Analysis of γ-hydroxy butyrate by combining capillary electrophoresis-indirect detection and wall dynamic coating: application to dried matrices.

γ-Hydroxybutyric acid (GHB) is a powerful central nervous system depressant, currently used in medicine for the treatment of narcolepsy and alcohol dependence. In recent years, it has gained popularity among illegal club drugs, mainly because of its euphoric effects as well as doping agent and date rape drug. The purpose of the present work was the development of a rapid analytical method for the analysis of GHB in innovative biological matrices, namely dried blood spots (DBSs) and dried urine spots (DUSs). The analytical method is based on capillary zone electrophoresis with indirect UV absorption detection at 210 nm and capillary wall dynamic coating. The background electrolyte is composed of a phosphate buffer containing nicotinic acid (probe for detection) and cetyltrimethylammonium bromide (CTAB, reversal of electroosmosis in wall dynamic coating). The influence of probe and CTAB concentration, together with buffer pH, on migration time and signal response was investigated. Under the optimized conditions, analytical linearity and precision were satisfactory; absolute recovery values were also high (>90 %); the use of dried matrices (DBSs and DUSs) was advantageous as an alternative matrix to classical ones. No interferences were found either from the most common exogenous or from endogenous compounds. This analytical approach can offer a rapid, precise and accurate method for GHB determination in innovative biological samples, which could be important for screening purposes in clinical and forensic toxicology. Graphical Abstract CE method, by combined indirect UV detection and dynamic coating, for GHB determination in DBSs and DUSs.

Fatal Cervical Spine Injury From Diving Accident.

Spinal cord injuries result after diving into shallow water, often after incautious jumps head first into water of unknown depth during recreational or sport activities. Mortality is generally due to upper cervical trauma. The authors present a case of a diving-related death in a young woman who underwent medicolegal investigations. The measured water depth at the supposed dive site was 1.40 m. Postmortem radiology and autopsy revealed fractures of the body and the posterior arch of the fifth cervical vertebra, a fracture of the right transverse process of the sixth cervical vertebra and hemorrhages involving the cervical paraspinal muscles. Neuropathology showed a posterior epidural hematoma involving the whole cervical region and a symmetric laceration of the spinal cord located at the fourth and fifth cervical vertebra level, surrounded by multiple petechial hemorrhages. Toxicology revealed the presence of ethanol in both blood and urine samples. The death was attributed to cervical spine fracture (C5-C6), spinal cord contusion, and subsequent drowning. This case highlights the usefulness of postmortem radiology, examination of the deep structures of the neck, toxicology, neuropathology, and a detailed research of signs of drowning to formulate appropriate hypotheses pertaining to the cause and mechanism of death.

PKCε contributes to chronic ethanol-induced steatosis in mice but not inflammation and necrosis.

BACKGROUND: Protein kinase C epsilon (PKCε) has been shown to play a role in experimental steatosis by acute alcohol. The "two-hit" hypothesis implies that preventing steatosis should blunt more advanced liver damage (e.g., inflammation and necrosis). However, the role of PKCε in these pathologies is not yet known. The goal of this current work was to address this question in a model of chronic alcohol exposure using antisense oligonucleotides (ASO) against PKCε. METHODS: Accordingly, PKCε ASO- and saline-treated mice were fed high-fat control or ethanol (EtOH)-containing enteral diets for 4 weeks. RESULTS: Chronic EtOH exposure significantly elevated hepatic lipid pools as well as activated PKCε. The PKCε ASO partially blunted the increases in hepatic lipids caused by EtOH. Administration of PKCε ASO also completely prevented the increase in the expression of fatty acid synthase, and tumor necrosis factor α caused by EtOH. Despite these protective effects, the PKCε ASO was unable to prevent the increases in inflammation and necrosis caused by chronic EtOH. These latter results correlated with an inability of the PKCε ASO to blunt the up-regulation of plasminogen activator inhibitor-1 (PAI-1) and the accumulation of fibrin. Importantly, PAI-1 has been previously shown to more robustly mediate inflammation and necrosis (vs. steatosis) after chronic EtOH exposure. CONCLUSIONS: This study identifies a novel potential mechanism where EtOH, independent of steatosis, can contribute to liver damage. These results also suggest that PAI-1 and fibrin accumulation may be at the center of this PKCε-independent pathway.

