Forensic Science International

Editorial Board

2012-02-10

Editorial

2012-02-10

Detection of the synthetic drug 4-fluoroamphetamine (4-FA) in serum and urine

J. Röhrich, J. Becker, T. Kaufmann, S. Zörntlein, R. Urban — 2011-05-05

Abstract: 4-Fluoroamphetamine (4-FA) was detected in the blood and urine of two individuals suspected for driving under the influence (DUI). The test for amphetamines in urine subjected to immunoassay screening using the CEDIA DAU assay proved positive. Further investigations revealed a 4-FA cross-reactivity of about 6% in the CEDIA amphetamine assay. 4-FA was qualitatively detected in a general unknown screening for drugs using GC/MS in full scan mode. No other drugs or fluorinated phenethylamines were detected. A validated GC/MS method was established in SIM mode for serum analysis of 4-FA with a limit of detection (LOD) of 1ng/mL and a lower limit of quantification (LLOQ) of 5ng/mL. Intra-assay precision was approx. 4% and inter-assay precision approx. 8%. Applying this method, the 4-FA serum concentrations of the two subjects were determined to be 350ng/mL and 475ng/mL, respectively. Given the pharmacological data of amphetamine, 4-FA psychoactive effects are to be expected at these serum levels. Both subjects exhibited sympathomimetic effects and psychostimulant-like impairment accordingly.

Identification of 1-butyl-3-(1-(4-methyl)naphthoyl)indole in a herbal mixture

F. Westphal, F.D. Sönnichsen, S. Thiemt — 2011-05-05

Abstract: Besides the cannabinoid mimetic JWH-073, a novel 4 methylnaphthoyl homologue of JWH-073 was detected in a herbal mixture. The structure of the compound was elucidated after thin layer chromatographic enrichment from the herbal mixture by nuclear magnetic resonance (NMR) and gas chromatographic mass spectrometric (GC–MS) analysis. The paper outlines data after GC–MS, liquid chromatography mass spectrometry (LC–MS) and NMR spectroscopy, and describes the structure elucidation.

The prevalence of drugs in injured drivers

2011-03-07

Abstract: In mid 2009 Victoria introduced compulsory drug testing of blood taken from all injured drivers taken to hospital. Δ9-Tetrahydrocannabinol (THC), methylamphetamine (MA) and 3,4-methylenedioxy-methylamphetamine (MDMA) are prohibited and if drivers are positive to any amount an automatic penalty is enforced. Laboratory screens were conducted on preserved blood using ELISA testing for cannabis metabolite and methylamphetamines and a fully validated LC–MS/MS method for 105 drugs including THC, amphetamines, opioids, benzodiazepines, antidepressants and antipsychotics and a number of other psychoactive substances using a minimum of two transitions per drug. Conventional GC-testing for ethanol was used to screen and quantify the presence of alcohol. 1714 drivers were tested and showed alcohol in 29% (≥0.01g/100mL) and drugs in 35%. The positive rate for the three drugs prohibited by legislation was 12.5%. The prevalence of THC, MA and MDMA was 9.8%, 3.1%, and 0.8%, respectively. The range of THC concentrations in blood was 2–42ng/mL (median 7) of which 70% had a concentration of 10ng/mL or higher. The range of concentrations for MA and MDMA was 0.02–0.4 and 0.03–0.3mg/L (median for both drugs was 0.05mg/L). Drugs of any type were detected in 35% of cases. The other drugs were largely prescribed drugs such as the antidepressants (9.3%) and benzodiazepines (8.9%). Neither 6-acetylmorphine nor cocaine (or benzoylecgonine) was detected in these cases.

Concentrations of free-morphine in peripheral blood after recent use of heroin in overdose deaths and in apprehended drivers

A.W. Jones, A. Holmgren, J. Ahlner — 2011-02-25

Abstract: The concentration of free-morphine was determined in peripheral (femoral) blood from heroin-related deaths and compared with the concentration in venous blood from impaired drivers. The presence of 6-MAM in blood or urine served as a biomarker for recent use of heroin. Males dominated over females (p<0.001) in both the autopsy cases (88%) and the drivers (91%), although their mean age was about the same 33–35 y (p>0.05). Concentrations of free-morphine in blood were not associated with age of heroin users in Sweden (p>0.05). The median concentration of free-morphine was higher in autopsy cases (0.24mg/L, N=766) compared with apprehended drivers with 6-MAM in blood (0.15mg/L, N=124, p<0.05), and appreciably higher than in drivers with 6-MAM in urine but not in blood (0.03mg/L, N=1823, p<0.001). The free-morphine concentration was above 0.20mg/L in 65% of autopsy cases, 36% of drivers with 6-MAM in blood but only 1.4% of drivers with 6-MAM in urine. Poly-drug deaths had about the same concentrations of free-morphine in blood (0.24mg/L, N=703) as heroin-only deaths (0.25mg/L, N=63). The concentration of morphine in drug overdose deaths (median 0.25mg/L, N=669) was about the same as in traumatic deaths among heroin users (0.23mg/L, N=97). However, the concentration of morphine was lower when the deceased had consumed alcohol (0.18mg/L, N=104) compared with taking a benzodiazepine (0.32mg/L, N=94). The concentration distributions of free-morphine in blood in heroin-related deaths overlapped with the concentrations in impaired drivers, which makes the interpretation of toxicology results difficult without knowledge about tolerance to opiates in any individual case.

Amphetamine, cocaine and cannabinoids use among truck drivers on the roads in the State of Sao Paulo, Brazil

2011-04-21

Abstract: Drugs are important risk factors for traffic accidents. In Brazil, truck drivers report using amphetamines to maintain their extensive work schedule and stay awake. These drugs can be obtained without prescription easily on Brazilian roads. The use of these stimulants can result in health problems and can be associated with traffic accidents. There are Brazilian studies that show that drivers use drugs. However, these studies are questionnaire-based and do not always reflect real-life situations. The purpose of this study was to demonstrate the prevalence of drug use by truck drivers on the roads of Sao Paulo State, Brazil, during 2009. Drivers of large trucks were randomly stopped by police officers on the interstate roads during morning hours. After being informed of the goals of the study, the drivers gave written informed consent before providing a urine sample. In addition, a questionnaire concerning sociodemographic characteristics and health information was administered. Urine samples were screened for amphetamines, cocaine, and cannabinoids by immunoassay and the confirmation was performed using gas chromatography–mass spectrometry (GC–MS). Of the 488 drivers stopped, 456 (93.4%) provided urine samples, and 9.3% of them (n=42) tested positive for drugs. Amphetamines were the most commonly found (n=26) drug, representing 61.9% of the positive samples. Ten cases tested positive for cocaine (23.8%), and five for cannabinoids (11.9%). All drivers were male with a mean age of 40±10.8 years, and 29.3% of them reported some health problem (diabetes, high blood pressure and/or stress). A high incidence of truck drivers who tested positive for drug use was found, among other reported health problems. Thus, there is an evident need to promote a healthier lifestyle among professional drivers and a need for preventive measures aimed at controlling the use of drugs by truck drivers in Brazil.

