Challenges for cocaine detection in smuggling samples.

Screening of seized cocaine powders is routinely performed by means of colour tests. An alternative fast screening technique is Mid-InfraRed (MIR) spectroscopy. In the context of smuggling cases, however, drugs are often processed to circumvent detection. In this study, the current screening techniques (cocaine colour test and MIR spectroscopy using libraries and chemometrics) were applied to five smuggling cases. For each case, all samples were first screened with a cocaine colour test and MIR analysis, followed by confirmation analyses with GC-MS and GC-FID to identify and quantify cocaine and cutting agents. Finally, Scanning Electron Microscopy-Energy Dispersive X-ray spectroscopy (SEM-EDX) analyses were performed for additional characterization. All smuggling samples tested negative, both on-site as in the laboratory, for cocaine with the cocaine colour test. Four smuggling cases consisted of coloured samples. Consequently the colour test result was influenced because discolouration of the test showed almost the same colour as the colour of the powders (brown, green, red or black). In contrast, the (coloured) powders could be measured with MIR, but the MIR spectra showed no hit for cocaine using a reference library search. Moreover, cocaine was not detected in four out of the five cases after application of a chemometric classification model. GC-MS analysis, the golden standard for identification, resulted in a positive identification of cocaine in all cases. These samples contained cocaine ranging between 0.8w% and 35w%. Taking into account the results of the screening, the chromatographic and the SEM-EDX analyses, it could be presumed that cocaine was masked. False negative screening results were caused by chemically modified cocaine and/or cocaine mixed with coloured powders. In additional experiments, a sample extraction step prior to the screening techniques was performed. Two sample preparation methods (acetone and ethyl acetate) were tested and resulted in a positive screening of cocaine with the colour test and/or MIR spectroscopy. It can be concluded that the outcome of screening techniques such as colour tests and MIR spectroscopy is only presumptive and should always be confirmed.

Evaluation of data preprocessings for the comparison of GC-MS chemical profiles of seized cannabis samples.

Cannabis is the most frequently used illicit drug in Belgium, where it is mainly cultivated indoor. To improve the fight against this drug, cannabis-profiling methods are required. Cannabis is a natural product and its chemical composition depends on many factors, which cause a high heterogeneity and variability in the secondary metabolites, and make this study challenging. The aim of this study is to combine cannabis profiling with statistical methodology to evaluate the intra (within)- and inter (between)-plantation variabilities with the goal to define a suitable approach linking seized marijuana to given plantations. The data set used contains 46 samples from 9 locations. The chemical profiles, consisting of data from eight cannabinoids, are obtained by gas chromatography - mass spectrometry. The raw data (peak areas) is pretreated with different preprocessing methods. The Pearson correlation coefficients between intra-location profiles were calculated after each pre-treatment, and the 95 and 99 % confidence limits determined. All preprocessed data were then compared with the internal standard normalization reference method with the aim to minimize the overlap between intra- and inter-location results, i.e. to reduce the number of false positives, and to obtain the best discrimination. Furthermore, cross-validation was used to evaluate the model originating from the most efficient data pre-treatment technique. The best results were obtained, when the peak areas were normalized to the internal standard with subsequent calculation of the fourth root. It results in a reduction of false positives for both confidence limits to 11 % and 14 % compared to 21 % and 27 % for the reference method. Cross-validation reveals similar false positive results as for the calibration set. In conclusion, when preprocessing the data, an improved model is obtained resulting in a significant decrease in the number of false positives. After studying the predictive performance of the model, it appears to be representative for the entire plantation information.

Evaluation of a calibration transfer between a bench top and portable Mid-InfraRed spectrometer for cocaine classification and quantification.

