Interpretation of a highly positive ethyl glucuronide result together with negative fatty acid ethyl esters result in hair and negative blood results

Considering the widespread nature of alcohol-related problems, the diagnosis of excessive alcohol consumption is an important task from medical and legal viewpoints. Alcohol abuse can be documented by usual blood [e.g., carbohydrate deficient transferrin (CDT)] and liver function [e.g., γ-GT or mean corpuscular volume (MCV)] tests, and, over a long-term basis, by hair analysis. Major markers of ethanol consumption in hair are ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEEs). Detection of EtG in hair is said to be associated with excessive alcohol consumption, whereas a negative result does not unambiguously exclude alcohol abuse. Investigations on FAEEs can also be used to monitor excessive alcohol consumption. Four FAEEs (ethyl myristate, ethyl palmitate, ethyl oleate, and ethyl stearate) are the most suitable markers for the detection of heavy alcohol consumption and show different concentrations in hair of children, adult teetotalers, and social drinkers in comparison with FAEEs concentrations found in the hair of alcoholics. The Society of Hair Testing has provided guidelines for hair testing for chronic excessive alcohol consumption, and states positive cutoffs for a 0–3 cm hair segment as 30 pg/mg and 0.5 ng/mg for EtG and FAEEs, respectively. This study reviews the difficulties in interpreting blood and hair results of a 41-year-old woman involved in a child custody legal dispute. Her EtG and FAEEs concentrations in a 0–3 cm segment were 203 pg/mg and 0.29 ng/mg, respectively, and blood parameters were in the normal range (0.6 %, 93 fl, 14 IU/l for CDT, MCV, and γ-GT, respectively). Although the high EtG concentration suggested excessive drinking behavior, the second hair marker, FAEEs, and the three bloods tests were inconspicuous and in accordance with the claim of abstinence from alcohol by the subject. The woman declared having dyed her hair more than 6 months prior to sampling, use of weight-loss medication, and consumption of about four energy drinks per day. A hair lotion based on ethanolic plant extracts was regularly used by the subject, and could have been a source of contamination. Therefore, we recommend that possible external sources of hair contamination always be taken into consideration, especially if contradictory biological results are obtained and if the subject denies any alcohol intake.     

Fatty acid ethyl esters in hair as alcohol markers: estimating a reliable cut-off point by evaluation of 1,057 autopsy cases

Alcohol abuse is a widespread problem, especially in Western countries. Therefore, it is important to have markers of alcohol consumption with validated cut-off points. For many years research has focused on analysis of hair for alcohol markers, but data on the performance and reliability of cut-off values are still lacking. Evaluating 1,057 cases from 2005 to 2011, included a large sample group for the estimation of an applicable cut-off value when compared to earlier studies on fatty acid ethyl esters (FAEEs) in hair. The FAEEs concentrations in hair, police investigation reports, medical history, and the macroscopic and microscopic alcohol-typical results from autopsy, such as liver, pancreas, and cardiac findings, were taken into account in this study. In 80.2 % of all 1,057 cases pathologic findings that may be related to alcohol abuse were reported. The cases were divided into social drinkers (n = 168), alcohol abusers (n = 502), and cases without information on alcohol use. The median FAEEs concentration in the group of social drinkers was 0.302 ng/mg (range 0.008–14.3 ng/mg). In the group of alcohol abusers a median of 1.346 ng/mg (range 0.010–83.7 ng/mg) was found. Before June 2009 the hair FAEEs test was routinely applied to a proximal hair segment of 0–6 cm, changing to a routinely investigated hair length of 3 cm after 2009, as proposed by the Society of Hair Testing (SoHT). The method showed significant differences between the groups of social drinkers and alcoholics, leading to an improvement in the postmortem detection of alcohol abuse. Nevertheless, the performance of the method was rather poor, with an area under the curve calculated from receiver operating characteristic (ROC curve AUC) of 0.745. The optimum cut-off value for differentiation between social and chronic excessive drinking calculated for hair FAEEs was 1.08 ng/mg, with a sensitivity of 56 % and a specificity of 80 %. In relation to the “Consensus on Alcohol Markers 2012” by the SoHT, an increase in the cut-off value for FAEEs in the proximal hair segment 0–3 cm from 0.5 to 1 ng/mg may be advisable to avoid excessive numbers of false positive results.     

