Comparison of heroin and cocaine concentrations in saliva with concentrations in blood and plasma

Saliva is an alternate biological matrix for drug testing that has several advantages over more traditional fluids such as blood and urine. Collection is rapid, noninvasive, and relatively easy to obtain. Several reports have detailed the appearance of drugs of abuse in saliva, but few have compared the excretion profiles of drugs administered by different routes. In this study, subjects were administered three smoked and three intravenous doses of heroin in an ascending dose design. Blood and saliva were collected periodically after drug administration and analyzed by gas chromatography-mass spectrometry (GC-MS) for heroin, 6-acetylmorphine, and morphine. In a second study, subjects were administered a single, smoked dose of 40 mg cocaine base and an intravenous dose of 44.8 mg cocaine HO on separate occasions. Plasma and saliva were collected and analyzed by CC-MS for cocaine, anhydroecgonine methyl ester (AEME), and seven additional metabolites. Heroin and 6-acetylmorphine were detected in the first saliva sample collected (2 min) following drug administration by both routes. Peak heroin concentrations were achieved quickly, between 2 and 5 min after intravenous administration and at 2 min after smoke heroin. Peak heroin concentrations in saliva after smoking heroin base ranged from 3534 (2.6 mg) to 20,580 ng/mL (5.2 mg), and after intravenous administration, concentrations ranged from 6 (10 mg heroin HCl to 30 ng/mL (12 mg heroin HCl. Saliva concentrations of heroin declined rapidly after intravenous administration, reaching the limit of sensitivity of the assay (1 ng/mL) by 60 min. Heroin concentrations in saliva after smoking declined slowly; detection times ranged from 4 to 24 h. Cocaine was the major analyte detected in saliva and plasma after smoked and intravenous administration. Peak saliva cocaine concentrations after intravenous administration ranged from 428 to 1927 ng/mL (N = 7); after smoking, they ranged from 15,852 to 504,880 ng/mL (N = 7). Peak plasma cocaine concentrations after intravenous administration ranged from 122 to 442 ng/mL A = 7), and after smoking, concentrations ranged from 46 to 291 ng/mL A = 7). The thermal degradation product of cocaine, AEME, was detected in saliva but not in plasma after smoking. Peak saliva AEME concentrations were achieved at 2 min and ranged from 558 to 4374 ng/mL (N = 7). These are the first reported observations of heroin and metabolites in saliva following heroin smoking and of AEME in saliva after smoking cocaine base. The presence of AEME in saliva may be useful as a marker of the smoked route following cocaine administration.     

Crab clocks rewound

The effect of amphetamine (AMPH), codeine (COD), methamphetamine (MEPH), morphine (MORP), and benzoylecgonine (BE) on the binding of cocaethylene (CE) and cocaine (COC) to human serum in vitro was investigated by equilibrium dialysis at 4 degrees C. Each compound was added individually at concentrations of 500, 1,000, or 2,000 nM to pooled human serum containing COC or CE at 500 nM concentration. For COC, the addition of COD, MEPH, and CE enhanced serum binding whereas MORP and BE decreased it. Variable effects on COC binding were noted for AMPH. For CE, the addition of COD and COC generally increased binding whereas MORP decreased it. No appreciable effect on CE binding was observed after adding AMPH, MEPH, and BE. Except for CE, AMPH, and MEPH in the presence of COC, the binding of COC and CE tended to be less with 2,000 nM of each added drug than at lower concentrations of them, presumably because of mass-action displacement of COC and CE at the higher concentration. These findings should be clinically important because these drugs are frequently found together in patients.     

Trace elements in spinocerebellar degeneration

Cocaine and cocaethylene concentrations in blood and tissues at early stages postmortem (0-6 h) were investigated using alcohol-treated rats. Gas chromatography/mass spectrometry following a liquid/liquid extraction procedure was employed to detect these drugs. Calibration curves showed good linearity in the range of 0 to 2,500 ng/mL with correlation coefficients of 0.9999 and 0.9998 for cocaine and cocaethylene, respectively. In a group treated with cocaine and ethanol orally, the liver lost over 25% of the cocaine present at death after 1 h. Conversely, the hepatic cocaethylene concentrations at this time reached more than twice those at death. Thereafter, the hepatic concentrations of cocaine and cocaethylene were maintained at a constant level until 6 h postmortem. Similar results were obtained with rats given cocaine intramuscularly. No changes in the cocaine and cocaethylene concentrations in any other tissues during the 6-h of postmortem period were observed. The forensic pathologist and toxicologist should be aware of these phenomena when selecting postmortem specimens for the analysis of cocaine and cocaethylene and take them into account when interpreting the results.     