Assistance of ethyl glucuronide and ethyl sulfate in the interpretation of postmortem ethanol findings.

Postmortem ethanol formation is a well-known problem in forensic toxicology. The aim of this study was to interpret findings of ethanol in blood, in a large collection of forensic autopsy cases, by use of the nonoxidative ethanol metabolites, ethyl glucuronide (EtG), and ethyl sulfate (EtS). In this study, according to previously published literature, antemortem ethanol ingestion was excluded in EtS-negative cases. Among 493 ethanol-positive forensic autopsy cases, collected during the study period, EtS was not detected in 60 (12 %) of the cases. Among cases with a blood alcohol concentration (BAC) of ≤ 0.54 g/kg, antemortem ethanol ingestion was excluded in 38 % of the cases, while among cases with a BAC of ≥ 0.55 g/kg, antemortem ethanol ingestion was excluded in 2.2 % of the cases. For all cases where ethanol was measured at a concentration >1.0 g/kg, EtS was detected. The highest blood ethanol concentration in which EtS was not detected was 1.0 g/kg. The median concentrations of EtG and EtS in blood were 9.5 µmol/L (range: not detected (n.d.) 618.1) and 9.2 µmol/L (range: n.d. 182.5), respectively. There was a statistically significant positive correlation between concentration levels of ethanol and of EtG (Spearman's rho=0.671, p<0.001) and EtS (Spearman's rho=0.670, p<0.001), respectively. In conclusion, this study showed that in a large number of ethanol-positive forensic autopsy cases, ethanol was not ingested before the time of death, particularly among cases where ethanol was present in lower blood concentrations. Routine measurement of EtG and EtS should therefore be recommended, especially in cases with BAC below 1 g/kg.

Disposition and metabolism of cumene in F344 rats and B6C3F1 mice.

Cumene is a high-production volume chemical that has been shown to be a central nervous system depressant and has been implicated as a long-term exposure carcinogen in experimental animals. The absorption, distribution, metabolism, and excretion of [(14)C]cumene (isopropylbenzene) was studied in male rats and mice of both sexes after oral or intravenous administration. In both species and sexes, urine accounted for the majority of the excretion (typically ≥ 70%) by oral and intravenous administration. Enterohepatic circulation of cumene and/or its metabolites was indicated because 37% of the total dose was excreted in bile in bile duct-cannulated rats with little excreted in normal rats. The highest tissue (14)C levels in rats were observed in adipose tissue, liver, and kidney with no accumulation observed after repeat dosing up to 7 days. In contrast, mice contained the highest concentrations of (14)C at 24 h after dosing in the liver, kidney, and lung, with repeat dosing accumulation of (14)C observed in these tissues as well as in the blood, brain, heart, muscle, and spleen. The metabolites in the expired air, urine, bile, and microsomes were characterized with 16 metabolites identified. The volatile organics in the expired air comprised mainly cumene and up to 4% α-methylstyrene. The major urinary and biliary metabolite was 2-phenyl-2-propanol glucuronide, which corresponded with the main microsomal metabolite being 2-phenyl-2-propanol.

An enzymatic method to determine γ-hydroxybutyric acid in serum and urine.