Mumie ohne Geheimnisse – Identifizierung, Eingrenzung der Liegezeit und Nachweis von Morphin

In 2010 the mummified corpse of a 25-year-old male drug user was found in his apartment after a 2-year postmortem interval and had died just after being released from an 11-month prison sentence. The initial postmortem examination revealed skeletization and advanced stage mummification. The autopsy revealed surprisingly well preserved thigh muscles in addition to disintegration of the parenchymatous organs and nearly complete desiccation. The comprehensive toxicological investigations provided evidence of morphine (0.41 ng/mg in hair, 0.05 µg/g in muscle, 1.95 µg/g in liver, 0.54 µg/g in kidneys, 70 ng/g in bone and 7.57 ng/g in bone marrow), 6-acetylmorphine (0.15 ng/mg in hair) and codeine (0.05 ng/mg in hair and 0.01 µg/g in muscle) due to heroin consumption. The concentrations measured in the postmortem specimens indicated lethal heroin intoxication and the distribution ratio of the concentrations contributed to clarifying the cause of death. The state of preservation of the body, the scope of investigations and the results make the case just as newsworthy as the rarity of comparable cases.     

Specific interpretation of hair concentrations in 2 fatal metformin intoxication cases

Hair analysis is very useful for toxicological investigations since, by providing a wider detection window, it gives the possibility to perform a retrospective study on the historical consumption of a substance. Unfortunately, there are no data available for hair concentrations in metformin-related deaths. In this study, the authors present 2 cases of fatal metformin intoxication in which, for the first time, hair analysis was performed using a specific GC–MS/MS method. Metformin was tested positive in femoral blood (112.3 mg/L and 64.7 mg/L respectively) and cardiac blood (226.9 and 203.2 mg/L) of the two subjects. For case 1, other samples were also tested positive, including vitreous humor (31.1 mg/L) and gastric contents (773.5 mg/L). In case 2, metformin was measured at 844.9 mg/L in urine. Metformin hair concentrations were 28.3–44.8 and 22.5 ng/mg for both cases, respectively. The concentrations found in the 2 fatal cases are clearly higher than those obtained in a previous study with subjects under treatment (0.3–3.8 ng/mg) or those found in 3 post-mortem cases where metformin death was excluded (0.6–1.4 ng/mg). Excessive sweating during the agonal phase due to fatal hypoglycemia could explain these elevated concentrations as sweat can have contaminated the hair.     

A review of the pharmacology, pharmacokinetics and behavioral effects of procaine in thoroughbred horses.

Since procaine has both local anaesthetic and central stimulant actions its presence in the blood or urine of racing horses is forbidden. After rapid intravenous injection of procaine HC1 (2.5 mg/Kg) in thoroughbred mares plasma levels of this drug fell rapidly (t 1/2 alpha = 5 min) and then more slowly (t 1/2 beta = 50.2 min). These kinetics were well fitted by a two compartment open model (Model I). This model gave an apparent Vdbeta for procaine in the horse of about 3,500 litres. Since procaine was about 45% bound to equine plasma protein this gives a true Vdbeta for procaine of about 6,500 litres. After subcutaneous injection of procaine HC1 (3.3 mg/Kg) plasma levels peaked at about 400 ng/ml and then declined with a half-life of about 75 minutes. These data were well fitted by Model I when this was modified to include simple first order absorption (K = 0.048 min-1) from the subcutaneous injection site (Model II). After intramuscular injection of procaine penicillin (33,000 I.U./Kg) plasma levels reached a peak at about 270 ng/ml and then declined with a half-life of about 9 hours. These data were approximately fitted by Model II assuming a first order rate constant for absorption of procaine of 0.0024 min-1. After intramuscular injection of procaine HC1 (10 mg/Kg) plasma levels of procaine peaked rapidly at about 600 ng/ml but thereafter declined slowly (+ 1/2 = 2 hours). A satisfactory pharmaco-kinetic model for this intramuscular data could not be developed. An approximation of these data was obtained by assuming the existence of two intramuscular drug compartments, one containing readily absorbable drug and the other poorly absorbable drug (Model III). After intra-articular administration of procaine (0.33 mg/Kg) plasma levels of this drug reached a peak at about 17 ng/ml and then declined with a half-life of about 2 hours. These data were not modelled.     