Improved long-term stability of blood cocaine in evacuated collection tubes

A study was undertaken to determine if a relatively minor modification of our existing specimen collection tubes could enhance the long-term stability of blood cocaine. We added cocaine, benzoylecgonine (BE) and ethanol to whole sheep blood in glass tubes that were prepared to contain one of several combinations of preservatives and anticoagulant. On day 1 and at intervals of up to one year, the drugs were measured by gas chromatography-mass spectrometry (cocaine and BE) or headspace gas chromatography (ethanol). Storage of blood containing 200 ng/mL cocaine at 4 degrees C for one year resulted in 100% loss of the drug using our normal 10 mL specimen collection tubes containing 100 mg sodium fluoride and 20 mg potassium oxalate. The substitution of oxalic acid for potassium oxalate reduced this loss to 76% without any significant effect on the benzoylecgonine or ethanol concentrations. Further addition of 10 mg echothiophate iodide, a quaternary ammonium compound, brought the cocaine loss down to 60% of the original concentration by one year. Further work will be required to determine if oxalic acid and/or echothiophate iodide could be used in blood collection vials intended for forensic toxicological purposes without any detrimental effect on other assays.     

Time course of cocaine in rabbit hair

The accurate interpretation of analytical results from hair testing for drugs of abuse continues to be a complex and difficult problem since many questions still remain unanswered. In this paper an animal model was developed to ascertain the time course for the appearance and disappearance of cocaine and its metabolite benzoylecgonine (BE) in hair. Female Fauve Bourgogne red-haired rabbits (n = 6) were intraperitoneally administered a single dose of cocaine at 5 mg/kg. Animal hair was shaved just before drug administration and the newly grown back hair was subsequently shaved and collected daily over a period of two weeks. Samples were analyzed for cocaine and BE by gas chromatography-mass spectrometry (GC-MS). The profiles were quite similar for parent drug and metabolite. Cocaine and BE appeared in the first sampling (day 1), with peak concentration appearing that same day. 1.01 ng/mg and 0.51 ng/mg for cocaine and BE, respectively. Levels declined rapidly on day 2, remaining detectable for ten days after drug administration. This study demonstrates that the initial incorporation of cocaine compounds in rabbit hair is very rapid (24 h). A small fraction of the drug is detected ten days after exposure, at a time when concentrations in other biological specimens (blood or urine) are not detectable.     

Direct determination of benzoylecgonine in serum by EMIT d.a.u. cocaine metabolite immunoassay

An EMIT d.a.u. immunoassay for urine testing was applied on the Syva ETS Plus analyzer for the detection of the cocaine metabolite, benzoylecgonine (BE), in human serum. Serum was analyzed without prior extraction, concentration, or matrix modification. Calibrators and serum controls were prepared from EMIT d.a.u. calibrators that were reconstituted and diluted with EMIT Tox serum calibrator. The assay cutoff concentration for BE was 50 ng/mL. The within-run and between-run precisions of the assay were both less than 5%. Analysis of 162 patient serums yielded 43 BE positive results. All EMIT positive serum BE results were confirmed by gas chromatography-mass spectrometry. All patients with positive BE serums also had BE positive urine samples. Serum bilirubin and triglycerides as high as 38 mg/dL and 319 mg/dL, respectively, did not interfere with the assay. Modification of the EMIT urine assay allowed for a simple, rapid, and reliable method for the detection of BE in serum.     

Detection of cocaine exposure in the neonate. Analyses of urine, meconium, and amniotic fluid from mothers and infants exposed to cocaine

Cocaine and its metabolites were measured in urine, meconium, and amniotic fluid specimens collected from 30 maternal-infant pairs with histories of prenatal cocaine use. Cocaine, benzoylecgonine, and ecgonine methyl ester were measured by isotope dilution gas chromatography-mass spectrometry. Mothers were interviewed at delivery regarding their cocaine use during pregnancy. There was qualitative agreement between the results of drug determinations in maternal urine, amniotic fluid, infant urine, and meconium. Although all of the mothers in this study admitted to using cocaine during their pregnancy, cocaine or its metabolites were detected only in the 20 cases in which cocaine was used within 3 weeks before delivery. We conclude that when sufficiently sensitive analytic methods are used, maternal urine, infant urine, and meconium analyses yield equivalent results for detection of prenatal cocaine exposure. Importantly, neither meconium nor urinary drug measurements detected cocaine exposure when the last reported use was prior to 3 weeks before delivery.     