The incidence of drugs of impairment in oral fluid from random roadside testing

2011-06-13

Abstract: Oral fluid (OF) has become a popular specimen to test for presence of drugs, particularly in regards to road safety. In Victoria, OF specimens from drivers have been used to test for the presence of methylamphetamine (MA) and Δ9-tetrahydrocannabinol (THC) since 2003 and 3,4-methylenedioxy-N-methylamphetamine (MDMA) since 2006. LC–MS/MS has been used to test the most recent 853 submitted OF specimens from Victoria Police for 31 drugs of abuse including those listed in the Australian Standard AS4760-2006. At least one proscribed drug was detected in 96% of drivers, of which MA was the most common (77%), followed by THC (42%), MDMA (17%) and the combination of all three (3.9%). Opioids were detected in 14% of drivers of which 4.8% were positive for 6-acetylmorphine and 3.3% for methadone. The incidence of the opioids tramadol (1.2%) and oxycodone (1.1%) were relatively low. Cocaine (8.0%) was as commonly detected as benzodiazepines (8.0%), and was almost always found in combination with MA (7.9%). Samples positive to benzodiazepines were largely due to diazepam (3.5%) and alprazolam (3.4%), with only 0.2% of drivers combining the two. Ketamine was also detected in 1.5% of cases. While the incidences of the proscribed drugs itself are concerning, it is clear that many drivers are also using other drugs capable of causing impairment.

Significantly increased detection rate of drugs of abuse in urine following the introduction of new German driving licence re-granting guidelines

Ronald Agius, Thomas Nadulski, Hans-Gerhard Kahl, Bertin Dufaux — 2011-11-18

Abstract: In this paper we present the first assessment of the new German driving licence re-granting medical and psychological assessment (MPA) guidelines by comparing over 3500 urine samples tested under the old MPA cut-offs to over 5000 samples tested under the new MPA cut-offs. Since the enzyme multiplied immunoassay technique (EMIT) technology used previously was not sensitive enough to screen for drugs at such low concentrations, as suggested by the new MPA guidelines, enzyme-linked immunosorbent assay (ELISA) screening kits were used to screen for the drugs of abuse at the new MPA cut-offs. The above comparison revealed significantly increased detection rates of drug use or exposure during the rehabilitation period as follows: 1.61, 2.33, 3.33, and 7 times higher for 11-nor-delta-9-tetrahydrocannabinol-9-carboxylic acid (THC-COOH), morphine, benzoylecgonine and amphetamine respectively. The present MPA guidelines seem to be more effective to detect non-abstinence from drugs of abuse and hence to detecting drivers who do not yet fulfil the MPA requirements to regain their revoked driving licence.

Validation of LUCIO®-Direct-ELISA kits for the detection of drugs of abuse in urine: Application to the new German driving licence re-granting guidelines

Ronald Agius, Thomas Nadulski, Christine Moore — 2011-11-10

Abstract: LUCIO®-Direct-enzyme linked immunosorbent assay (ELISA) tests were validated for the screening of drugs of abuse cannabis, opiates, amphetamines and cocaine in urine for the new German medical and psychological assessment (MPA) guidelines with subsequent gas chromatographic–mass spectrometric (GC–MS) confirmation. The screening cut-offs corresponding to 10ng/mL 11-nor-delta-9-tetrahydrocannabinol-9-carboxylic acid (THC-COOH), 50ng/mL amphetamine, 25ng/mL morphine and codeine and 30ng/mL benzoylecgonine were chosen at the point where the number of false negatives was lower than 1%. Due to their accuracy, ease of use and rapid analysis, these ELISA tests are very promising for cases where a large proportion of the tests are expected to be negative such as for abstinence monitoring as part of the driving licence re-granting process.

Prevalence of drug abuse among workers: Strengths and pitfalls of the recent Italian Workplace Drug Testing (WDT) legislation

2011-04-11

Abstract: Background: In 2008 a Workplace Drug Testing (WDT) law became effective in Italy for workers involved in public/private transportation, oil/gas companies, and explosives/fireworks industry with the aim to ensure public safety for the community.Aims: To examine and elaborate WDT data collected on a large group of workers (over 43,500) during March 2009–February 2010 in order to highlight pros and cons and to draw suggestions for policies in the field.Setting: Northern Italy.Methods: After ≤24h notification, workers provided a urine sample screened for opiates, methadone, buprenorphine, cocaine, amphetamines, ecstasy, and cannabinoids (THC) by immunoassay. Positives were confirmed by GC–MS.Results: The positive rate was 2.0%, THC being most frequent drug (1.3%; cocaine, 0.4%; opioids, 0.3%). 6.9% of the positive workers tested positive for ≥2 classes (most often THC+cocaine). Gender ratio and mean age were significantly lower in positives (F/M=0.007; 35.5±8.3years) than negatives (0.016 and 40.7±9.5, respectively). No decline in rates of positives and an increase of diluted samples over time were observed. The highest rates of positives were detected when sampling was performed just before/after week-end and during morning hours. Possible correlation between job type and drugs used were observed (e.g. more cocaine positives among road vehicle-drivers than among lift truck-drivers). Declared use of medicine/illicit drugs during the preceding week showed that illicit drug use was likely not always detected in urine and that almost 4% workers declared use of medicine drugs possibly affecting performance.Conclusions: This survey enabled to evidence relevant pitfalls of the law and to define strategies to improve the outcomes of WDT policies.

Ethyl glucuronide in human hair after daily consumption of 16 or 32g of ethanol for 3 months

Robert Kronstrand, Linda Brinkhagen, Fredrik H. Nyström — 2011-03-02

Abstract: The overall objectives of the study were to develop a sensitive method for ethyl glucuronide (EtG) determination in hair and then investigate if a low or moderate intake of ethanol could be differentiated from total abstinence. Forty-four subjects were included in the study, 12 males (7 drinkers and 5 abstinent) and 32 females (14 drinkers and 18 abstinent). The study lasted 3 months and the female drinkers consumed one glass (16g of ethanol) and the males consumed two glasses (32g of ethanol) of wine (13.5–14%) daily. Hair samples were collected as close as possible above the skin and the proximal 2cm were analyzed for EtG. Hair was cut into pieces of about 0.5cm length and washed before incubation overnight in water and then extracted on Clean Screen EtG Carbon columns. The LC/MS/MS system consisted of a Waters ACQUITY UPLC connected to an API 4000 triple quadrupole instrument. Two transitions for EtG and one for the internal standard EtG-D5 were measured. The method was linear from 60 to 10,000pg/sample. Imprecision studies were performed at three levels as well as with an authentic sample. Total imprecision was 16% at 200pg/sample, 8% at 1000pg/sample, 6% at 8000pg/sample and 13% at 29pg/mg in the authentic sample. Of those who drank two glasses of wine every day, four had measurable amounts of EtG in their hair (5–11pg/mg), and in only one of the females drinking one glass of wine EtG was quantified (3pg/mg). Among the 23 abstinent subjects two had traces of EtG in the hair. We conclude that persons who ingested 16 or 32g of ethanol daily for 3 months presented with low concentrations of EtG in hair, well below the proposed threshold for overconsumption set at 30pg/mg. In addition, none of those who ingested 16g/day had concentrations over the proposed abstinence threshold of 7pg/mg.

