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In clinical and forensic toxicology, hair analysis offers a larger window for detecting drug exposure than blood or urine. Drug measurements are generally carried out using a segmented lock of hair, but few articles report the use of a single hair to document drug exposure. Nevertheless, single hair analysis can be very useful, particularly if only small amounts of biological matrices are available. More data on analyzing new synthetic opioids (NSOs) in hair are needed to help interpretation in future cases. In this study, segmental single hair analysis is compared with segmental hair lock analysis to document an ocfentanil-related death. The hair lock and single hair analyses were performed using the LC-MS/MS method after decontamination and incubation. Ocfentanil (OcF) concentrations ranged from 42 to 150 pg/mg in the segmented hair lock, depending on the segments. The hair lock and single hair analyses showed similar results: the highest concentrations were measured in the first two centimeters and decreased from root to tip. The similar profiles obtained from both the lock of hair and the single hair demonstrate the relevance of single hair analysis in cases where very few data are available. This article describes OcF concentrations in an authentic hair sample after a documented intake of this molecule in a fatality.

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Tamponade due to rupture of the chambers of the heart, in particular the left ventricle, after blunt thoracic trauma is described only sparsely in the literature. Most cases involve multiple thoracic trauma following motor vehicle accidents. We present the case study of a five-year old victim of a household accident, in which two concrete basins apparently fell on him. He died rapidly, despite attempted resuscitation. The autopsy showed essentially a hemorrhagic extravasation of the diaphragm and mediastinum, hemopericardium, and massive damage to the apex of the left ventricle. Pathological exam confirmed the traumatic origin of the cardiac rupture, with no underlying pathology. We will discuss the mechanisms described in the literature that result in such lesions, the mechanism which we believe most probable in this case, and the importance of background information. In our case study, lack of specific information concerning the accident prevents a definitive conclusion of the exact mechanism that caused this massive trauma particularly due to the fact that the external examination couldn't find any lesion in favor of a thoracic or abdominal traumatism. To our knowledge, in context of a household accident, such an isolated lesion causing almost immediate death has not previously been described in the literature.

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AIMS: This study aimed at comparing the cerebral cytotoxicity of ethanol and its main metabolite acetaldehyde after acute or chronic exposures of rat astrocytes in primary culture. METHODS: Cytotoxicity was evaluated on the cell reduction of viability (MTT reduction test) and on the characterization of DNA damage by single cell gel electrophoresis (or comet assay). RESULTS: Changes in astrocyte survival and in DNA integrity only occurred when the astrocytes were chronically exposed to ethanol (20 mM; 3, 6 or 9 days). On the other hand, viability and DNA integrity were deeply affected by acute exposure to acetaldehyde. Both effects were dependent on the concentration of acetaldehyde. The cytotoxic effect of acetaldehyde was also indirectly evaluated after modifications of the normal ethanol metabolism by the use of different inducers or inhibitors. In presence of ethanol, the concomitant induction of catalase (i.e. by glucose oxidase) and inhibition of aldehyde dehydrogenase (i.e. by methylene blue) led to acetaldehyde accumulation within cells. It was followed by both a reduction in viability and a substantial increase in DNA strand breaks. CONCLUSIONS: These data were thus consistent with a possible predominant role of acetaldehyde during brain ethanol metabolism. On the other hand, the effects observed after AMT could also suggest a possible direct ethanol effect and a role for free radical attacks. These data were thus consistent with a possible predominant role of acetaldehyde during brain ethanol metabolism. On the other hand, the effects observed after AMT could also suggest a possible direct ethanol effect and a role for free radical attacks.

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The central nervous system is vulnerable to oxidative stress, especially when a toxicant can modify the physiological balance between anti- and pro-oxidant mechanisms. Among brain cells, astrocytes seem less vulnerable than neurons, but their impairment can dramatically affect neurons because of their protective role toward neurons. Ethanol is able to stimulate the formation of reactive oxygen species and modify the activity of most of the antioxidant agents. However, ethanol can react with the OH* radical to form the alpha-hydroxyethyl radical, which is considered to be less toxic. Ethanol also can stimulate H2O2 degradation through catalase activation. This study, therefore, sought to determine whether ethanol affected the sensitivity of astrocytes exposed to various free radical-generating systems. The cellular impact of such exposure was assessed by assays exploring cytotoxicity (i.e., NR (neutral red) and MMT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetiazolium bromide) reduction assays) and genotoxicity (comet assay) induced by these treatments. DNA alterations were evaluated by single-cell gel electrophoresis (comet assay), considered a precocious biomarker of intracellular alterations. After concomitant exposure to H2O2 and ethanol, the viability of astrocytes decreased significantly whereas the mean percentage of DNA in the tail increased,reflecting DNA damage (H2O2 was either directly added to the culture medium or endogenously produced from menadione). Ethanol also reduced the loss of viability and DNA alterations after exposure to OH* radicals produced by a Fenton system. The exposure to a xanthine/xanthine oxidase system had the same effect.