Comparison of the reinforcing and anxiogenic effects of intravenous cocaine.

People report that ethanol improves the experience produced by cocaine. This effect may be attributable to cocaethylene (CE), a cocaine metabolite formed only in the presence of ethanol. To test this, rats were trained to run an alley for a single intravenous dose of either cocaine (0.5–2.0 mg/kg) or an equimolar dose of CE (0.75–2.88 mg/kg). The rats' start latency and running speed measured the reinforcing effects of the drugs and the number of times rats approached but failed to enter the goal box (i.e., approach-avoidance retreats) indexed anxiety. Rats reinforced with CE had shorter start latencies and faster running speeds and exhibited fewer "retreats" than cocaine-reinforced rats. These results suggest that CE is more reinforcing and less anxiogenic than cocaine and hence may account for the combined effects of cocaine and ethanol in humans. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Changes in substance P in the jejuna of rats after burns

Ecgonine is formed by the hydrolysis of both ester linkages in cocaine and is of interest as both a potential metabolite and a postmortem and postcollection artifact in biological specimens. Its extreme polarity and water solubility make its isolation very difficult, particularly from complex matrices such as postmortem whole blood. This procedure describes an initial protein precipitation followed by two derivatization steps, the first of which is to propylate organic acids and primary and secondary amines and the second of which is to form the p-nitrobenzoyl ester of organic alcohols. The resultant derivatized products are then subjected to cleanup using a standard extraction with n-butyl chloride as the solvent. Analysis of the extracts is performed by gas chromatography-mass spectrometry using deuterated internal standards, although the adducts would also be suitable for analysis by high-performance liquid chromatography. Significant quantities of ecgonine have been identified in postmortem whole blood samples using this method.     

The incidence of sudden unexpected death in epilepsy (SUDEP) in South Dublin and Wicklow

The binding of cocaine (COC) and cocaethylene (CE) to whole human liver homogenates in vitro was studied by equilibrium dialysis. Drugs were measured by high-pressure liquid chromatography. Up to 32% of COC and up to 43% of CE were bound. Scatchard analysis suggested a high-affinity, low-capacity binder for both COC (Ka, 4.69 x 10(4) L/mol; Bo, 1.08 x 10(-5) mol/L) and CE (Ka, 4.38 x 10(4) L/mol; Bo, 1.54 x 10(-5) mol/L). In addition, low-affinity, high-capacity binders for COC (Ka, 2.93 x 10(3) L/mol; Bo, 1.32 x 10(-4) mol/L) and CE (Ka, 6.50 x 10(3) L/mol; Bo, 1.11 x 10(-4) mol/L) were noted. Finally, for both compounds, very low-affinity, high-capacity binding, which was likely nonspecific in nature, was defined as follows: COC, Ka, 8.00 x 10(2) L/mol; Bo, 5.45 x 10(-4) mol/L and CE, Ka, 2.10 x 10(3) L/mol; Bo, 3.71 x 10(-4) mol/L. The binding profiles of COC and CE in liver were compared with those in human serum and placenta studied previously by this laboratory.     

Cell density-dependent changes in the insulin action pathway: evidence for involvement of protein-tyrosine phosphatases

Three gas chromatographic procedures for the determination of ethanol in postmortem blood using alternative internal standards to n-propanol are presented: a direct injection procedure using t-butanol, and two headspace methods using t-butanol and methyl ethyl ketone. t-Butanol and methyl ethyl ketone were well resolved from ethanol, acetone, methanol and other commonly observed putrefactive volatiles using direct injection or headspace analysis. CVs for the direct injection method were below 5% for ethanol and below 10% for the other volatiles. The lower limits of detection (LOD) were 25-50 mg/L. The CVs for the headspace methods were below 5% for ethanol and below 6% for the other volatiles. The LODs were 10 mg/L using either t-butanol or methyl ethyl ketone as internal standards. The use of t-butanol or methyl ethyl ketone as alternatives to n-propanol avoids the possibility of error in the quantitation of ethanol due to the presence of n-propanol and allows for the identification of other volatiles that may aid in distinguishing antemortem ingestion from postmortem production of ethanol.     