Analysis of drugs of abuse in hair: Evaluation of the immunochemical method VMA-T vs. LC–MS/MS or GC–MS

Markus R. Baumgartner, Rosetta Guglielmello, Monique Fanger, Thomas Kraemer — 2011-10-24

Abstract: Hair analysis is an elaborate and time-consuming multi-step process. The immunometric test VMA-T from Comedical has been evaluated as screening assay for hair analysis. From routine work, authentic samples were selected that were positive for opiates, cocaine, MDMA-type drugs, amphetamines, methadone or THC. These hair samples were investigated by LC–MS or GC–MS and the VMA-T procedure, respectively. Using the cut-off values recommended by the Society of Hair Testing, the VMA-T method discriminates with good sensitivity between negative and positive hair samples and is an expedient screening method for drugs in keratinized matrices such as hair. In order to save time and resources, the residue of the VMA-T extraction solution can be reused for confirmation analysis by LC–MS except for cocaine.

Evaluation of two immunoassay procedures for drug testing in hair samples

F. Musshoff, K.M. Kirschbaum, K. Graumann, C. Herzfeld, H. Sachs, B. Madea — 2011-05-25

Abstract: A preliminary initial enzyme-linked immunosorbent assay (LUCIO®-Direct ELISA kit) and a preliminary DRI® enzyme immunoassay were evaluated for drug detection in head hair with respect to lowered cutoff values recommended in Germany for the control of abstinence in cases of re-granting of drivers’ licences. Following drug classes were included: cannabinoids, opiates, cocaine like substances, amphetamine, methamphetamine (and methylenedioxyamphetamines), methadone, and benzodiazepines. 759 analyses were performed using LUCIO®-Direct ELISA kits and 936 analyses using DRI® enzyme immunoassay tests. Sample size for each drug group and immunoassay test reached from 74 to 178.The LUCIO®-Direct ELISA kit revealed a sensitivity of 91% for amphetamine up to 98% for methadone (methamphetamine 92%, cocaine 94%, opiates 94%, benzodiazepines 96%) and values of specificity of 72% for methadone up to 89% for amphetamine and benzodiazepines. The test was not useful for a preliminary screening for tetrahydrocannabinol (sensitivity of 65%) in consideration of a suggested cutoff of 0.02ng/mg. The DRI® enzyme immunoassay test was only useful for morphine and cocaine testing at low recommended new cutoff values (0.1ng/mg) revealing sensitivities of 94% and 99%, respectively.

Detection and validated quantification of 21 benzodiazepines and 3 “z-drugs” in human hair by LC–MS/MS

Kristina Y. Rust, Markus R. Baumgartner, Natascha Meggiolaro, Thomas Kraemer — 2011-08-29

Abstract: A method for detection and quantification of 21 benzodiazepines and the pharmacologically related “z-drugs” in human hair samples was developed and fully validated using liquid chromatography coupled to tandem mass spectrometry (LC–MS/MS). After methanolic and methanolic/aqueous extraction, the analytes were separated using two different LC–MS systems (AB Sciex 3200 QTRAP and AB Sciex 5500 QTRAP). Separation columns, mobile phases and MS modes for both systems were: Phenomenex Kinetex, 2.6μm, 50/2.1; 5mM ammonium formate buffer pH 3.5/methanol, total flow 0.75mL/min; electrospray ionization (ESI), multiple reaction monitoring (MRM), information dependent acquisition (IDA), enhanced product ion scan (EPI). The assays were found to be selective for the tested compounds (alprazolam, 7-aminoclonazepam, 7-aminoflunitrazepam, bromazepam, chlordiazepoxide, clonazepam, N-desalkylflurazepam, diazepam, flunitrazepam, flurazepam, alpha-hydroxymidazolam, lorazepam, lormetazepam, midazolam, nitrazepam, nordazepam, oxazepam, phenazepam, prazepam, temazepam, triazolam, zaleplon, zolpidem and zopiclone), all validation criteria were in the required ranges according to international guidelines, except for bromazepam. Matrix effects, and process efficiencies were in the acceptable ranges evaluated using the post-extraction addition approach. Lower limits of quantification were between 0.6 and 16pg/mg of hair. The LC–MS/MS assay has proven to be applicable for determination of the studied analytes in human hair in numerous authentic cases (n=175).

Segmental hair analysis can demonstrate external contamination in postmortem cases

Pascal Kintz — 2011-02-28

Abstract: Excluding laboratory mistakes, a false positive hair result can be observed in case of contamination from environmental pollution (external contamination) or after drug incorporation into the hair from the individual body fluids, such as sweat or putrefactive fluid (post mortem artifact). From our 20 years experience of hair testing, it appears that artifact(s) cannot be excluded in some post mortem cases, despite a decontamination procedure. As a consequence, interpretation of the results is a challenge that deserves particular attention. Our strategy will be reviewed in this paper, based on six cases.In all cases, a decontamination procedure with two washes of 5ml of dichloromethane for 5min was performed and the last dichloromethane wash was negative for each target drug. From the histories, there was no suspicion of chronic drug use. In all six cases, the concentrations detected were similar along the hair shaft, irrespective of the tested segment. We have considered this as indicative of external contamination and suggested to the forces or the judges that it is not possible to indicate exposure before death. In contrast to smoke, it seems that contamination due to aqueous matrices (sweat, putrefactive fluid, blood) is much more difficult to remove.To explain potential incorporation of 7-aminoflunitrazepam via putrefactive material, the author incubated negative hair strands in blood spiked at 100ng/ml and stored at +4°C, room temperature and +40°C for 7, 14 and 28 days. After routine decontamination, 7-aminoflunitrazepam tested positive in hair, irrespective of the incubation temperature, as early as after 7 days (233–401pg/mg). In all periods, maximum concentrations were observed after incubation at room temperature. The highest concentration (742pg/mg) was observed after 28 days incubation at room temperature.It is concluded that a standard decontamination procedure is not able to completely remove external contamination in case of post mortem specimens. Homogenous segmental analyses can be probably indicative of external contamination and therefore a single hair result should not be used to discriminate long-term exposure to a drug. Nor should the presence of a metabolite be considered as a discrimination tool, as it can also be present in putrefactive material.