A fatal case of MAM-2201 poisoning

A 59-year-old man was found dead in his house, where three sachets containing herbal blends were found on a table. The sachet contents were analyzed by gas chromatography–mass spectrometry and found to contain [1-(5-fluoropentyl)-1H-indol-3-yl](4-methyl-1-naphthalenyl)methanone (MAM-2201). The deceased was subjected to forensic autopsy. There were neither external injuries nor endogenous diseases judged by macroscopic and microscopic observations. Liquid chromatography–electrospray ionization–tandem mass spectrometry was used to quantitate the concentrations of MAM-2201 in postmortem samples using deuterated MAM-2201 as internal standard. The MAM-2201 concentrations were: 12.4 ng/ml in whole blood; 18.1 ng/g in the liver; 11.2 ng/g in the kidney; 4.3 ng/g in the brain; and 1,535 ng/g in the adipose tissue. We concluded that the man’s death was caused by acute intoxication with MAM-2201. In addition, we propose that the adipose tissue is the specimen of choice to detect MAM-2201 in the unchanged form. To our knowledge, this is the first report of a fatal MAM-2201 poisoning case. In addition, this report is also the first to describe the distribution of the drug in postmortem human tissues and blood.     

噻吩诺啡及其活性代谢产物的药代动力学研究

目的建立灵敏、专一的可同时测定比格犬血浆中噻吩诺啡及其代谢产物葡萄糖醛酸结合物含量的方法。方法采用HPLC-MS/MS法测定比格犬血中原型和代谢产物的浓度,分析在比格犬中的药代动力学参数。结果噻吩诺啡和代谢产物线性范围分别为0.02～50 ng/ml和0.2～500 ng/ml,定量下限分别为0.02 ng/ml和0.2 ng/ml,日内和日间精密度均小于9%,回收率均大于60%,在样品贮存、处理和分析过程中稳定性良好。比格犬单次口服噻吩诺啡0.2、0.6、1.8 mg/kg后,原型的Cmax分别为1.42、2.08、4.82 ng/ml,AUC分别为8.60、13.24和26.10μg.h/L,Tmax约为0.55～0.65 h,t1/2为10～13 h,MRT约为10～12 h;其代谢产物(噻吩诺啡葡萄糖醛酸结合物)的Cmax分别为1.78、5.03、7.09μg/L,AUC分别为9.37、22.34、41.40μg.h/L,Tmax为0.47～1.1 h,t1/2为18～40 h,MRT在11～17 h。比格犬口服0.2 mg/kg噻吩诺啡生物利用度为12.65%。结论本研究首次报道了噻吩诺啡及其葡萄糖醛酸结合物在比格犬中药代动力学研究,证明药物在犬体内的吸收和转化较快,单次口服盐酸噻吩诺啡低、中、高3种剂量后原型及其葡萄糖醛酸结合物药代动力学过程均符合一级吸收二房室模型。     

Determination of phosphatidyl ethanol (PEth) 16:0/18:1 in dried blood samples of drivers involved in traffic accidents: A pilot study