A case of axonal form of Guillain-Barré syndrome associated with anti-GM1b IgG antibody following Penner 4 Campylobacter jejuni infection

A 48-year-old man with an extensive history of alcoholism was found dead at home. He was lying face down on a carpet. There was evidence of gastric aspiration at autopsy and histologic examination. The distribution of ethanol was very unusual (concentrations in mg/100 mL or mg/100 g): femoral blood, 257 and 273 (two samples); heart blood, 643; vitreous humor, 763; urine, 84; bile, 616; liver, 250; and gastric, 4660 (2470 mg/53 g). In addition, this man ingested isopropanol, and, according to the history, may also have ingested acetone in the form of nail polish remover. The distribution of both isopropanol and acetone was as expected, which was approximately in proportion to the aqueous content of the respective tissues. It is proposed that agonal or postmortem aspiration of the ethanol-rich vomitus and postmortem fermentation could account for the apparently elevated concentrations of ethanol in heart blood and bile. The elevated vitreous ethanol could be explained if ethanol diffused across the eye in the agonal phase or postmortem from gastric aspirate in the carpet. The relatively low urinary ethanol concentration would be consistent with a recent binge-drinking episode, which allowed only a limited time period for excretion into an already partially full, but relatively ethanol-free, bladder.     

A fatal drug interaction between clozapine and fluoxetine

A case is presented of a fatal drug interaction caused by ingestion of clozapine (Clozaril) and fluoxetine (Prozac). Clozapine is a tricyclic dibenzodiazepine derivative used as an "atypical antipsychotic" in the treatment of severe paranoid schizophrenia. Fluoxetine is a selective serotonin reuptake inhibitor used for the treatment of major depression. Clinical studies have proven that concomitant administration of fluoxetine and clozapine produces increased plasma concentrations of clozapine and enhances clozapine's pharmacological effects due to suspected inhibition of clozapine metabolism by fluoxetine. Blood, gastric, and urine specimens were analyzed for fluoxetine by gas chromatography/mass spectrometry (GC/MS) and for clozapine by gas-liquid chromatography (GLC). Clozapine concentrations were: plasma, 4.9 micrograms/mL; gastric contents, 265 mg; and urine, 51.5 micrograms/mL. Fluoxetine concentrations were: blood, 0.7 microgram/mL; gastric contents, 3.7 mg; and urine 1.6 micrograms/mL. Norfluoxetine concentrations were: blood, 0.6 microgram/mL, and none detected in the gastric contents or urine. Analysis of the biological specimens for other drugs revealed the presence of ethanol (blood, 35 mg/dL; vitreous, 56 mg/dL; and urine 153 mg/dL) and caffeine (present in all specimens). The combination of these drugs produced lethal concentrations of clozapine and high therapeutic to toxic concentrations of fluoxetine. The deceased had pulmonary edema, visceral vascular congestion, paralytic ileus, gastroenteritis and eosinophilia. These conditions are associated with clozapine toxicity. The combined central nervous system, respiratory and cardiovascular depression of these drugs was sufficient to cause death. The death was determined to be a clozapine overdose due to a fatal drug interaction.     

On-line dialysis solid-phase extraction coupled to capillary electrophoresis

A fully automated dialysis solid-phase extraction (SPE) sample preparation procedure is coupled on-line to capillary electrophoresis (CE) for the first time. The system is used to determine sulfonamides in serum and urine. The dialysis unit serves to remove proteins and particulate matter. Reconcentration of the analytes is performed with a small SPE column while (in)organic salts and other interferences are removed simultaneously. Finally, the analytes are desorbed and injected, via a homemade interface, into the CE system. Limits of detection (LOD) of 0.05-0.1 and 0.05-0.3 microg/mL are obtained in urine and serum, respectively. The within-day and between-day precisions are in the range of 2-6% and 3-8%, respectively, for a concentration of five times the LOD. The dialysis SPE-CE system was used over a period of six months for the analysis of over 500 serum and urine samples without problems such as clogging of the CE capillary or SPE column.     