Hair testing and self-report of cocaine use

Claudia Vignali, Cristiana Stramesi, Micol Vecchio, Angelo Groppi — 2011-06-06

Abstract: Hair analysis is a useful tool in both clinical and forensic fields: it allows information about drugs of abuse (DOA) consumption to be obtained. However, in spite of analytical results, sometimes patients continue to deny using drugs or, on the contrary, insist on describing themselves as severe drug addicts; indeed there are often considerable difficulties in getting truthful statements about the real amount of drugs used.In this study we have tried to compare cocaine concentration in hair samples with self-reported drug intake. We enrolled 113 subjects (61 Africans, 52 Caucasians) who had been recently sent to jail. They were asked to tell about their use of illicit drugs during the last three months and then submitted to hair analysis. Hair segments (3cm) were analyzed by GC–MS for amphetamines, cocaine and opiates. Useful data was obtained from 82 subjects, separated into two main groups on account of ethnic origin (African or Caucasian) and divided further into daily, weekly and monthly users. The results showed qualitative results and self-reported consumption to be in good agreement, although the correlation between frequency of consumption and concentration in hair revealed sometimes higher concentrations in contrast with the admission of low consumption. There was a definite separation between occasional and daily use (especially in Caucasian people), while concentrations found where weekly use was reported were more variable. Concentrations of cocaine measured in Africans’ hair were much higher than in Caucasians’.Even if this study is exclusively based on self-report, it provides some interesting information in order to differentiate the frequency of consumption, and especially underlines the great importance of ethnic bias on hair analysis.

Simultaneous analysis of psychotropic phenylalkylamines in oral fluid by GC–MS with automated SPE and its application to legal cases

2011-03-07

Abstract: Phenylalkylamine derivatives, such as methamphetamine (MA), amphetamine (AM), 3,4-methylenedioxymethamphetamine (MDMA), 3,4-methylenedioxyamphetamine (MDA), phentermine (PT), fenfluramine (FFA) and phenmetrazine (PM), and ketamine (KT) are widely abused recreational or anorectic drugs in Korea and are regulated under the Controlled Substance Act in Korea. Phenylalkylamines and ketamine analysis is normally performed using both urine and hair samples but there is no established method for the simultaneous analysis of all these phenylalkylamines and ketamine in oral fluids. Oral fluid is easy to collect/handle and can provide an indication of recent drug abuse. In this study, to confirm the presence of phenylalkylamine derivatives and ketamine in oral fluid after screening with an immunoassay, an analytical method using automated solid phase extraction (SPE) and gas chromatography–mass spectrometry (GC–MS) was developed and fully validated according to international guidelines. The applicability of the assay was demonstrated by analyzing of authentic oral fluid samples and the results of oral fluid analysis were compared with those in urine and hair to to evaluate the feasibility of oral fluid in forensic cases. The recovery of phenylalkylamines and ketamine from oral fluid collection devices was also assessed. Oral fluid specimens from 23 drug abuse suspects submitted by the police were collected using Salivette ® (Sarstedt, Nümbrecht, Germany), Quantisal™ (Immunalysis, Pomona, CA) or direct expectoration. The samples were screened using a biochip array analyzer (Evidence InvestigatorTM, Randox, Antrim, UK). For confirmation, the samples were analyzed by GC–MS in selected-ion monitoring (SIM) mode after extraction using automated SPE (RapidTraceTM, Zymark, MA, USA) with a mixed-mode cation exchange cartridge (CLEAN SCREEN®, 130mg/3ml, UCT, PA, USA) and derivatization with trifluoroacetic anhydride (TFA). The results from the immunoassay were consistent with those from GC–MS. Twenty oral fluid samples gave positive results for MA, AM, PT and/or PM among the 23 cases, which gave positive results in urine and/or hair. Although large variations in the MA, AM, PT and PM concentrations were observed in three different specimens, the oral fluid specimen was useful for demonstrating phenylalkylamines and ketamine abuse as an alternative specimen for urine.

Oral fluid results compared to self reports of recent cocaine and heroin use by methadone maintenance patients

Edward J. Cone — 2011-03-07

Abstract: Introduction: Although self reports of illicit drug use may not be reliable, this information is frequently collected and relied upon by national drug surveys and by counselors in drug treatment programs. The addition of oral fluid testing to these programs would provide objective information on recent drug use.Aim: The goal of this study was to compare oral fluid tests for cocaine, benzoylecgonine, 6-acetylmorphine, morphine, codeine and 6-acetylcodeine to self reports of recent cocaine and heroin use by patients in an outpatient methadone treatment program.Methods: Patients (n=400) provided an oral fluid specimen and completed a short questionnaire on illicit drug use over the last seven days. Oral fluid was collected with the Intercept® Oral Fluid Collection device. Oral fluid was analyzed by a validated assay using liquid chromatography coupled with tandem mass spectrometry. The presence of an analyte was confirmed if all identification criteria were met and its concentration (ng/mL) was ≥LOQ (cocaine, 0.4; benzoylecgonine, 0.4; morphine, 2; codeine, 2; 6-acetylmorphine, 0.4; and 6-acetylcodeine, 1).Results: Analyses of oral fluid specimens collected from the 400 methadone maintained patients revealed that a majority (95%) of subjects who admitted to recent cocaine use were confirmed positive, whereas slightly more than 50% were confirmed positive who admitted to heroin over the last seven days. For those patients who denied recent cocaine and heroin use, approximately 30% were positive for cocaine and 14% were positive for heroin.Conclusion: Oral fluid testing provides an objective means of verifying recent drug use and for assessment of patients in treatment for substance use disorders.