In routine forensic toxicology practices, blood alcohol concentration (BAC) levels are measured in traffic accidents that ended up in emergency departments. Nevertheless, since the elimination of ethanol from the blood is fast and the detection time is short, BAC cannot indicate the occurrence of chronic excessive alcohol consumption. Phosphatidylethanol (PEth) is a unique ethanol direct biomarker that occurs only in the presence of phospholipase D enzyme in erythrocyte membranes during alcohol intake, and it indicates alcohol intake.In this study, both whole blood and dried blood samples were collected from 50 patients who were admitted to Cukurova University Hospital Emergency Department due to a traffic accident. While studying BAC in whole blood samples, PEth 16:0/18:1 analysis was performed on dried blood samples by LC-MS/MS.According to the BAC (50 mg/dL) value, the legal limit in Turkey, the optimal threshold PEth 16:0/18:1 value was set as 160 ng/mL and over. This study determined that 15 people with above PEth 16:0/18:1 concentrations above 160 ng/mL were classified as excessive alcohol consumption.The data obtained in this study showed a positive correlation between BAC and PEth concentration when driving under the influence of ethanol.     

Analysis of oseltamivir active metabolite, oseltamivir carboxylate, in biological materials by HPLC-UV in a case of death following ingestion of Tamiflu

An HPLC-UV method was established for determination of oseltamivir carboxylate, an active metabolite of oseltamivir, in biological materials. Oseltamivir carboxylate and floropipamide (internal standard) were extracted by using a mixed mode cation exchange extraction cartridge. The recovery yields of oseltamivir carboxylate from spiked blood at concentrations 0.1microg/ml and 1.0microg/ml were 103.8+/-16.9% and 95.0+/-4.3%, respectively. Linearity was obtained in the concentration from 0.1 to 3.0microg/ml. The detection limit was 0.04microg/ml (S/N=3). This method was applied to a case of death from falling after ingestion of a Tamiflu capsule containing 75mg of oseltamivir. Concentration of oseltamivir carboxylate in the femoral blood was 0.4microg/ml. The highest concentration in all specimens was in the liver at 18.3microg/g. In all brain samples, the concentrations were not higher than the quantification limit (0.1microg/g).     

A fatal case of 3,4-methylenedioxypyrovalerone poisoning: coexistence of α-pyrrolidinobutiophenone and α-pyrrolidinovalerophenone in blood and/or hair

We report a fatal case of 3,4-methylenedioxypyrovalerone (MDPV) poisoning. A 35-year-old woman was taken to a nearby hospital after she was found unconscious. MDPV (1,200 ng/ml) and α-pyrrolidinobutiophenone (α-PBP; 200 ng/ml) were detected in her cardiac blood. Hair segments from the victim were also analyzed to confirm chronic drug use. MDPV and α-pyrrolidinovalerophenone (α-PVP) were found in many of the segments tested with the highest MDPV concentration at 22 ng/10-mm hair. We concluded that the woman died of acute MDPV poisoning, and that she had abused MDPV and α-PVP for a long time. To our knowledge, this is the first report dealing with MDPV in human hair segments, and with α-PBP in human specimens.     

Death after excessive propofol abuse

Abuse of the anaesthetic agent propofol (2,6-diisopropylphenol) is rare, but we report a case of a 26-year-old male nurse in which the autopsy showed unspecific signs of intoxication and criminological evidence pointed towards propofol abuse and/or overdose. Intravenously administered propofol is a fast and short-acting narcotic agent, therefore it seemed questionable whether the deceased would have been able to self-administer a lethal overdose before losing consciousness. The blood and brain concentrations corresponded to those found 1–2 min after bolus administration of a narcotic standard dose of 2.5 mg propofol/kg body weight. Extremely high propofol concentrations were found in the urine indicating excessive abuse before death. However, due to the short half-life of propofol, the cumulative effects of repeated injections should not be relevant for toxicity, since this would result in a blood level increase of only 1–2 μg/ml. Furthermore, the detection and quantitation of propofol in three different hair segments indicated chronic propofol abuse by the deceased. The results of the investigation suggest that death was not caused by a propofol overdose but by respiratory depression resulting from overly rapid injection. Received: 1 July 1999 / Accepted: 25 November 1999     