Human energy and work in a European village

In a mouse model of transplacental cocaine exposure we have demonstrated alterations in brain structure and function of offspring including disturbances of brain growth, disruption of neocortical cytoarchitecture, and transient as well as persistent behavioral deficits. One mechanism by which cocaine may alter fetal brain development is through cocaine-induced alpha-adrenergic-mediated (uterine) arterial vasoconstriction. In this study pregnant Swiss Webster (SW) mice were injected with cocaine HCl (20 or 40 mg/kg, SC) without any changes evident in mean arterial blood pressure (MAP) measurements. These physiology results suggest that in our mouse model, cocaine's transplacental effects on the fetus are not due to cocaine-induced maternal vasoconstriction, nor concomitant hypoperfusion of the fetus. In a separate series of experiments, pregnant SW dams were administered cocaine HCl at 40 mg/kg/day (COC 40), 20 mg/kg/day (COC 20), or 10 mg/kg/day (COC 10) [SC, divided in two daily doses, from embryonic day (E) 8 to E17 inclusive]. Additional groups of cocaine-treated dams were administered phentolamine (5 mg/kg, SC), a short-acting alpha-adrenergic antagonist, 15 min prior to each cocaine dose (Phent COC 40, Phent COC 20, Phent COC 10). Animals born to Phent COC 40 dams demonstrated transient postnatal brain growth retardation and behavioral deficits in first-order conditioning of P9 mice comparable to mice born to COC 40 dams, which received the same regimen of cocaine injections without phentolamine pretreatment. Like COC 40 offspring, Phent COC 40 offspring also demonstrated a persistent deficit in the blocking paradigm. The behavioral and growth findings confirm and extend the physiology data, and imply that in our rodent model, alpha-adrenergic mechanisms (including maternal vasoconstriction) are unlikely to mediate these toxic effects of transplacental cocaine exposure on developing brain.     

Perflubron as a gastrointestinal MR imaging contrast agent in the pediatric population

A microplate enzyme immunoassay (EIA) for the detection of lysergic acid diethylamide (LSD) in human urine was developed. The assay kit is designed around an LSD derivative coated on the wall of microplate wells with preservatives and stabilizers. Sample and rabbit anti-LSD are added to the microplate well. The immobilized LSD and LSD present in specimens compete for the opportunity to bind to the anti-LSD antibodies. An anti-rabbit antibody labeled with horseradish peroxidase is used to provide the assay signal, which is inversely proportional to the concentration of LSD in the sample. The assay requires a 25-microL urine sample and three consecutive incubation periods of 60, 30, and 30 min at room temperature. The assay was tested with a variety of drugs, including ergot alkaloids spiked into drug-free urine at up to 100,000 ng/mL without cross-reaction. Nor-LSD was shown to cross-react between 16% and 28%, depending on its concentration. Of the other compounds tested, only ergonovine demonstrated slight cross-reactivity at approximately 0.0008%. The assay is designed to be used with a qualitative cutoff of 0.5 ng/mL. Precision testing at 0.5 ng/mL gave a coefficient of variation (CV) of 6% based on 20 replicates. The CV at 0.375 ng/mL (cutoff, -25%) was 5.2% and at 0.625 ng/mL was 6.6%. Precision at other concentrations within the range of the calibration curve gave similar results both intra- and interassay. Clinical performance of the assay was compared with that of a commercial radioimmunoassay (RIA). Comparable performance was observed with both methods, each screening a total of 458 samples as negative and 17 samples as positive relative to a 0.5 ng/mL cutoff. The EIA found an additional three positive samples that were negative by RIA. The EIA is suitable for the screening of urine samples for the presence of LSD. Preliminary indications are that the assay is also suitable for use with whole blood specimens. The assay can be performed manually or be fully automated and without the need for radioactivity; it can be used in any laboratory.     