BACKGROUND: Gamma-hydroxybutyric acid (GHB) has become one of the most dangerous illicit drugs of abuse today. It is used as a recreational and date rape drug because of its depressant effect on the central nervous system, which may cause euphoria, amnesia, respiratory arrest, and coma. There is an urgent need for a simple, easy-to-use assay for GHB determination in urine and blood. In this article, a rapid enzymatic assay adapted to clinical chemistry analyzers for the detection of GHB is presented. METHODS: The described GHB enzymatic assay is based on a recombinant GHB dehydrogenase. The full validation of the assay was performed on a Konelab 30 analyzer (Thermo Fisher Scientific). RESULTS: The analytical sensitivity was <1.5 mg/L, whereas the functional sensitivity was 4.5 mg/L in serum and 2.8 mg/L in urine. The total imprecision coefficient of variation (CV) was <9.8% in serum and <7.9% in urine. The within-run imprecision showed a CV of <3.8% in serum and <4.6% in urine. The assay was linear within the range 5-250 mg/L. Mean recoveries were 109% in serum and 105% in urine. No cross-reactivity was observed for tested GHB analogues and precursors. Comparison of GHB-positive samples showed an excellent correlation with ion chromatography, gas chromatography-mass spectrometry, and liquid chromatography associated to tandem mass spectrometry. Except for ethanol, no substantial interference from serum constituents and some drugs was observed. CONCLUSIONS: This automated GHB assay is fully quantitative and allows the accurate measurement of GHB in serum and urine. It can be used as a rapid screening assay for the determination of GHB in intoxicated or overdosed patients.

Recent heavy alcohol consumption at death certified as ischaemic heart disease: correcting mortality data from Kaunas (Lithuania).

AIMS: To assess the proportion of deaths assigned to ischaemic heart disease (IHD) which in fact were caused by the toxic effects of alcohol, and how this may affect the official statistics of mortality from IHD in Lithuania. METHODS: Using the IHD register in Kaunas, Lithuania, and verifying underlying causes of death using standard international methodology, 3061 cases were found in Kaunas city who had died from IHD at age 25-64 during 1993-2007. Out-of-hospital sudden deaths accounted for 2467 cases (81%), including 1498 where forensic autopsy was conducted and post-mortem concentration of alcohol in blood and urine was available. RESULTS: In total, 78.4% of all initial IHD diagnoses were verified, while in 8.7% of deaths the underlying cause of death was corrected into an alcohol-related cause and in 12.9% to other diseases. Alcohol was found in about half (50.3%) of out-of-hospital death cases subjected to autopsy. In 18.0% of cases, the alcohol concentration was 3.5% or higher. Alcohol was more likely to be present in winter months and at weekends. CONCLUSION: A significant number of alcohol-attributable deaths in Lithuania were misclassified as coronary deaths, accounting for almost one-tenth of officially registered deaths from IHD in ages 25-64. A high prevalence of positive post-mortem blood or urine alcohol tests suggests that the proportion of alcohol-related deaths among out-of-hospital IHD deaths may be actually even higher. A similar situation may be present in some other countries where high levels of alcohol consumption and binge drinking patterns are observed.

Urinary ethyl glucuronide and ethyl sulfate testing for recent drinking in alcohol-dependent outpatients treated with acamprosate or placebo.

AIMS: Ethyl glucuronide (EtG) and ethyl sulfate (EtS) are sensitive and specific biomarkers for recent alcohol ingestion. This study compared urinary EtG and EtS measurement with self-reports for detection of prior drinking in alcohol-dependent outpatients treated with the anti-craving medication acamprosate or placebo. METHODS: Alcohol-dependent outpatients (26 women, 30 men) were randomized to 21 days of oral acamprosate (2 g/day) or placebo treatment in a double-blind design. They were instructed to refrain from drinking during the study. Return visits to the ward for blood and urine sampling and filling out questionnaires were made on Day 7, 14 (urine sample optional) and 22 (urine sample mandatory). EtG and EtS were determined by liquid chromatography-mass spectrometry. RESULTS: On the first day (Day 0), 72% of all patients (acamprosate 65%, placebo 78%) tested positive for recent drinking according to urinary EtG (reporting limit ≥ 0.5 mg/l) and EtS (≥ 0.1 mg/l). On the final day (Day 22), the frequency of positive tests was significantly reduced to 30% in the acamprosate group (P = 0.0374) and 33% for placebo (P = 0.0050). However, there was no difference between the treatment groups. When both groups were combined, the EtG (P = 0.025) and EtS (P = 0.015) concentrations were lower on the final day. Altogether, EtG and EtS were detected in 76 of 156 (49%) urine samples. When drinking in the day before sampling was admitted, 93% of urines tested positive; when drinking was denied, still 28% of the samples were positive. CONCLUSION: These results confirmed the value of urinary EtG and EtS as reliable indicators of recent drinking during outpatient treatment of persons with alcohol-related problems, and as objective outcome measures when evaluating new alcohol treatment strategies and pharmacotherapies.