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Ethanol consumption has long been associated with brain damage. However, the mechanism underlying this deleterious effect remains unclear. Among different hypotheses, acetaldehyde is regarded by certain authors as playing a major role in the expression of ethanol toxicity, but there are still some uncertainties about the exact nature of its implication. We therefore tried to characterize the profile of the alterations of neuronal viability and DNA integrity obtained after either a direct exposure to ethanol or to acetaldehyde. Ethanol at concentrations within the range of blood alcohol levels in intoxicated humans (< or = 100 mmol/L) induced DNA alterations without any apparent effect on cell viability. Acetaldehyde (< or = 1000 micromol/L) can also induce DNA alterations but with a different profile of the DNA cellular alterations. The comparison between the distributions of the comet tail DNA indicated that ethanol induced strong breaks (tail DNA > or = 60 a.u.) generation whereas acetaldehyde rather induced lower breaks (20 < or = tail DNA < or = 50 a.u.) formation but affecting a greater number of neurones. Acetaldehyde had thus a different genotoxic potential which may suggest a different mode of action or a different cellular target. Furthermore, when a single 100 mmol/L ethanol exposure did not lead to any loss of cell viability, the addition of an inhibitor of aldehyde dehydrogenase was followed by a significant loss in viability. In contrast, the inhibition of catalase, which suppresses acetaldehyde synthesis, led to no reduced viability in the same exposure conditions. ROS also reduced viability, but this was observed only after both cytochrome P450 stimulation and catalase inhibition. These combined results could suggest that acetaldehyde may play a significant role in the expression of ethanol toxicity in brain.

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AIMS: Ethanol can create progressive neuropathological and functional alterations of neurones. However, the influence of exposure duration is still debated. It is difficult to specify the level of alcohol consumption leading to alcohol-induced brain damage. Moreover, the mechanism of toxicity is assumed to combine direct and metabolically induced effects, although numerous uncertainties remain. Finally, the genotoxic power of ethanol has not fully been investigated in the brain. In the experiment reported herein, primary cultures of neurones were exposed either chronically or acutely to doses of ethanol within the range of blood alcohol levels in intoxicated humans. The impact on the integrity of neurones was assessed by cytotoxicity tests and DNA alterations by single-cell gel electrophoresis (Comet assay) and flow cytometry. Chronic ethanol exposure, even at a low dose, was more harmful to neurones than acute exposure. Both significant reductions in cell viability and DNA alterations were observed in this condition. On the other hand, DNA repair capacities seemed to be preserved as long as the viability measured by specific tests was not affected. Instead, neurones entered a death cell process compatible with apoptosis.

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A sensitive and specific method using reversed-phase liquid chromatography coupled with electrospray ionization-mass spectrometry (LC-ESI-MS) has been developed for the quantitative determination of flunitrazepam (F) and its metabolites 7-aminoflunitrazepam (7-AF), N-desmethylflunitrazepam (N-DMF) and 3-hydroxyflunitrazepam (3-OHF) in biological fluids. After the addition of deuterium labelled standards of F,7-AF and N-DMF, the drugs were isolated from urine or plasma by automated solid-phase extraction, then chromatographed in an isocratic elution mode with a salt-free eluent. The quantification was performed using selected ion monitoring of protonated molecular ions (M+H(+)). Experiments were carried out to improve the extraction recovery (81-100%) and the sensitivity (limit of detection 0.025 ng/ml for F and 7-AF, 0.040 ng/ml for N-DMF and 0.200 ng/ml for 3-OHF). The method was applied to the determination of F and metabolites in drug addicts including withdrawal urine samples and in one date-rape plasma and urine sample.

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In a previous study, the production of acetaldehyde and free radicals derived from ethanol was characterized in astrocytes in primary culture. In the present study, the effects of chronic exposure on the production of both compounds as well as on the main antioxidant system were compared with those of an acute exposure. This was done to better understand the different ways the brain reacts to these modes of exposure. Under these conditions, both a time-dependent increase in the accumulation of acetaldehyde and a decreased formation of the alpha-hydroxyethyl radical were shown. This was associated with increased activities of catalase, superoxide dismutase (SOD), and glutathione peroxidase (GPX) and with decreased glutathione (GSH) content. These effects, which counteract reactive oxygen species (ROS) formation by stimulating the main enzymes of the antioxidant system, were also associated with the reduced amount of radicals derived from ethanol. This could be a beneficial effect, but this was counter-balanced by the increased rate of acetaldehyde accumulation, whose high toxicity is well known. All these effects underline the crucial role played by catalase which, on one hand converts hydrogen peroxide to water and, on the other hand, ethanol to acetaldehyde.