A rapid and sensitive method for the identification of delta-9-tetrahydrocannabinol in oral fluid by liquid chromatography–tandem mass spectrometry

Anna Molnar, John Lewis, Philip Doble, Glyn Hansen, Tatiana Prolov, Shanlin Fu — 2011-03-02

Abstract: A fast and sensitive method was developed for detecting delta-9-tetrahydrocannabinol (THC) in oral fluid by liquid chromatography–tandem mass spectrometry (LC–MS/MS). The method is suitable for samples of small volume and low concentration. For method development and validation, neat oral fluid (200μL) spiked with THC and d3-THC (internal standard) was extracted via liquid–liquid extraction (LLE). The LLE method had an extraction efficiency of 75% with no significant matrix effects observed in either diluted or neat oral fluid samples. LC was performed on a Zorbax Eclipse XDB-C18 Rapid Resolution HT column (2.1mm×50mm, 1.8μm particle size) with positive electrospray ionisation and selected reaction monitoring. The total run time was an efficient 3.5min in isocratic elution mode. The limit of quantification was 1ng/mL and the analysis was linear over the range of 1–500ng/mL with a correlation coefficient of 0.9998. The imprecision (RSD) of the method was 13% and inaccuracy (MRE) was 4%. The method was subsequently applied to two neat oral fluid samples taken from a chronic cannabis smoker. It was also applied to buffer diluted residual oral fluid samples (n=48) collected using the Cozart RapiScan® system through the Roadside Drug Testing Program (RDTP) in NSW, Australia. A stability study was performed that revealed freezing or refrigerating resulted in comparable decreases in THC recovery from neat oral fluid at the end of two weeks of storage. Storage at room temperature even for one day invoked significant losses and is not recommended.

Sexual abuse and anti-wrinkle cream: Evidence from octocrylene

Carole Jamey, Antoine Tracqui, Bertrand Ludes — 2011-03-03

Abstract: We report an alleged case of sexual assault in which an anti-wrinkle cream could have been used as a lubricant. Three anorectal samples taken from the victim were given to us in an attempt to document the presence of remains of the cream involved. After examining the composition of the cream, octocrylene (OCT) was selected as the most relevant marker for this analysis.An ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) method was developed for identification of OCT. Anorectal samples were diluted with methanol and injected onto an Acquity BEH C18 column using a gradient mode with 0.1% formic acid/acetonitrile as the mobile phase. Data were acquired using positive electrospray ionization and multiple reaction monitoring. Three transitions were selected for OCT (m/z 362.2>250.0, m/z 362.2>232.0 and m/z 362.2>204.0).The analysis of the cream seized at the offender's home confirmed the presence of OCT as an ingredient, and the analysis of extracts from the anorectal samples also allowed the formal identification of OCT. These results strongly suggest that a cosmetic containing octocrylene as an ingredient has in fact been applied to the anus of the alleged victim.

Fatal tolperisone poisoning: Autopsy and toxicology findings in three suicide cases

Frank Sporkert, Christophe Brunel, Marc P. Augsburger, Patrice Mangin — 2011-06-20

Abstract: Tolperisone (Mydocalm®) is a centrally acting muscle relaxant with few sedative side effects that is used for the treatment of chronic pain conditions. We describe three cases of suicidal tolperisone poisoning in three healthy young subjects in the years 2006, 2008 and 2009. In all cases, macroscopic and microscopic autopsy findings did not reveal the cause of death.Systematic toxicological analysis (STA) including immunological tests, screening for volatile substances and blood, urine and gastric content screening by GC–MS and HPLC–DAD demonstrated the presence of tolperisone in all cases. In addition to tolperisone, only the analgesics paracetamol (acetaminophen), ibuprofen and naproxen could be detected. The blood ethanol concentrations were all lower than 0.10g/kg. Tolperisone was extracted by liquid–liquid extraction using n-chlorobutane as the extraction solvent. The quantification was performed by GC–NPD analysis of blood, urine and gastric content. Tolperisone concentrations of 7.0mg/l, 14mg/l and 19mg/l were found in the blood of the deceased.In the absence of other autopsy findings, the deaths in these three cases were finally explained as a result of lethal tolperisone ingestion. To the best of our knowledge, these three cases are the first reported cases of suicidal tolperisone poisonings.

Determination of venlafaxine in post-mortem whole blood by HS-SPME and GC-NPD

2011-06-06

Abstract: Venlafaxine is a phenethylamine derivative widely prescribed for the treatment of depression which inhibits both serotonin and norepinephrine reuptake (SNRI). In treatment with antidepressants of patient with depression and other psychiatric disorders there is also increased risk of suicidal thought and behaviour. Several lethal intoxications involving venlafaxine usually among psychotic patients have been reported in the literature. Sample preparation is of the greatest significance for a successful toxicological analysis. The development of simple, effective and rapid extraction procedures of drugs from post-mortem biological samples is a challenge. Headspace-solid phase microextraction (HS-SPME) offers significant advantages such as simplicity, low cost, compatibility with analytical systems, automation and solvent-free extraction. The aim of our work was the optimization of a HS-SPME procedure for the determination of venlafaxine in post-mortem biological samples by gas chromatography (GC) with nitrogen-phosphorous detection (NPD). Venlafaxine was extracted on 100μm Polydimethylsiloxone Coating-Red (PDMS) SPME fiber and determined by GC-NPD. Salt addition, extraction temperature, preheating and extraction time were optimized to enhance the recovery of the extraction from aqueous solution spiked with venlafaxine.Finally the developed procedure was applied to post-mortem biological samples of a fatally poisoned woman by venlafaxine. The drug was quantified in post-mortem blood gastric and oesophagus contents of the deceased woman.A simple and rapid procedure using HS-SPME was developed for sample preparation of venlafaxine in post-mortem biological samples prior to GC-NPD determination. Validation data was satisfactory, thus enabling application in the toxicological analysis of forensic samples.

A fatal intoxication by chloroprene

Annette Rickert, Benno Hartung, Bernd Kardel, Johanna Teloh, Thomas Daldrup — 2011-04-21

Abstract: Objective Chloroprene, 2-chloro-1,3 butadiene, is a volatile synthetic liquid. The chloroprene monomer is extremely reactive and is used for the production of latexes and synthetic rubber such as Neoprene®. Up to now an acute lethal human exposure has been described only once in the literature . The intoxication is associated with nervous system depression, pulmonary edema, narcosis, and respiratory arrest. Case report A 29-year-old chemistry company worker was found unconscious in an empty vessel (depth: 3m) used for chloroprene. The man was dressed in shoes, trousers, a helmet and a respiratory mask. The upper part of the body was unclothed. In spite of reanimation, the man died three hours later in a hospital. Material and methods: All analyses were performed by headspace gas chromatography (HS/GC/FID). In addition, brain, muscle and myocardial muscle were analysed by headspace GC–MS. Results and discussion Autopsy findings: The cause of death could not be determined as the macromorphological findings were unspecific. Toxicology findings The calibration curve of chloroprene in serum shows linearity from 1.0 to 200μg/ml (r2=0.9999) using benzene as internal standard. The LOD is 0.28μg/ml, the LLOQ is 0.99μg/ml. Tissues and body fluids were stored at −20°C till the analysis. Chloroprene was quantified after addition of benzene as the internal standard. It was found in nearly all tissues and body fluids except in the urine and lung. The highest concentrations were detected in the kidney, liver, myocardial muscle and especially in the brain. Furthermore, hexanal was found in all samples except in the urine. The amount of hexanal in some specimens is high, especially in the lung, bile, gastric content and myocardial muscle. Conclusion We assume that a significant amount of chloroprene was not only inhaled but also absorbed through the skin because the man wore a respiratory mask. Presumably the accident would not have happened if the works safety protocols had been followed. The reason why high concentrations of hexanal were found in the tissues could not be clarified.