Double suicide.

Most suicides are solitary and private; but few result from a pact between 2 people to die together. Two young men made a pact to commit suicide by hanging from a tree. It was the first case among 1320 single suicides in more than 2 decades in eastern Croatia. Double suicide between people of the same gander is an unusual event. It is predominantly made by male-female partners and by less violent methods. One of the 2 members experienced depression. He may have been the initiator and the other partner may have been the dependent. The initiator usually plans the act and stimulates the other party. Whether the decision was evenly shared by both partners and initiative came from one of the two or it was result of 2 independent decisions remains open for analysis. Double suicide is also very interesting from the perspective of medical examiners. They need to make comprehensive postmortem examination to finally conclude the proper cause and the manner of death.

Endogenous melatonin is not obligatory for the regulation of the rat sleep-wake cycle.

STUDY OBJECTIVES: Though melatonin and melatonin receptor agonists are in clinical use and under development for treating insomnia, the role of endogenous melatonin in the regulation of the sleep-wake cycle remains uncertain. Some clinical case reports suggest that reduced nocturnal melatonin secretion is linked to sleep disruption, but pineal-gland removal in experimental animals has given variable results. DESIGN: The present study examined the effects of pinealectomy on the diurnal sleep-wake cycle of rats implanted with a radiotransmitter to allow continuous measurement of cortical electroencephalogram, electromyogram, and core temperature (Tc) without restraint in their home cages. MEASUREMENTS AND RESULTS: Tc was slightly (0.2 degrees C) but significantly lower after pineal removal. The total amount and diurnal distribution of locomotor activity, wake, non-rapid eye movement (NREM) sleep, and rapid eye movement (REM) sleep were unaltered in pinealectomized rats compared to sham-operated controls. Sleep consolidation measured by determining wake, NREM sleep, and REM sleep bout length and frequency was also unchanged. The EEG power spectrum during NREM sleep was unchanged, but a significant decrease in theta power (5-8 Hz) during REM sleep episodes was found. CONCLUSIONS: Our data provide no evidence that endogenous circulating melatonin plays a role in regulating the sleep-wake cycle in rats. However, because cortical theta oscillations are generated in the CA1-3 layer of the hippocampus, neurons known to express melatonin receptors, this suggests that a lack of melatonin following pineal removal influences the function of these neurons and is consistent with previous work suggesting that endogenous melatonin is an important regulator of hippocampal physiology.

Forensic toxicology findings in deaths involving gamma-hydroxybutyrate.

Concentrations of the illicit drug gamma-hydroxybutyrate (GHB) were determined in femoral venous blood and urine obtained at autopsy in a series of GHB-related deaths (N = 49). The analysis of GHB was done by gas chromatography after conversion to gamma-butyrolactone and quantitation of the latter with a flame ionization detector. The cutoff concentration of GHB in femoral blood or urine for reporting positive results was 30 mg/L. The deceased were mainly young men (86%) aged 26.5 +/- 7.2 years (mean +/- SD), and the women (14%) were about 5 years younger at 21.4 +/- 5.0 years. The mean, median, and highest concentrations of GHB in femoral blood (N = 37) were 294, 190, and 2,200 mg/L, respectively. The mean urine-to-blood ratio of GHB was 8.8, and the median was 5.2 (N = 28). In 12 cases, the concentrations of GHB in blood were negative (<30 mg/L) when the urine contained 350 mg/L on average (range 31-1,100 mg/L). Considerable poly-drug use was evident in these GHB-related deaths: ethanol (18 cases), amphetamine (12 cases), and various prescription medications (benzodizepines, opiates, and antidepressants) in other cases. Interpreting the concentrations of GHB in postmortem blood is complicated because of concomitant use of other psychoactive substances, variable degree of tolerance to centrally acting drugs, and the lack of reliable information about survival time after use of the drug.