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A 49-year-old male chemical industry worker was admitted to intensive care with a 24-hour history of respiratory failure, vomiting, headache, stupor, arterial hypotension, and cyanosed face and limbs. He had acute haemolysis (3.9 g/L plasma haemoglobin concentration) and 30% methaemoglobinaemia. Whereas the search for alcohol, barbiturates and opiates was negative, benzodiazepines and tricyclic antidepressants were present. The patient was in fact being treated with fluvoxamine, amitryptiline, and alprazolam. As the clinical and biological signs suggested chlorate poisoning, chlorate was looked for by using an aniline color reaction. It was found in gastric content and urine. Treatment consisted in mechanical ventilation, vasoactive amines, methylene blue, plasma exchange, exchange transfusion, and haemodialysis. Despite this, the patient had several cardiac arrests and refractory metabolic acidosis. He died 12 h after his admission. Specific ion chromatography was used afterhand to assay the chlorate in various body fluids. The technique was based on a separation on an ion exchange Dionex AS 12A column coupled with conductivity detection. A quantitative estimation was carried out by using external calibration with a four-point calibration curve which was linear between 1 and 15 mg/L. The measured plasma levels of chlorate were 78 and 29 mg/L respectively before and after exchange transfusion. Gastric-lavage liquid contained 1300 mg/L of chlorate and urine 4300 mg/L. Ion chromatography, which is routinely used in environmental studies helped to confirm a massive oral intake of chlorate by measuring the corresponding blood and urine chlorate concentrations, data which had only rarely been reported previously.

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The nervous system is one of the main targets of ethanol toxicity. Astrocytes might play an important role in ethanol-induced brain toxicity, because their integrity is essential for the normal growth and functioning of neurons. On the other hand, acetaldehyde has been implicated as a mediator in some of the biochemical, pharmacological, and behavioral effects of ethanol. The present study aimed at demonstrating the ability of astrocytes in culture to produce acetaldehyde from ethanol. Significant metabolization of ethanol with production of acetaldehyde was demonstrated in the primary culture of astrocytes. This production was quite low, compared with that usually observed in hepatocytes, but was in the same range as that measured in whole brain homogenates and corresponded to biologically active levels. Such a demonstration could bring new elements for understanding of ethanol neurotoxicity.

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Ethanol evaporation from the culture medium is a potential source of misinterpretation of long-term exposure of cells. Different methods have been proposed to prevent this evaporation, the most effective being the saturation of the atmosphere over the culture medium with ethanol. Unfortunately, no simple predictive method has been devised to determine the appropriate concentration of ethanol in the system avoiding either evaporation or contamination of the culture medium. We present some keys to a solution adapted to the culture of astrocytes, which allow for the first time a direct evaluation of ethanol absorption by these cells. The system described remains compatible with normal growth and viability.

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Formation of the alpha-hydroxyethyl radical (CH3 degree CHOH) has already been extensively demonstrated after ethanol metabolism in the liver. Despite favourable conditions, this formation in the brain has remained speculative since there is no direct experimental evidence in intact brain cells. In this preliminary study, the formation of such a radical was observed after exposure of astrocytes and astrocytic C6 glioma cells to ethanol. These cells were studied because astrocyte integrity is essential for normal growth and functioning of neurons. The free radicals were detected by EPR spectroscopy using the spin trapping technique. Astrocytes appeared to be more sensitive than the C6 cells to free radical formation as the intensity of the signal was higher after exposure of the astrocytes and increased with time, a fact not observed after exposure of the C6 cells.

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A gas chromatographic technique with flame ionization detection, which is based on a solid-phase extraction (SPE) procedure using mixed-mode SPE columns, for the simultaneous quantitation of dextropropoxyphene and norpropoxyphene in urine is presented. Urine is treated with sodium hydroxide in order to rearrange, by base catalysis, norpropoxyphene to norpropoxyphene amide, which is then extracted with these columns and chromatographed. The method is specific, linear over the range 0-2000 ng/mL, sensitive, and reproducible. The extracts are cleaner than those obtained with traditional liquid-liquid extraction procedure, which is an important feature in view of further mass spectrometric confirmation of narcotics and other drugs.

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The nervous system is one of the main targets of ethanol toxicity and it has been suggested that astrocytes might play an important role as their integrity is essential for the normal growth and functioning of neurons. Morphological variations of astrocyte cultures were therefore examined after exposure to various doses of ethanol (0.5, 1 and 2%) for different durations (24, 48, 72 and 96 h). The percentage of cell viability and the cell density were calculated and the changes in astrocyte morphology were assessed by an image analysis system (Samba 2005) allowing the characterization of 5 parameters (perimeter, surface, elongation factor, convexity factor and the form factor) of a great number of cells (over 6500). This was necessary because of the high variability in normal cultured astrocyte morphology. A two-way statistical approach (2-factors ANOVA completed by stepwise discriminant analysis) was adopted to emphasize the differences between control and exposed cells. In such conditions, ethanol treated cells became more elongated, less circular and more concave and did not grow like non-exposed cells. The mean pooled values of these parameters tended to be modified as a function of the dose of ethanol. The relationships between parameters clearly separated the groups as a function of the different doses. Finally no significant difference was observed in cell viability and cell density despite lower scores in the groups exposed to the highest dose of ethanol for the longest time. Our results suggest that ethanol might affect astrocytes in two different but probably complementary ways by modifying the cell shape and by altering normal cell development.

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We report the two first cases of neuropsychic side-effects with zipeprol (non opiate antitussive) given at therapeutic doses. In both cases, the patients showed confusion, whereas, hallucinations occurred only in one case. The anticholinergic activity of zipeprol might explain these effects. These two cases are in agreement with zipeprol central nervous system action.

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