Reporting a sudden death due to accidental gasoline inhalation

María Antonia Martínez, Salomé Ballesteros, Rafael Alcaraz — 2011-02-28

Abstract: The investigation of uncertain fatalities requires accurate determination of the cause of death, with assessment of all factors that may have contributed to it. Gasoline is a complex and highly variable mixture of aliphatic and aromatic hydrocarbons that can lead to cardiac arrhythmias due to sensitization of the myocardium to catecholamines or acts as a simple asphyxiant if the vapors displace sufficient oxygen from the breathing atmosphere. This work describes a sudden occupational fatality involving gasoline. The importance of this petroleum distillate detection and its quantitative toxicological significance is discussed using a validated analytical method.A 51 year-old Caucasian healthy man without significant medical history was supervising the repairs of the telephone lines in a manhole near to a gas station. He died suddenly after inhaling gasoline vapors from an accidental leak. Extensive blistering and peeling of skin were observed on the skin of the face, neck, anterior chest, upper and lower extremities, and back. The internal examination showed a strong odor of gasoline, specially detected in the respiratory tract. The toxicological screening and quantitation of gasoline was performed by means of gas chromatography with flame ionization detector and confirmation was performed using gas chromatography–mass spectrometry. Disposition of gasoline in different tissues was as follows: heart blood, 35.7mg/L; urine, not detected; vitreous humor, 1.9mg/L; liver, 194.7mg/kg; lung, 147.6mg/kg; and gastric content, 116,6mg/L (2.7mg total).Based upon the toxicological data along with the autopsy findings, the cause of death was determined to be gasoline poisoning and the manner of death was accidental. We would like to alert on the importance of testing for gasoline, and in general for volatile hydrocarbons, in work-related sudden deaths involving inhalation of hydrocarbon vapors and/or exhaust fumes.

Evidence of Haldol (haloperidol) long-term intoxication

Enrico Gerace, Alberto Salomone, Sergio Pellegrino, Marco Vincenti — 2011-03-03

Abstract: A case of intoxication by haloperidol is reported. Haloperidol is a butyrophenone derivative commonly used in many hospital units as an antipsychotic agent. Adverse reactions due to haloperidol intoxication include drowsiness, blurred vision, extrapyramidal effects, tardive dyskinesia, tachycardia, hypotension and muscular rigidity. In August 2008, a 49 year-old female nurse started feeling various symptoms such as muscular rigidity, drowsiness and buccal dyskinesia. After 3 months, she was hospitalized for the worsening of these symptoms. Four months later, she showed once more the same symptoms. Two open water bottles from which the nurse used to drink in the hospital were confiscated and analyzed. Moreover, the nurse was asked to give a sample of her hair for executing the inherent toxicological analyses. Haloperidol was found in both bottles 1 and 2 at a concentration of 31.5μg/mL and 43.6μg/mL, respectively. Based on segmental hair analysis, it was deduced that the nurse consumed haloperidol in the approximate period from August 2008 to March 2009. The higher levels of haloperidol in hair were found in accordance with the periods of most severe appearance of symptoms, requiring the hospitalization of the nurse. The analysis of preservatives and excipients led us to conclude that the pharmaceutical drug was probably added to the water bottles as “Haldol 2mg/mL oral solution”.

Characterization of pentoxyverine metabolites in urine using GC/MS after intoxication with Silomat® cough drops

F. Westphal, Th. Junge, L. Radünz, J. Stegk, G. Rochholz — 2011-03-31

Abstract: A nearly two and a half year old boy was hospitalized after showing symptoms of disorientation and hallucination. The parents remembered the child playing with a bottle of Silomat® cough drops, so that an intoxication was taken into consideration. After liquid/liquid extraction of a urine sample collected in hospital, the underivatized and the acetylated extracts were analyzed by gas chromatography–mass spectrometry (GC/MS) using electron ionization (EI) as well as chemical ionization (CI). In the urine sample high amounts of pentoxyverine (carbetapentane) and several of its metabolites, e.g., different hydrolyzed, desalkylated and ring-hydroxylated products have been identified. The correlation of the results, the observed symptoms, and the access to the Silomat® cough drops reveal an intoxication after ingestion of an unknown amount of the antitussive pentoxyverine. Corresponding EI- and CI-GC/MS spectra are presented characterizing the structure of its metabolites.

Development and validation of a sensitive UPLC–MS/MS method for the analysis of narcotic analgesics in urine and whole blood in forensic context

Ruth Verplaetse, Jan Tytgat — 2011-02-28

Abstract: Narcotic analgesics are widely (ab) used and sometimes only occur in low concentrations in biological samples. Therefore, a highly sensitive liquid chromatography tandem mass spectrometry method was developed for simultaneous analysis of 9 narcotic analgesics and metabolites (buprenorphine, O-desmethyltramadol, fentanyl, norbuprenorphine, norfentanyl, pethidine, piritramide, tilidine and tramadol) in urine and whole blood. Sample preparation was performed on a mixed-mode cation exchange solid phase extraction cartridge with an additional alkaline wash step to decrease matrix effects and thus increase sensitivity. Ionization with electrospray ionization was found to be more efficient than atmospheric pressure chemical ionization. The use of a mobile phase of high pH resulted in higher electrospray ionization signals than the conventional low pH mobile phases. In the final method, gradient elution with 10mM ammonium bicarbonate (pH 9) and methanol was performed on a small particle column (Acquity C18, 1.7μm, 2.1mm×50mm). Selectivity, matrix effects, recovery, linearity, sensitivity, precision, accuracy and stability were validated in urine and whole blood. All parameters were successfully evaluated and the method showed very high sensitivity, which was the major aim of this study. The applicability of the method was demonstrated by analysis of several forensic cases involving narcotic analgesics.

Method validation of a survey of thevetia cardiac glycosides in serum samples

Sarah Kohls, Barbara Scholz-Böttcher, Jürgen Rullkötter, Jörg Teske — 2011-03-07

Abstract: A sensitive and specific liquid chromatography tandem mass spectrometry (HPLC–ESI+-MS/MS) procedure was developed and validated for the identification and quantification of thevetin B and further cardiac glycosides in human serum. The seeds of Yellow Oleander (Thevetia peruviana) contain cardiac glycosides that can cause serious intoxication. A mixture of six thevetia glycosides was extracted from these seeds and characterized. Thevetin B, isolated and efficiently purified from that mixture, is the main component and can be used as evidence. Solid phase extraction (SPE) proved to be an effective sample preparation method. Digoxin-d3 was used as the internal standard. Although ion suppression occurs, the limit of detection (LOD) is 0.27ng/ml serum for thevetin B. Recovery is higher than 94%, and accuracy and precision were proficient. Method refinement was carried out with regard to developing a general screening method for cardiac glycosides. The assay is linear over the range of 0.5–8ng/ml serum. Finally, the method was applied to a case of thevetia seed ingestion.