Determination of common drugs of abuse in body fluids using one isolation procedure and liquid chromatography--atmospheric-pressure chemical-ionization mass spectromery

A method for determining opiate agonists (morphine, morphine-3-glucuronide, morphine-6-glucuronide, 6-monoacetylmorphine, codeine, codeine-6-glucuronide, dihydrocodeine, dihydromorphine, buprenorphine, methadone, tramadol, and ibogaine), cocaine and its metabolites (benzoylecgonine and ecgonine methyl ester) and lysergic acid diethylamide in serum, blood, urine and other biological matrices is presented. Aliquots (0.5-1.5 mL) of biological fluids were spiked with appropriate deuterated internal standards and extracted using a common solid-phase extraction method (C18 cartridges). The extracts were subjected to liquid chromatographic-atmospheric-pressure chemical-ionization mass spectrometric examination using selected ion monitoring procedures. These procedures were developed after analysis of full-scan mass spectra of examined compounds. The extraction method appeared very universal; the recoveries were high for almost all drugs and the extracts were very clean. The procedure was applied for routine forensic casework.     

Validation of an automated microplate enzyme immunoassay for screening of postmortem blood for drugs of abuse

The objective of this study was to compare the sensitivity and specificity of an enzyme immunoassay employing antibodies bound to a microtiter plate (MPEIA) with those of two radioimmunoassays for screening postmortem blood from selected coroner's cases for drugs of abuse. The radioimmunoassays were a coated-tube radioimmunoassay (CTRIA) and a double antibody radioimmunoassay (DARIA). Specimens consisted of 260 postmortem blood specimens from coroner's cases. Immunoassay results (positive or negative) were compared with confirmed results on those cases by gas chromatography-mass spectrometry, alone or in combination with gas-liquid chromatography using either a nitrogen-phosphorus or flame-ionization detector. Sensitivity was calculated as the true-positive rate using chromatographic confirmation as the reference standard. Specificity was calculated as the true-negative rate. Sensitivity and specificity were calculated for 5-7 potential cutoff concentrations for the drug classes opiates, amphetamines, cocaine and metabolites, and barbiturates. For opiates, the sensitivity and specificity were 99% and 93%, respectively, for the MPEIA at a cutoff of 20-ng/mL morphine, compared with 94% and 96% for the CTRIA at a cutoff of 5-ng/mL morphine and > 99% and 96% for the DARIA at 20-ng/mL morphine. For cocaine and metabolites, the sensitivity and specificity were 96% and 93%, respectively, for the MPEIA at 50-ng/mL benzoylecgonine, compared with 93% and 96% for CTRIA at 50-ng/mL benzoylecgonine and 98% and 97% for the DARIA at 50-ng/mL benzoylecgonine. For amphetamines, the sensitivity and specificity were >99% and 91%, respectively, for the MPEIA at 25-ng/mL methamphetamine, compared with 93% and 86% for the CTRIA at 25-ng/mL methamphetamine and 83% and 89% for the DARIA at 50-ng/mL methamphetamine. For barbiturates, the sensitivity and specificity were > 99% and 92%, respectively, for the MPEIA at 50-ng/mL secobarbital, compared with 91% and 87% for the CTRIA at 500-ng/mL secobarbital and 79% and 95% for the DARIA at a cutoff of 1000-ng/mL phenobarbital.     

Buprenorphine-related deaths among drug addicts in France: a report on 20 fatalities

This paper reports a series of 20 fatalities involving a high-dose, sublingual buprenorphine (BUP) formulation recently marketed in France for the substitutive therapy of opiate addicts. The files were recorded over a 16-month period from five different urban areas in France. All subjects but one were male, aged 14-48 (mean 26.6). BUP and its primary metabolite norbuprenorphine (norBUP) were assayed in postmortem fluids and viscerae by HPLC-MS. Blood levels for BUP and norBUP ranged from 1.1 to 29.0 ng/mL (mean 8.4 ng/mL) and 0.2 to 12.6 ng/mL (mean 2.6 ng/mL), respectively, that is, within or slightly over the therapeutic range. BUP exhibited extensive tissue distribution, with average postmortem concentrations of 6.0, 35.0, 45.5, and 80.0 ng/g in the myocardium, kidney, brain, and liver, respectively. In blood, as in viscerae, norBUP levels were generally lower than BUP. The highest concentrations were found in the bile for both BUP (range 575-72,650 ng/mL) and norBUP (range 41-30,000 ng/mL). Therefore, bile may represent a sample of choice for postmortem screening. BUP was identified in 9 of the 11 hair samples assayed at concentrations ranging from 6 to 597 ng/g (mean 137 ng/g), whereas norBUP was never detected. Intravenous injection of crushed tablets, a concomitant intake of psychotropics (especially benzodiazepines), and the high dosage of the BUP formulation available in France appear to be the major risk factors for such fatalities.