A portable Fourier Transform Mid-InfraRed (FT-MIR) spectrometer using Attenuated Total Reflectance (ATR) sampling is used for daily routine screening of seized powders. Earlier, ATR-FT-MIR combined with Support Vector Machines (SVM) algorithms resulted in a significant improvement of the screening method to a reliable and straightforward classification and quantification tool for both cocaine and levamisole. However, can this tool be transferred to new (hand-held) devices, without loss of the extensive data set? The objective of this study was to perform a calibration transfer between a newly purchased bench top (BT) spectrometer and a portable (P) spectrometer with existing calibration models. Both instruments are from the same brand and have identical characteristics and acquisition parameters (FT instrument, resolution of 4 cm-1 and wavenumber range 4000 to 500 cm-1). The original SVM classification model (n = 515) and SVM quantification model (n = 378) were considered for the transfer trial. Three calibration transfer strategies were assessed: 1) adjustment of slope and bias; 2) correction of spectra from the new instrument BT to P using Piecewise Direct Standardization (PDS) and 3) building a new mixed instrument model with spectra of both instruments. For each approach, additional cocaine powders were measured (n = 682) and the results were compared with GC-MS and GC-FID. The development of a mixed instrument model was the most successful in terms of performance. The future strategy of a mixed model allows applying the models, developed in the laboratory, to portable instruments that are used on-site, and vice versa. The approach offers opportunities to exchange data within a network of forensic laboratories using other FT-MIR spectrometers.

Practical tool for sampling and fast analysis of large cocaine seizures.

Large quantities of illicit drugs are frequently seized by law enforcement. In such cases, a representative number of samples needs to be quickly examined prior to destruction. No procedure has yet been set up which rapidly provides information regarding the homogeneity of the samples, the presence of controlled substances, and the degree of purity. This study establishes a protocol for fast analysis of cocaine and its most common cutting agent, levamisole, in large seizures. The protocol is based on a hypergeometric sampling approach combined with Fourier-transform infrared (FTIR) spectrometry and support vector machines (SVM) algorithms as analysis methods. To demonstrate the practical use of this approach, 5 large cocaine seizures (consisting between 45 and 85 units) were analysed simultaneously with gas chromatography-mass spectrometry (GC-MS), gas chromatography-flame ionisation detector (GC-FID), and a portable FTIR spectrometer using attenuated total reflectance (ATR) sampling combined with SVM models. According to the hypergeometric sampling plan of the guidelines of the Drugs Working Group (DWG) of the European Network of Forensic Science Institutes (ENFSI), the required number of subsamples ranged between 19 and 23. Considering the identification analyses, the SVM models detected cocaine and levamisole in all subsamples of Cases 1 to 5 (100% correct classification), which was confirmed by GC-MS analysis. Considering the quantification analyses, the SVM models were able to estimate the cocaine and levamisole content in each subsample, compared to GC-FID data. The developed strategy is easy, cost effective, and provides immediate information about both the presence and concentration of cocaine and levamisole. By using this new strategy, the number of confirmation analyses with laborious and expensive chromatographic techniques could be significantly reduced.

Prevalence of new psychoactive substances and prescription drugs in the Belgian driving under the influence of drugs population.

Driving under the influence of drugs (DUID) is a worldwide problem. Several countries have adopted DUID legislations which prove their deterrent effect and impact on road safety. However, the use of new psychoactive substances (NPS) and prescription drugs is not known, as the applied roadside screening tests have not yet been adapted for these compounds. In this study, 558 blood samples obtained during roadside controls in Belgium (January to August 2015) after a positive Drugwipe 5S® test and 199 oral fluid (OF) samples obtained from negatively screened test pads were analyzed. The NPS positivity rate was 7% in blood, while it reached 11% in OF. NPS detected were: diphenidine, ketamine, 4-fluoroamphetamine, 2-amino-indane, methoxetamine, α-PVP, methiopropamine, a mix of 5-MAPB/5-EAPB, TH-PVP, mephedrone, methedrone, 4-methylethylcathinone, 5-MeO-DALT, 4-Acetoxy-DiPT, AB Fubinaca, FUB-JWH018, JWH020, trifluoromethylphenylpiperazine, and ethylphenidate. Moreover, 17% of blood samples (and 5% of OF) contained an analgesic drug, 10% (0.5%) a benzodiazepine/hypnotic, 5% (2%) an antidepressant, 2% (3%) an antipsychotic, 2% an antiepileptic drug, and 1% methylphenidate. The presence of NPS in the young (and predominately male) DUID population is proven. Furthermore, a high level of poly-drug use including combinations of NPS, licit, and drugs of abuse was observed. Further research concerning the development of on-site NPS detection techniques should be established. Meanwhile, the effects of combined drug use on driving ability and the physical/psychological signs after NPS use should be performed to improve the on-site DUID detection of NPS by police officers, so they can engage in blood sampling for a general unknown screening.

Roadside drug testing: comparison of two legal approaches in Belgium.

