmortem blood concentrations of fentanyl was detected (trace-280 ng/mL), with a minimum concentration of 7 ng/mL of fentanyl strongly associated with illicit use of fentanyl in poly-drug cases. The most commonly detected opiates/opioids in illicit fentanyl users were: morphine (29%), oxycodone (14.5%), and methadone (14.5%). Ethanol, cannabinoids, diazepam, citalopram, and diphenhydramine were each detected in greater than 10% of the licit fentanyl cases. Most fentanyl abusers died at their own home and their deaths were most often classified as accidental. Mapping of primary residences of decedents revealed conspicuous clustering of the illicit fentanyl use cases, as opposed to the random pattern in licit use cases. Fentanyl misuse is a public health problem in Massachusetts.

Topics: Accidents; Adult; Aged; Aged, 80 and over; Cannabinoids; Cause of Death; Central Nervous System Depressants; Cocaine; Dopamine Uptake Inhibitors; Drug Overdose; Ethanol; Female; Fentanyl; Forensic Toxicology; Gas Chromatography-Mass Spectrometry; Humans; Male; Middle Aged; Narcotics; Selective Serotonin Reuptake Inhibitors; Substance-Related Disorders

Analyzing hair for many substances can be tedious and expensive, and a rapid screening method should prove helpful. Generally, screening has been performed using immunological tests, mainly in workplace drug testing, where the number of samples has been high. The aim of this study was to develop an LC-MS-MS method for the simultaneous analysis of several drugs of abuse in human hair as an alternative to immunological screening tests. In 75 randomly selected autopsy cases, hair was analyzed in addition to the usual specimens of blood and urine. The method included nicotine, cotinine, morphine, codeine, 6-acetylmorphine, ethylmorphine, amphetamine, methamphetamine, MDA, MDMA, benzoylecgonine, cocaine, 7-aminoflunitrazepam and diazepam. The LC-MS-MS analysis was performed on a SCIEX API 2000 MS-MS instrument equipped with an electrospray interface. To 20-50 mg of hair, 0.5 ml of mobile phase A (acetonitril:methanol:20 mM formate buffer, pH 3.0 (10:10:80)) and 25 microl of internal standard were added and the sample was incubated in a water bath at 37 degrees C during 18 h. Using a threshold of 20 ng/sample, equivalent to 1 ng/mg if 20mg hair is used, 26 positive results were found in 16 cases. Three of the 26 positive detections could not be confirmed by GC-MS. Two of the cases were not previously known as drug users. Of the 59 negative cases, only one case had a positive blood sample showing 0.01 and 0.07 microg/g femoral blood of 6-acetylmorphine and morphine, respectively. This might indicate drug abstinence resulting in decreased tolerance or even a "first time" use of heroin resulting in death. We conclude that the use of hair analysis in postmortem cases can reveal both unknown drug use, as well as confirm a period of drug abstinence prior to an acute fatal overdose. The proposed LC-MS-MS method showed high sensitivity, was very easy to perform and seemed appropriate for screening purposes.

Topics: Amphetamines; Anti-Anxiety Agents; Central Nervous System Stimulants; Chromatography, Liquid; Cocaine; Cotinine; Diazepam; Dopamine Uptake Inhibitors; Drug Overdose; Flunitrazepam; Forensic Medicine; Gas Chromatography-Mass Spectrometry; Hair; Humans; Morphine; Morphine Derivatives; Narcotics; Nicotine; Nicotinic Agonists; Reproducibility of Results; Spectrometry, Mass, Electrospray Ionization; Substance Abuse Detection

Hair samples of eight postmortem cases were analyzed in segments of 1 to 3 cm for cocaine, benzoylecgonine and cocaethylene. Samples were prepared for analysis by digestion in 0.1 M HCl and subsequent extraction with mixed-mode solid-phase extraction columns. Measurement was made by reversed-phase, narrow-bore HPLC and fluorescence detection using two laboratory-made internal standards. The concentrations were in the region of 0.29-316 ng/mg of hair for cocaine, 0.43-141 ng/mg of hair for benzoylecgonine and 0.93-1.83 ng/mg of hair for cocaethylene. All eight investigated cases had cocaine-positive segments. In six of the cases, all segments were positive, suggesting regular cocaine use and two showed in-between negative segments indicating an interruption or a change of the abuse intensity. The results showed a second, remarkable observation, i.e. enormous concentration differences (factor >150) for both cocaine and benzoylecgonine between the different subjects. Furthermore, interindividual cocaine/benzoylecgonine ratios ranged from 0.02 to 8.43. We believe these observations could in part be attributed to both some of the still existing limitations in the analytical approach(es), especially the mandatory hair washing steps, and in our still too limited knowledge of the hair incorporation processes. Nevertheless, in some cases, segmental analysis proved to be an important tool to distinguish, together with postmortem examination, deadly chronic abuse from single acute drug overdosage.

Topics: Chromatography, High Pressure Liquid; Cocaine; Cocaine-Related Disorders; Drug Overdose; Forensic Medicine; Hair; Humans

A procedure is presented for the simultaneous identification and quantification of morphine (MOR), codeine (COD), ethylmorphine (EM), 6-monoacetylmorphine (6-MAM), cocaine (COC), benzoylecgonine (BZE), ecgonine methylester (EME) and cocaethylene (CE), contained in the hair of opiates and cocaine addicts. The method involves decontamination in dichloromethane, pulverization in a ball mill, heat-acid hydrolysis, addition of deuterated internal standards, liquid-liquid extraction and gas chromatography/mass spectrometry (GC/MS) after silylation. The limit of detection (LOD) was approximately 0.1-0.8 ng/mg for each drug, using a 30-mg hair sample. The method is reproductible, with a coefficient of variation (CV) of approximately 8-17%. Cocaine and 6-monoacetylmorphine were the major compounds detected in cases of cocaine (14 cases) and heroin (68 cases) intake. Concentrations were in the range 0.4-78.4 ng/mg (COC), 0.0-36.3 ng/mg (BZE), 0.0-1.6 ng/mg (EME), 0.0-2.1 ng/mg (CE), 0.0-84.3 ng/mg (6-MAM), 0.2-27.1 ng/mg (MOR) and 0.1-19.6 ng/mg (COD). An application in forensic sciences, involving multi-sectional analysis, is given.

Topics: Adolescent; Adult; Calibration; Cocaine; Dopamine Uptake Inhibitors; Drug Overdose; Ethylmorphine; Female; Forensic Medicine; Gas Chromatography-Mass Spectrometry; Hair; Heroin; Humans; Male; Morphine; Morphine Derivatives; Narcotics; Reproducibility of Results

Intestinal ischemia induced by cocaine abuse is a rare condition. To this date, only three cases have been described. The diagnosis of bowel ischemia should be suspected whenever a cocaine addict has severe abdominal pain. A pathological examination of the resected bowel segment was performed in one case, and the diagnosis was confirmed microscopically. However, the existence of pathologic alterations of the intestinal vessels was not confirmed. Why the intestinal injury is segmental and whether it is related to the dose ingested, the administration route, or the combination of cocaine with alcohol, caffeine, or marijuana remain unclear. The authors report one fatal case associated with cocaine-alcohol overdose. The postmortem examination demonstrated the existence of segmental intestinal ischemia. Microscopic study failed to demonstrate thrombosis in the mesenteric vessels; however, we found an unusual lesion affecting the arterioles located in the intestinal submucosa of the hemorrhagic areas.

Topics: Adult; Alcoholic Intoxication; Arterioles; Cocaine; Drug Overdose; Ethanol; Humans; Intestine, Small; Ischemia; Male; Substance-Related Disorders