Analysis of clinical forensic examination reports on sexual assault.

Medical-forensic examination of sexual assault victims and alleged offenders is a common task of many forensic institutes. In the current study, the results from samples taken at the Institute of Legal Medicine, Hanover Medical School, during a period from 2005 to 2007 were retrospectively evaluated. In total, 292 victims (283 females and nine males) and 88 suspects were examined. At the time of the assault, 41.8% of the victims and 43.2% of the alleged perpetrators were under the influence of alcohol. Injuries were found in 84.9% of the victims and 39.8% of the suspects. Thirty victims (10.3%) reported having been choked or strangled. Cytology was performed in 218 victims. In 81 cases (38.0%), sperm could be detected in vaginal swabs up to 3 days post-assault. In seven (18.9%) out of 37 anal samples, evidence of sperm could be found 24 h post-assault. None of 22 oral samples was positive for sperm. Out of 301 sexual assault cases, 171 could be proved by means of medical-forensic examination. In summary, our evaluation shows that an early medical-forensic examination of both victim and suspect can secure numerous medical findings. Furthermore, persons intoxicated by alcohol, handicapped persons and persons with psychiatric disorders are more vulnerable to become a sexual assault victim.

An avoidable death in police custody?

An obviously inebriated 36-year-old man was found in a helpless condition on a pavement. When he was brought to a police station, a doctor certified the individual to be fit to be held in custody. He was unresponsive when the officers tried to wake him the next morning but was allowed to sleep on until he was found dead in his cell at mid day. An autopsy revealed as cause of death an epidural hematoma and cerebral contusion with fracture of the calvarium. The expert report stressed the point that the monitoring of the state of consciousness had been objectively insufficient throughout the entire time period, both in terms of frequency as well as in terms of the monitoring methods. However, neither the police officers involved nor the doctor, was subjected to prosecution mainly because of the high burden of proof required by German criminal law. The article will discuss conclusions for the avoidance of such deaths (such as training of police officers, legal regulations on monitoring procedures, etc).

The role of ethanol in fatalities in Jeddah, Saudi Arabia.

In Saudi Arabia, alcohol consumption is prohibited by law, but interpreting postmortem ethanol can be complicated by its postmortem production. This study developed and validated a method using headspace gas chromatography with flame ionization detection and liquid chromatography tandem mass spectroscopy to detect ethanol and its polar metabolites (ethyl glucuronide [EtG] and ethyl sulfate [EtS]) in postmortem blood and urine specimens, respectively. All calibration curves were linear with coefficients of determination greater than 0.999. The limits of detection ranged 4.5-5.0mg/dL for ethanol and 0.05-0.06mg/L for EtG and EtS. The limits of quantification were 10.0mg/dL for ethanol and 0.075mg/L for EtG and EtS. Within-run precision was less than 11% for all analytes of interest. Matrix effects for EtG and EtS ranged 3-47%. After excluding matrix effects, analytical recoveries ranged 72-100%. This validated method was then used for routine postmortem forensic toxicology analyses in 592 routine postmortem cases to distinguish between antemortem ethanol consumption and its postmortem microbial formation. Among them, 98 blood samples (17%) were positive for ethanol or its polar metabolites. Thirty-two of these cases (33%) were positive for EtG and EtS and therefore due to antemortem ethanol consumption. The remaining 66 (67%) cases were negative for both EtG and EtS and therefore due to postmortem ethanol synthesis. Because this is the first study to report the problem of alcohol consumption in Saudi Arabia, further studies are essential for validating these findings.