BACKGROUND: Internationally, urine on-site testing has been used for detecting drivers under the influence of drugs (DUID) but more and more countries, such as Belgium, are switching to oral fluid screening. OBJECTIVE: To compare the previous (published in 1999) and current (published 2009) enforcement procedures of DUID in Belgium. The two evaluated procedures differ in the way the drivers are screened by the police (signs of impairment versus signs of recent drug use), the matrix for screening (urine versus oral fluid) and the analytical cut-off concentrations in plasma. METHODS: Data on positive screening and confirmation results were gathered from 1st April 2008 to 30th September 2010, when urine screening (Dipro Druglab panels test) was performed; and from 1st October 2010 to 31st March 2013, when an on-site oral fluid test (Securetec Drugwipe 5(+)) was used. RESULTS: Approximately 4100 data sets related to urine screening and 3900 data sets related to oral fluid screening were studied. Eighty-eight percent of positive urine on-site tests yielded positive results in plasma for cannabis, 21% for cocaine, 20% for amphetamines and 7% for opiates. Sixty-six percent of the positive oral fluid on-site tests yielded positive results in plasma for cannabis, 30% for cocaine, 28% for amphetamines and 8% for opiates. For cannabis, opiates and amphetamines more negative results in plasma were observed in the period of urine screening. CONCLUSIONS: The percentage of plasma samples of tested drivers, in which none of the positive screened target drugs were present in a concentration above the legal cut-off value, has decreased from 17% to 8% since the introduction of the current legislation involving oral fluid screening.

Susac-like syndrome in a chronic cocaine abuser: could levamisole play a role?

INTRODUCTION: Toxic leukoencephalopathy is a possible but rare complication of chronic cocaine abuse. The role of adulterants, mainly levamisole, is still debated. CASE REPORT: We describe an atypical case of fatal leukoencephalopathy mimicking Susac syndrome in a 22-year-old man who was chronically abusing cannabis and cocaine. Exposure to levamisole as adulterant to cocaine was proven by hair analysis. Despite cessation of exposure to cocaine and aggressive immunosuppressive therapy, the patient remained in a minimally conscious state until death. DISCUSSION: Susac syndrome is a rare entity, and its etiology is not yet fully elucidated. The toxic etiologies have been poorly investigated to date. Further observations are required to determine if cocaine and/or adulterants might play a significant role.

Bio-analysis of forensically relevant drugs in alternative matrices by liquid chromatography-tandem mass spectrometry.

Cannabis is the most frequently detected illicit drug in the Western world, e.g. in cases of driving under the influence of drugs (DUID), whereas benzodiazepines comprise the most abused licit drugs and have been linked with drug-facilitated sexual assault cases (DFSA). In recent years, remarkable advances in sensitive analytical techniques have enabled the analysis of drugs in alternative matrices such as oral fluid and hair. These specimens allow easy, non-invasive sampling, which can be achieved under close supervision to prevent adulteration or substitution of the samples. The volume is often limited and to achieve the required analytical sensitivity, liquid chromatography-tandem mass spectrometry (LC-MS-MS) methods for the detection of cannabis and benzodiazepines in oral fluid and hair were developed. After validation, these methods were applied to genuine samples to assess: (a) the validity of oral fluid to detect recent cannabis consumption, (b) the Dräger Drug Test as an on-site oral fluid test, and (c) the applicability of hair testing in forensic cases. The latter led to new insights into metabolic conversions between benzodiazepines; this knowledge may avoid potentially erratic conclusions regarding DFSA. Finally, benzodiazepines are also frequently encountered in post-mortem cases. An LCMS-MS method to detect benzodiazepines in larvae and puparia of insects rearing on corpses was developed and validated. In conclusion, this research aimed at combining the usefulness of alternative matrices with the analytical power of LC-MS-MS. Final outcome is a number of sensitive and validated methods ready for use in routine analysis.

Acute effects of nocturnal doses of MDMA on measures of impulsivity and psychomotor performance throughout the night.