Assessment of the stability of 30 antipsychotic drugs in stored blood specimens

Eva Saar, Dimitri Gerostamoulos, Olaf H. Drummer, Jochen Beyer — 2011-03-28

Abstract: The stability of 30 common antipsychotics (APs) in spiked whole blood was investigated over ten weeks in a preliminary experiment (designated “P experiment”). Pools of blank blood spiked with drugs at two different therapeutic levels were stored at four different temperatures: 20°C, 4°C, −20°C, and −60°C and extracted once weekly in duplicate, using a previously published method. A loss of >15% of the initial drug concentration was considered to indicate possible instability and the respective drugs were selected for further investigation in a final experiment (designated “F experiment”).Eight APs (chlorpromazine, chlorprothixene, fluspirilene, droperidol, olanzapine, thioridazine, triflupromazine, and ziprasidone) were incorporated into the F experiment. The same conditions were used in both experiments, however only a high therapeutic drug concentration was chosen for the F experiment and the storage time was extended to 20 weeks.All drugs of interest in the F experiment showed significant losses after 20 weeks of storage under at least one storage condition. The most notable results involved olanzapine, where losses of almost 100% in all storage temperatures were observed. Drug degradation in fluspirilene samples was significant after 20 weeks under all storage conditions. Overall, extensive degradation was seen with approximately 80% drug loss when stored at 20°C and 4°C with samples also seriously affected by degradation of up to 50% when stored at −20°C and −60°C, respectively. Ziprasidone remained stable when stored at 4°C, −20°C, and −60°C over 9 weeks, however significant degradation was observed when stored at 20°C, with a loss of almost 100% after 20 weeks of storage.The time period and temperature of storage of biological samples can have a significant influence on the stability of several APs. It is therefore important to be aware of potential changes in drug concentrations during storage when interpreting analytical results.

Stability of diazepam in blood samples at different storage conditions and in the presence of alcohol

2011-05-06

Abstract: Diazepam is one of the mostly used benzodiazepines and it is frequently analyzed in different biological samples, especially blood samples. The diazepam stability in the sample matrices is an important factor regarding reliable data obtaining. The storage is the main factor determining the stability of diazepam in blood samples and it is the object of the study presented. Remaining diazepam amount in spiked whole blood and plasma samples were tested at different storage temperatures, in the absence or presence of sodium fluoride as stabilizer as well as the influence of ethanol on diazepam stability was evaluated. The results of the study indicated that the temperature is the main storage factor affecting diazepam stability. In the fluoride stabilized blood samples the amount of diazepam decreases up to 85% of initial level when stored at −20°C for the period of testing (12 weeks). The presence of low (0.5g/L) or high (3g/L) ethanol concentrations influences the stability of diazepam at −20°C. In whole blood samples, the combination of sodium fluoride and ethanol decreases additionally (15–25%) the concentration of the analyte. Freeze–thaw experiments of whole blood samples show about 5–9% decrease in diazepam concentration after the first cycle. The freeze–thaw experiments on plasma samples, containing ethanol and/or fluoride show insignificant decreases of analyte concentration. Further experiments on benzodiazepines stability at different storage conditions or in combination of different factors should be undertaken in forensic toxicology to ensure the data quality, their reliability and reproducibility.

Characteristics of cannabinoids composition of Cannabis plants grown in Northern Thailand and its forensic application

2011-06-03

Abstract: The Thai government has recognized the possibility for legitimate cultivation of hemp. Further study of certain cannabinoid characteristics is necessary in establishing criteria for regulation of cannabis cultivation in Thailand. For this purpose, factors affecting characteristics of cannabinoids composition of Thai-grown cannabis were investigated. Plants were cultivated from seeds derived from the previous studies under the same conditions. 372 cannabis samples from landraces, three different trial fields and seized marijuana were collected. 100g of each sample was dried, ground and quantitatively analyzed for THC, CBD and CBN contents by GC–FID. The results showed that cannabis grown during March–June which had longer vegetative stages and longer photoperiod exposure, had higher cannabinoids contents than those grown in August. The male plants grown in trial fields had the range of THC contents from 0.722% to 0.848% d.w. and average THC/CBD ratio of 1.9. Cannabis in landraces at traditional harvest time of 75 days had a range of THC contents from 0.874% to 1.480% d.w. and an average THC/CBD ratio of 2.6. The THC contents and THC/CBD ratios of cannabis in second generation crops grown in the same growing season were found to be lower than those grown in the first generation, unless fairly high temperatures and a lesser amount of rainfall were present. The average THC content in seized fresh marijuana was 2.068% d.w. while THC/CBD ratios were between 12.6 and 84.09, which is 10–45 times greater than those of similar studied cannabis samples from the previous study. However, most Thai cannabis in landraces and in trial fields giving a low log10 value of THC/CBD ratio at below 1 may be classified as intermediate type, whereas seized marijuana giving a higher log10 value at above 1 could be classified as drug type. Therefore, the expanded information provided by the current study will assist in the development of criteria for regulation of hemp cultivation in Thailand.

THCVA-A – A new additional marker for illegal cannabis consumption

Lars Radünz, Folker Westphal, Edmund Maser, Gertrud Rochholz — 2011-04-01

Abstract: The aim of the present investigations was to find markers for differentiating between the consumption of illegal cannabis products and legal medication containing fully synthetic Δ9-tetrahydrocannabinol (Δ9-THC), e.g., Marinol® capsules. Δ9-Tetrahydrocannabinolic acid A (Δ9-THCA-A) and Δ9-tetrahydrocannabivarinic acid A (Δ9-THCVA-A) were taken into consideration for analysis, because these substances are the precursors of Δ9-THC and Δ9-tetrahydrocannabivarin (Δ9-THCV) in plant material of Cannabis sativa and are not contained in medical THC formulations. Whereas Δ9-THCA-A is an already well investigated substance, there is little analytical data on Δ9-THCVA-A.The reason for the presented investigations was a case in which a man was tested positive for Δ9-THC during a routine traffic control claiming that the positive serum sample resulted from the intake of a THC medication (Marinol®) and not from consuming illegal cannabis products.Sample preparation consisted of a protein precipitation with acetonitrile. Analysis was carried out on a Thermo Fisher LCQ Deca ion trap LC–MS–MS-system using electron spray ionization (ESI) in negative mode. MS2- and MS3-full scan spectra were recorded for Δ9-THCA-A and Δ9-THCVA-A starting from [M−H]−. Reference spectra were obtained by measuring a Δ9-THCA-A reference solution and an ethanolic cannabis extract for Δ9-THCVA-A as there is no reference material for this cannabinoid available on the market yet.Main transitions for Δ9-THCA-A were m/z 357→313 and 339 in the MS2-spectrum and m/z 313→245 and 191 in the MS3-spectrum. Fragmentation pattern of Δ9-THCVA-A was identical with a difference of 28amu less for the precursor ion as well as the fragments due to a shorter alkyl side chain in the molecule (MS2: m/z 329→285 and 311; MS3: m/z 285→217 and 163).The two plant cannabinoids Δ9-THCA-A and Δ9-THCVA-A could be detected in the serum sample by LC–MS–MS which proved the intake of illegal cannabis products derived from plant material of C. sativa in the described case.