A case of fatal triazolam overdose

A 57-year-old man was found dead lying down in a bamboo thicket. Moderate to severe petechiae were present on his conjunctivae, buccal mucosa, and laryngeal mucosa at autopsy. Cardiac chambers contained a normal volume of fluid blood. Moderate atherosclerosis and fatty liver were observed. No remarkable changes, other than congestion in other organs, were observed. Gas chromatographic screening of the stomach contents, blood and urine was positive for triazolam and alpha-hydroxytriazolam that were confirmed by gas chromatography-mass spectrometry. Blood concentrations of triazolam and free alpha-hydroxytriazolam were 62-251 and 10-66 ng/ml, respectively. A substantial amount of triazolam was detected in bile (1130 ng/ml), but not in urine. Free and total alpha-hydroxytriazolam concentrations were 3920 and 7050 ng/ml, respectively, in the bile and 3710 and 9670 ng/ml, respectively, in urine. Organs contained 216-583 ng/g triazolam. The concentration of free alpha-hydroxytriazolam in the kidney (246 ng/g) was higher than in any other organ. Free alpha-hydroxytriazolam was not detected in the liver. The concentrations of total alpha-hydroxytriazolam in the liver and kidney were 784 and 381 ng/g, respectively. Free to total ratios of alpha-hydroxytriazolam were 0.14-0.56 in fluid samples and 0.56-0.92 in tissue samples, except for the liver. A large quantity of triazolam (8.4 mg) remained in the stomach. The victim probably died of postural asphyxia caused by triazolam poisoning.     

Hair analysis of an unusual case of Chloroquine intoxication

A dead body of middle aged man was exhumed from 6.5 month earth-grave. Autopsy findings were non-specific as the body was completely putrefied. Deceased’s scalp hair and kidney was sent for toxicological analysis. Hair sample (50 mg) was incubated with 1 M NaOH (2 ml). Chloroquine was detected in hair and kidney during basic drug screen performed on GC/MS. For confirmation and quantitation, chloroquine was extracted using Hypersep verify CX SPE cartridges while mass detector was operated in SIM mode using the ions of m/z 245.0, 290.1, 319.0 for chloroquine while ions of m/z 260 and 455 were monitored for nalorphine (internal standard). Chloroquine was present in high concentration in hair (211 ng/mg) as well as in kidney (37.3 mg/kg). Moreover, chloroquine was not detected in the wash solvents, suggesting ingestion of the drug rather than an external contamination of hair. These findings strongly suggested the acute exposure of higher doses of chloroquine to the deceased before death.     

Anesthetized by chloroform before hanging

We present a unique case of suicidal hanging. The deceased was a 31-year-old male who was found hanging from a tree in a dense thicket, with his lower limbs in contact with the ground (partial suspension). There was an apparatus similar to a facial mask placed around his nose and mouth. A strong chemical smell was emanating from the apparatus, which was identified as chloroform (Formyl trichloride/CHCl₃). A ligature with a soft cloth beneath it was around his neck. A ligature mark was present around the neck. The decedent’s blood alcohol levels were 112 mg/dl. The blood and stomach contents were negative for chloroform. A complete death investigation, including scene investigation and complete autopsy examination, confirmed the cause of death as hanging. The manner of death was suicide. This case highlights how the deceased had used several methods whilst committing suicide to minimize pain, including the inhalation of chloroform, which would have also resulted in the inability to engage in protective actions during the act.     