INTRODUCTION: Previous studies on the acute effects of MDMA on psychomotor performance and impulsivity showed that MDMA acts as a stimulant. These studies assessed performance during daytime, whereas in real life, dance-attendees leaving a party use the drug during the night. OBJECTIVES: The present study aimed to assess the effects of nocturnal doses of MDMA on psychomotor performance and impulsivity during the night and after a night of sleep deprivation. MATERIALS AND METHODS: Fourteen healthy subjects participated in a double-blind, placebo-controlled, two-way within-subject study. The treatment was MDMA (75 and 50 mg) divided over the evening or double placebo. Psychomotor and impulsivity tasks were conducted four times throughout the evening and night. A vigilance test was conducted once, at 5 A.M.,: and a sleepiness scale was presented to the subjects ten times throughout the evening and night. RESULTS: MDMA impaired tracking performance in a simple tracking task. Divided attention task performance was also impaired as indicated by a decrease in secondary task performance under the influence of MDMA compared with placebo. MDMA did not affect impulsivity measures. Vigilance performance decreased as a function of time on task, but this decrement was less during MDMA treatment compared to placebo. After the administration of MDMA, the sleepiness scale scores were lower during the night when compared with placebo. This difference between MDMA and placebo disappeared in the morning. CONCLUSION: It is concluded that nocturnal doses of MDMA may produce impairments of tracking performance and divided attention throughout the night that are additive to performance impairment produced by sleep loss.

Stimulant effects of 3,4-methylenedioxymethamphetamine (MDMA) 75 mg and methylphenidate 20 mg on actual driving during intoxication and withdrawal.

BACKGROUND: 3,4-methylenedioxymethamphetamine (MDMA) is currently one of the most popular drugs of abuse in Europe. Its increasing use over the last decade has led to concern regarding possible adverse effects on driving. The aims of the present study were to investigate the acute effects of MDMA on actual driving performance during the intoxication and withdrawal phase. METHODS: Eighteen recreational MDMA-users (nine males, nine females) aged 21-39 years participated in a double-blind, placebo-controlled, three-way cross-over study. MDMA 75 mg, methylphenidate 20 mg and placebo were administered on day 1 of treatment (intoxication phase). Driving tests were conducted between 3 and 5 hours post-drug. Subjects returned the following day for a repetition of the driving tests between 27 and 29 hours post-drug (withdrawal phase). On-the-road driving tests consisted of a road-tracking test and a car-following test. Its main parameters were standard deviation of lateral position (SDLP), time to speed adaptation (TSA), brake reaction time (BRT) and gain. FINDINGS: MDMA and methylphenidate significantly decreased SDLP in the road-tracking tests by about 2 cm relative to placebo on day 1 (intoxication phase). In addition, MDMA intoxication decreased performance in the car-following test as indicated by a significant rise in the 'overshoot' of the subjects' response to speed decelerations of the leading vehicle. Driving performance was not affected by treatments during withdrawal on day 2. CONCLUSION: Collectively, these data indicate that MDMA is a stimulant drug that may improve certain aspects of the driving task, such as road-tracking performance, but may reduce performance in other aspects of the driving task, such as accuracy of speed adaptation during car-following performance.

MDMA and alcohol effects, combined and alone, on objective and subjective measures of actual driving performance and psychomotor function.

RATIONALE: The party drug ecstasy is frequently used in combination with other drugs like marihuana and alcohol. In addition, a substantial proportion of the MDMA users has claimed to drive a car when under the influence of MDMA and/or other drugs. OBJECTIVE: To assess the effects of MDMA and alcohol, combined and alone, on actual driving performance and laboratory tasks related to driving. METHODS: Eighteen healthy subjects participated in a double-blind, placebo-controlled, six-way cross-over study. Treatments consisted of MDMA 0, 75, and 100 mg with and without alcohol, aiming at 0.06 mg/ml BAC. Laboratory tests (critical tracking task, object movement estimation task) were conducted between 1.5 and 2 h postdrug (0.5 and 1 h postalcohol). Actual driving tests (road tracking test, car-following test) were conducted between 3 and 5 h postdrug (2 and 4 h postalcohol). Subjects completed the addiction research center inventory (ARCI) and rated their driving quality and mental effort during driving. RESULTS: Alcohol alone impaired critical tracking performance, as well as a number of actual driving performance parameters [i.e., standard deviation of lateral position (SDLP), brake reaction time, and coherence]. MDMA alone reduced SDLP and standard deviation of speed. MDMA significantly moderated alcohol induced impairment of road tracking performance but did not affect alcohol impairments of car-following and laboratory task performance. Subjective data seemed to support objective data. CONCLUSION: MDMA moderated the impairing effects of a low dose of alcohol on road tracking performance but it could not overcome alcohol-induced impairment on other aspects of driving behavior or driving related performance.