Cross-examination of liquid–liquid extraction (LLE) and solid-phase microextraction (SPME) methods for impurity profiling of methamphetamine

2011-03-07

Abstract: Impurities in 48 methamphetamine (MA) samples were analyzed by liquid–liquid extraction (LLE) and headspace solid-phase microextraction (HS-SPME) methods. MPS-2 autosampler was used to improve reproducibility of SPME method, and nonadecane (C19) diluted with potassium bromide (KBr) powder was used as an internal standard for standardizing retention time. Impurities identified by SPME method showed different patterns compared with LLE method. Non-volatile impurities like methamphetamine dimer were not identified by SPME method, but some volatile impurities like diphenylketone, caprolactam and lots of unknowns were identified only by SPME method. 1-Phenyl-2-propanone (P2P), 1-phenyl-2-propanol and benzylcyanide peaks could be discriminated clearly by SPME method without interference of amphetamine, an artifact originates from MA degradation. Differences in the impurity patterns resulted in different clustering results. When 48 MA samples were classified into 5 LLE and 5 SPME clusters, cross-matching of the clusters resulted in 8 sub-clusters. It shows that combination of the different extraction methods can distinguish the differences which cannot be distinguished by LLE or SPME method alone, and can improve reliability of the profiling results.

Effects of synthetic cannabinoids on electroencephalogram power spectra in rats

2011-06-03

Abstract: Several synthetic cannabinoids have recently been distributed as psychoactive adulterants in many herbal products on the illegal drug market around the world. However, there is little information on pharmacology and toxicology of such compounds. Although Δ9-tetrahydrocannabinol (Δ9-THC), a psychoactive cannabinoid of marijuana, was reported to affect electroencephalograms (EEG) of rats, the effects of synthetic cannabinoids are unknown. We examined the pharmacological activities of three synthetic cannabinoids; cannabicyclohexanol (CCH), CP-47,497 and JWH-018; by analyzing EEG power spectra and locomotor activity after intraperitoneal administration to rats and compared them with those of Δ9-THC. The three compounds significantly increased the EEG power in the frequency range of 5.0–6.0Hz for the first 3h, while Δ9-THC decreased the power spectra in the wide range of 7.0–20.0Hz during the first hour. These results indicate that the effect of the three compounds on EEG is different from that of Δ9-THC. Additionally, CCH, CP-47,497 and JWH-018 significantly decreased the locomotor activity for 11.5h, 11h and 4.5h, respectively, after administration which was longer than that of Δ9-THC (3.5h). Furthermore, all three compounds significantly reduced the total amounts of locomotor activity during a 3-h, 6-h and 12-h period after injection, whereas no statistical difference was observed for the Δ9-THC injection. Among the three compounds, CCH and CP-47,497 exerted a longer duration of the change in the EEG power spectra and suppression of the locomotor activity than JWH-018.

Serotonin toxicity involving MDMA (ecstasy) and moclobemide

J.L. Pilgrim, D. Gerostamoulos, N. Woodford, Olaf H. Drummer — 2011-05-16

Abstract: The use of MDMA (ecstasy) in Australia is a widespread and growing problem, promoting acute toxicity and disease which can lead to premature death in users. We report four cases of fatal serotonin toxicity caused by the combination of MDMA and moclobemide, a reversible MAO-A inhibitor with potent serotonergic activity. Despite the highly reported toxicity of this drug combination, there are very few reports of fatalities attributed to a MDMA and moclobemide interaction. Pathology and toxicology reports, initial police reports and coroners’ findings were examined to determine the circumstances of the deaths. Symptoms of some of the four cases as reported by paramedics and medical staff included hyperthermia, hyperkalemia, profuse sweating, twitching and shaking. Two cases involved moclobemide concentrations consistent with common prescribed doses, while the other two cases involved much higher concentrations often associated with toxicity. Three of these cases presented with some form of heart disease.

Microbial ethanol production: Experimental study and multivariate evaluation

2011-04-07

Abstract: Ethanol can be produced from all the postmortem available substrates, though with higher rates and yields from carbohydrates, during the early stages of putrefaction. The so-called higher alcohols (1-propanol, isobutanol, 2-methyl-1-butanol and 3-methyl-2-butanol) and 1-butanol could be produced, from all the available postmortem substrates. However, a quantitative relationship between the produced ethanol and the potentially produced other alcohols is still missing.The objective of this study was the development of a simple, mathematical model which could be able to approximate the microbial produced ethanol in correlation with other produced alcohols. The selected bacterial species included two Gram+ spore-forming anaerobic bacteria and two (one Gram+ one Gram-) aerobic/facultative anaerobic bacteria, all being common commensals of the digestive tract and common colonizers of the corpse. The selected bacterial strains, Escherichia coli, Clostridium perfrigens, Clostridium sporogenes and Enterococcus faecalis, were cultured separately at 25°C, for 30days, under controlled anaerobic conditions. The produced ethanol and the previously referred alcohols were determined in the culture medium in 24h intervals. Using partial least squares (PLS) regression, the estimation of the relevance score for the available descriptors established the statistical model to assess the ethanol concentration produced by each studied microbe. E. coli, C. perfrigens, and C. sporogenes produced different patterns of ethanol and other alcohols, while E. faecalis produced negligible amounts of ethanol and higher alcohols. In constructing the mathematical models to predict the produced ethanol, 1-propanol, 1-butanol, and isobutanol were significant for C. perfrigens and C. sporogenes, while 1-butanol, 1-propanol, and methyl-butanol were significant for E. coli. The applicability of these models was tested in microbial, anaerobic cultures of normal human blood and plasma at 25°C. The results indicate that factors such as the type of microbe species, the glucose content and the medium composition apparently affect the procedure of microbial ethanol, and other alcohols production. However, the models can be applied with acceptable accuracy and they show potential for application in real postmortem cases.