Postoperative analgesic effects of an external cooling system and intra-articular bupivacaine/morphine after arthroscopic cruciate ligament surgery

The aim of this study was to evaluate the analgesic effect of an external cooling system with or without the combined effect of intra-articularly administered bupivacaine/morphine after arthroscopic anterior cruciate ligament (ACL) reconstruction. Fifty patients with isolated ACL insufficiency operated on under general anaesthesia were randomized to three different postoperative treatment groups. Group I was treated with the cooling system during the first 24 h after surgery and an intraarticular injection of 20 ml of physiological saline given at the completion of surgery; in group II, the cooling system was combined with an intra-articular injection of 20 ml bupivacaine 3.75 mg/ml and 1 mg of morphine at the end of the operation; while group III (placebo group) received an intra-articular injection of 20 ml of physiological saline at the completion of surgery. Pain was assessed using a visual analogue scale (VAS) at 1, 2, 4, 6, 24 and 48 h postoperatively. Supplementary analgesic requirements were registered. In group I 80% (16/20) and in group II 90% (18/20) of the patients were satisfied with the postoperative pain control regimen (NS). This was significantly better than in group III, where 30% (3/10) were satisfied. The pain scores were significantly lower in the two treatment groups compared with the placebo group during the entire postoperative period. The pain score was significantly lower in group II than in group I at 24 and 48 h after surgery. The supplementary analgesic requirements were also lower in the two treatment groups compared with the placebo group. No complications due to the use of the cooling system or the intra-articular injections of bupivacaine/morphine were observed. The external cooling system used in this study provides an effective method of obtaining pain relief after arthroscopic surgery. The combination with an intra-articular injection of morphine and bupivacaine results in a slightly greater analgesic effect than the cooling system alone.     

Ethyl glucuronide concentrations in beard hair after a single alcohol dose: evidence for incorporation in hair root

Background

Despite the growing importance of ethyl glucuronide (EtG) in hair for detection of chronic excessive alcohol consumption, the mechanism of incorporation is not yet clear. Deposition from sweat is believed to be the main route. In order to get more information, EtG was determined in daily shaved beard hair after single higher alcohol doses.

Methods

Three volunteers drank within 5.5?h 153, 165 and 200?g ethanol followed by abstinence. Daily shaved beard hair was analysed for EtG using a validated liquid chromatography–tandem mass spectrometry method with a limit of quantification of 2?pg/mg.

Results

For all three volunteers, small concentrations of EtG were already detected 9?h after end of drinking. The concentrations increased to maxima of 182, 242 and 74?pg/mg on days 2 to 4 and then gradually decreased to limit of quantification on days 8 to 10.

Discussion

The time course of EtG is discussed based on literature data about anatomic dimensions of the hair root, physiology of hair growth, kinetics of EtG formation and elimination in blood, and in comparison to literature results about drugs in beard hair. It follows that for beard hair the predominant portion of EtG is incorporated in the upper part of the hair root between suprabulbar region and isthmus leading to a positive zone of about 3?mm (8–9?days) after a single drinking event. Deposition from sweat which is only possible into the residual hair stubble after shaving and in the infundibulum down to the sebaceous gland mouth was found to be of minor importance but could play a greater role in long hair.

Conclusion

It is concluded that EtG in hair fulfils the prerequisites for time-resolved interpretation of segmental concentrations and that a single excessive drinking can be well detected in sufficiently short hair segments. However, in the routinely investigated 3-cm proximal scalp hair segment and using the cutoff of 7?pg/mg, a negative result can be expected with high probability because of dilution by negative hair.     

High blood mirtazapine concentration in a newborn – A case of suspected postpartum infanticide

We report a sudden death of an infant due to mirtazapine poisoning. A 15-day-old newborn boy was found dead when he was sleeping beside his mother who had suffered from panic disorder for approximately 1 year. After giving birth, she complained of palpitations and shaky hands, and was prescribed mirtazapine. The deceased newborn weighed 3,282 g and his height was 55 cm. There were no autopsy findings related to the death. The mirtazapine concentration as quantitated by liquid chromatography-tandem mass spectrometry analysis was 620 ng/mL in right heart blood, and was approximately 10 times higher than the therapeutic level in adults. Because transfer of mirtazapine into breast milk is low, mirtazapine was likely administered intentionally to the newborn. Based on the newborn’s immature renal, liver, and blood–brain barrier function, the cause of death was attributed to mirtazapine poisoning. Poison-related homicide in the infant is rare. We report the first case of intentional mirtazapine poisoning case in a newborn.     