Concentrations and ratios of amphetamine, methamphetamine, MDA, MDMA, and MDEA enantiomers determined in plasma samples from clinical toxicology and driving under the influence of drugs cases by GC-NICI-MS.

Enantiomers of amphetamine (AM), methamphetamine (MA), 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxymethamphetamine (MDMA), and 3,4-methylenedioxyethylamphetamine (MDEA) exhibit different pharmacological properties. This may be important for the interpretation of analytical results. Plasma samples were analyzed using validated negative ion chemical ionization gas chromatography-mass spectrometry procedures. The results for clinical toxicology cases, divided into screening (SCR) and intoxication (ITX) cases, and those of driving under the influence of drugs (DUID) cases were compared. The concentrations of all enantiomers, except R-(-)-MDA and R-(-)- and S-(+)-MA, in the SCR samples were lower than in ITX and DUID samples. Differences between concentrations in ITX and DUID samples were only significant for both enantiomers of AM (DUID higher). These findings suggested impairment in drugged drivers. Different enantiomer ratios (R vs. S) were found for AM between DUID and SCR samples, for MDMA between ITX and SCR samples, and for MDA between DUID and ITX and DUID and SCR samples. Higher MDMA enantiomer ratios in SCR compared to ITX samples are in accordance with a previously described increase of those ratios over time, possibly allowing differentiation of recent from nonrecent ingestion. Pharmacokinetic analysis of a MDMA poisoning yielded elimination half-lives of 6.0 h for R-(-)-MDMA and 4.1 h for S-(+)-MDMA. The enantiomer ratios rose exponentially over time.

Development of a rapid and sensitive method for the quantitation of benzodiazepines in Calliphora vicina larvae and puparia by LC-MS-MS.

Liquid chromatography-tandem mass spectrometry (LC-MS-MS) is emerging as the tool of choice for rapid analysis and the detection of biologically active compounds in complex mixtures. We describe the development of a sensitive method for the simultaneous quantitation of 10 benzodiazepines in Calliphora vicina (Diptera: Calliphoridae) larvae and puparia. The use of larvae for toxicological analyses offers some technical advantages over putrefied tissue. Four sample pretreatment methods for isolating the benzodiazepines out of larvae were evaluated. A simple homogenization, followed by acetonitrile precipitation yielded the highest recoveries. Puparia were pulverized and extracted by ultrasonification in methanol. All extracts were subsequently analyzed using reversed-phase LC-MS-MS. Larvae and puparia calibrators containing benzodiazepines at concentrations ranging from 25 to 750 pg/mg and 50 to 500 pg/mg, respectively, were prepared and analyzed. The method was demonstrated to be linear over the ranges investigated. Limits of detection were from 1.88 to 5.13 pg/mg larva and from 6.28 to 19.03 pg/mg puparium. The developed method was applied to the determination of nordiazepam and its metabolite oxazepam in larvae and puparia of the Calliphora vicina fly that had been reared on artificial foodstuff (beef heart) spiked with 1 microg/g nordiazepam. The larvae were harvested at day 5 for analysis of drug content. The method was sufficiently sensitive to allow the detection of nordiazepam and oxazepam in a single larva or puparium.

Development of a rapid and sensitive method for the quantitation of amphetamines in human plasma and oral fluid by LC-MS-MS.

Target analysis of amphetamines in biological samples is of great importance for clinical and forensic toxicologists alike. At present, most laboratories analyze such samples by gas chromatography-mass spectrometry. However, this procedure is labor-intensive and time-consuming, particularly as a preliminary extraction and derivatization are usually unavoidable. Here we describe the development of an alternative method. Amphetamines were isolated from human plasma and oral fluid using a simple methanol precipitation step and subsequently analyzed using reversed-phase liquid chromatography-tandem mass spectrometry. Quantitation of the drugs was performed using multiple reaction monitoring. The developed method, which requires only 50 microL of biological sample, has a total analysis time of less than 20 min (including sample preparation) and enables the simultaneous quantitation of 3,4-methylenedioxymethamphetamine, 3,4-methylenedioxyamphetamine, 3,4-methylenedioxyethylamphetamine, amphetamine, methamphetamine, and ephedrine in a single chromatographic run. Limits of detection of 2 microg/L or better were obtained. The method has been validated and subsequently applied to the analysis of plasma and oral fluid samples collected from current drug users.