Review of the pattern of traumatic limb lesions sustained in cases of hanging.

This retrospective study sought to identify a regular pattern of limb bruising which occurs in association with suicidal or accidental hanging. Following exclusion of cases suspicious for homicide, 82 consecutive cases of hanging from a 10-year period were retrospectively reviewed to identify the pattern of traumatic limb injury in each case. Relevant information such as location, toxicology, and type of suspension was also noted. 72% of the reviewed cases had traumatic limb lesions, the majority of which occurred on the posterior upper limb and the anterior lower limb. Although the distribution of limb injury in our study mirrored that found in the literature, the incidence is much higher than in previous studies (7.4-20%). This could either be due to differences in confounding factors such as intoxication and location of hanging or differences in the practice of recording of limb trauma in hanging between centres. Neither type of suspension nor location of hanging were significantly associated with an increased incidence of traumatic limb injury. Positive toxicology was found to increase the likelihood of sustaining limb injury (p = .044084). In conclusion, the presence of this well documented pattern of traumatic limb lesions in cases of hanging should not always raise suspicion of foul play.

Evaluating the hip-flask defence using analytical data from ethanol and ethyl glucuronide. A comparison of two models.

AIM: Claimed intake of alcohol after a traffic incident, called the hip-flask defence, can be objectively assessed by different methods. One of them is the use of two consecutive ethanol concentrations in urine and the ratio between ethanol concentrations in urine and blood. Another one is the concentrations of ethyl glucuronide (EtG) and ethyl sulphate (EtS) in blood and their ratio to ethanol. The experimental basis for both these models is from single dose studies only. The aim of this study was therefore to describe the kinetics of ethanol, EtG and EtS after ingestion of two repeated doses of ethanol and to investigate the usefulness of the different models for the assessment of the hip-flask defence. METHODS: Thirty-five subjects ingested a first dose of 0.51 g of ethanol per kilo body weight, and two hours later a second dose (the hip-flask drink) of 0.25, 0.51 or 0.85 g of ethanol per kilo body weight. Ten urine and 17 blood samples were collected and analysed for ethanol, EtG and EtS using fully validated methods. It was investigated if all subjects fulfilled the criteria for recent drinking, according to the two different models, when using the samples collected 180-240 minutes after start of first dose drinking. According to the first model, increase in urinary ethanol concentrations and a ratio UAC/BAC below 1.3 indicated recent drinking. According to the second model, increase in blood EtG concentrations and a ratio ethanol (g/kg)/EtG (mg/L) above 1 indicated recent drinking. RESULTS: All subjects in the high dose group fulfilled all criteria for recent drinking. One subject in the medium dose group and nine subjects in the low dose group failed to show increasing UAC and/or a UAC/BAC ratio below 1.3. One subject in the low dose group failed to show increasing concentrations of blood EtG, but all subjects showed a ratio ethanol/EtG above 1. CONCLUSIONS: The present study showed, by the use of experimental data, that both two models used to investigate the hip-flask defence can be used, but only when the hip-flask dose is sufficiently high.

Parasailing fatalities in southwest Florida.

Parasailing is a recreational sport that is generally considered to be of little risk to the participants. Typically, the passenger launches from a motorboat with a specially designed winch that pulls him or her back to the boat at the end of the ride. The sport is not regulated at the federal, state, or county level. There have been few reports of injuries to parasailors. Additionally, there have been only 2 fatalities reported to the United States Coast Guard in a 10-year review. We report the details of these 2 deaths, those of a mother and daughter riding in a tandem parasail, which occurred on Fort Myers Beach in 2001, as well as an additional case of a parasailing fatality that occurred in southwest Florida in 1999. These cases illustrate the injuries seen in such fatalities and the hazards posed by adverse weather conditions and faulty equipment, as well as the impairment of passenger judgment by drugs and/or alcohol.