A drug rape case involving triazolam detected in hair and urine

In recent years, there has been heightened awareness regarding the use of drugs to modify a person’s behavior to facilitate crime. A drug rape case involving the potent, short-acting sedative triazolam will be presented. On three occasions, the victim consumed green tea and chocolate before being massaged and ultimately sexually abused. Screening for alcohol, commonly used drugs and illicit substances in blood and urine sampled during the forensic examination 20 h after the last incident, was negative. Consequently, hair samples for chemical analysis were taken from the assaulted individual 34 days after the last incidents. The hair was cut into three 2-cm segments (0–6 cm) that were washed, dissolved in extraction solvent and screened and verified by ultra performance liquid chromatography coupled with time of flight mass spectrometry (UPLC-TOF-MS) and with tandem mass spectrometry (UPLC-MS/MS), respectively. In the 2-cm hair segment corresponding to the period of the alleged assaults, the presence of the sedative triazolam was revealed at a concentration of 1.0 pg/mg hair. The preserved urine sample, taken 20 h after the last incident, was reanalyzed by UPLC-MS/MS for metabolites of triazolam, and 39 μg/l α-hydroxytriazolam was detected in the hydrolyzed urine. This case illustrates that hair is a valuable forensic specimen in situations where natural processes have eliminated the drug from typical biological specimens due to delays in the crime being reported. Furthermore, it was possible to verify the hair finding with a urine sample by detection of a metabolite of triazolam.     

Top ten defence challenges among drinking drivers in Sweden.

This paper gives a review and opinion about defence challenges arising during the prosecution of drinking drivers in Sweden. The statutory limit of blood-alcohol concentration (BAC) is two-tiered; 0.2 mg/g (21 mg/100 ml) and 1.5 mg/g (159 mg/100 ml). In rank order of occurrence, the top ten defence challenges are shown in Table I. The hip-flask defence occurs in more than 90% of all challenges. Because the burden of proof in Swedish law rests on the prosecution, the charge of driving under the influence of alcohol is often dropped when alleged drinking after the offence is an issue.     

Violence under the influence of methylphenidate as determined by hair analysis

A 26-year-old woman under the influence of alcohol assaulted another person at a party in late November 2008. Afterward, she claimed that her drink was spiked with Ritalin, which would account for her bizarre behavior. Neither urine nor blood was sampled, because of the elapsed time after the incident. Hair was collected in late February 2009 and sent to our laboratory for methylphenidate testing. The hair strand was decontaminated twice using methylene chloride and then segmented. Each segment was cut into small pieces and about 30 mg was incubated in an ultrasonic bath for 3 h in buffer solution at pH 5.5 in the presence of 100 ng of 3,4-methylenedioxymethamphetamine (MDMA)-d ₅ used as internal standard (IS). Each mixture was neutralized with NaOH and extracted with hexane/ethyl acetate (90:10). After evaporation, the residue was reconstituted in 200 μl of acetonitrile/formate buffer (5:95). Each sample was subjected to liquid chromatography-tandem mass spectrometry (LC-MS-MS) with a gradient elution of acetonitrile and formate buffer. For both methylphenidate and MDMA-d ₅, detection was performed with two product ions each (m/z 234 to 84 and 56 for methylphenidate; m/z 199 to 135 and 165 for IS). The method showed a linear response from 0.5 to 500 pg/mg with a limit of quantitation of 0.5 pg/mg. Hair collection was made 3 months after the incident, so it was decided that the hair be analyzed in three 4-cm segments. Methylphenidate was detected only in the proximal segment at 1.0 pg/mg, while the result was negative in the other two segments. These results support the suggestion of a single exposure to methylphenidate in the previous 4 months, which includes the time of the party.