The prevalence of drugs and alcohol found in road traffic fatalities: a comparative study of victims.

Researchers have studied the involvement of drugs and alcohol in fatal road traffic incidents, but with particular emphasis on the possible impairment of the driver. This paper describes a comparative study of drug and alcohol findings in various victim groups (drivers of cars, vans or lorries, car passengers, motorcyclists, motorcycle passengers, cyclists and pedestrians) between 2000 and 2006. Post-mortem blood and urine specimens submitted were analysed by immunoassay, GC-NPD, GC-FID, GC-MS and HPLC-DAD. The results of 1047 cases indicated 54% of all victims were positive for drugs and/or alcohol, with the highest percentage of positive findings occurring in pedestrians (63%). Males between the ages of 17-24 were most likely to be involved in a road traffic accident, whether being in control of a vehicle (driver) or involved indirectly (car passenger, pedestrian, motorcycle passenger). A wide range of drugs were detected (e.g., drugs of abuse, anti-convulsants, anti-histamines, anti-inflammatories, anti-psychotics, cardiac drugs and over-the-counter products), but alcohol and cannabinoids were the most frequent substances across the victim groups. When detected, alcohol was commonly above the legal driving limit in blood and urine (>63% in those in control and >60% not in control). Overall, the presence of drugs and/or alcohol was of similar frequency in those victims in control (55% of driver, 48% of motorcyclists, 33% of cyclists) and not in control of a vehicle (52% of car passengers, 63% of pedestrians). This degree of frequency strongly implicates the involvement of drugs and alcohol in road traffic incidents and infers an effect on driving ability and individual impairment.

Model cannons causing lethal traumata.

Model cannons are usually considered to be harmless toys; therefore no legal regulations are required. Nevertheless, suicidal intentions or playful experimentation can turn this toy into a lethal weapon. This paper describes two cases of misuse of model cannons, leading to severe injuries and consequently death: the first case was the result of a suicidal intention, where the cannon caused a bullet injury into the forehead with marked signs of close-firing and intensive brain damage. The second case was an accidental event, where the cannon induced an entry wound at the left chest wall with surrounding carbonization and a fatal rupture of the heart. Circumstances of the lethal situations and mechanisms of the cannons leading to fatal injuries including ballistic considerations are described. The report is supported by a thorough literature research.

Evaluating the hip-flask defence in subjects with alcohol on board: An experimental study.

Driving under the influence of alcohol is a major problem for traffic-safety and a popular defence argument is alleged consumption after driving, commonly referred to as the hip-flask defence. Forensic toxicologists are often called as expert witnesses in drinking and driving cases where the suspect has claimed the hip-flask defence, to assess the credibility of the explanation. Several approaches to help the expert have been introduced but the scientific data used to support or challenge this is solely based on data from controlled single doses of ethanol administered during a short time and in abstinent subjects. In reality, we believe that even in drinking after driving cases, the subject many times has alcohol on board at time of the hip-flask drink. This questions the applicability of the data used as basis to investigate the hip-flask defence. To fill this knowledge gap, we aimed to investigate how blood and urine ethanol kinetics vary after an initial drinking session of beer and then a subsequent hip-flask drink of three different doses of whiskey. Fifteen subjects participated in the study and each provided 10 urine samples and 17 blood samples over 7h. The initial drink was 0.51g ethanol/kg and the second was either 0.25, 0.51, or 0.85g/kg. Our data suggested that the difference between the ethanol concentrations in two consecutive urine samples is a more sensitive parameter than the ratio between urine and blood alcohol to detect a recent intake when ethanol from previous intakes are already present in the body. Twelve subjects presented results that fully supported a recent intake using the criteria developed from a single intake of ethanol. Three subjects showed unexpected results that did not fully support a recent intake. We conclude that data from one blood sample and two urine samples provide good evidence for investigating the hip-flask defence even if alcohol was on board at the time of the hip-flask drink.