Dissociable effects of a single dose of ecstasy (MDMA) on psychomotor skills and attentional performance.

Ecstasy (3,4-methylenedioxymethamphetamine, MDMA) is a psychoactive recreational drug widely used by young people visiting dance parties, and has been associated with poor cognitive function. The current study assessed the influence of a single dose of MDMA 75 mg and alcohol 0.5 g/kg on cognition, psychomotor performance and driving-related task performance. Twelve healthy recreational ecstasy users participated in an experimental study conducted according to a double-blind, double-dummy, placebo-controlled three-way cross-over design. MDMA improved psychomotor performance, such as movement speed and tracking performance in a single task, as well as in a divided attention task. MDMA impaired the ability to predict object movement under divided attention. However, the inability to accurately predict object movement after MDMA may indicate impairment of particular performance skills relevant to driving. There was no effect of MDMA on visual search, planning or retrieval from semantic memory.

On-site testing of saliva and sweat with Drugwipe and determination of concentrations of drugs of abuse in saliva, plasma and urine of suspected users.

Potential drug users participated voluntarily in a Belgian study on the usefulness of the non-instrumental immunoassay Drugwipe (Securetec, Germany) for the screening of cocaine, opiates, amphetamine and cannabinoids in saliva and sweat. If one of the screening assays (urine, oral fluid, sweat) showed a positive result, blood and saliva were collected. The on-site Drugwipe results were correlated with the Drugwipe results for saliva in the laboratory and with the GC/MS results of the corresponding saliva, plasma and urine samples and pharmacological effects at the time of sampling. The Drugwipe assay proved to be sufficiently sensitive for the detection of recent cocaine (n = 6) and amphetamine (n = 15) abuse, whether the device was wiped on the tongue or on the surface of the body, or when a saliva sample was applied to the wiping part. In five of the six potential cocaine users, the saliva concentrations of cocaine exceeded 1,000 ng/ml. In the amphetamine group, the saliva concentrations of amphetamine, MDMA or both were high (> 1,000 ng/ml) in 13 subjects. For cocaine and amphetamine, the positive scores for Drugwipe matched the GC/MS results for the three body fluids. Recent heroin abuse (n = 5) could be demonstrated to some extent with Drugwipe on samples from the tongue but only the two subjects with the highest saliva concentrations of MAM (> 500 ng/ml) and morphine (> 500 ng/ml) were positive. If the legal cut-off value for driving under the influence of opiates in Belgium (20 ng/ml of free morphine in plasma) was taken into account, only three subjects would have been legally positive. For cannabinoids (n = 15), false negatives and even some false positives were observed. Saliva can be considered as a useful analytical matrix for the detection of drugs of abuse after recent abuse when analysed with GC/MS.

Determination of "Ecstasy" components in alternative biological specimens.

This paper reviews procedures for the determination of methylenedioxyamphetamine derivatives, MDA, MDMA, MDEA and MBDB in saliva, sweat and hair. For this topic, the international literature appears very poor, particularly for saliva and sweat. MDMA was first reported in hair in 1993. All but one of the reviewed papers reported detection with GC-MS. No references seem to be available for both meconium and vitreous humor. As it has been already reported in these biological specimens, the parent drug is detected in higher concentrations than its metabolites. The main data on sample preparation, work-up, GC column, derivatization and analytical determination are listed. Several references, taken from the forensic practice are used to document the cases. Some new findings, based on the experience of the author, are also added. Some references, dealing with amphetamine and methamphetamine in alternative specimens are listed in the manuscript to give an overview on the stimulants detection.

Analysis of Drugs in Saliva.

Saliva is presented as an alternative matrix in the establishment of drug abuse. The ultimate salivary concentration is determined by the route of administration, the salivary pH, the degree of plasma protein binding, and the physico-chemical properties of the abused drug. Since the saliva/plasma ratio can exceed 1, saliva might be a better analytical tool than blood during roadside testing of potentially intoxicated drivers. Moreover, saliva can be obtained non-invasively and under supervision. Although drugs of abuse have been determined in saliva for more than a decade, the use of saliva drug testing for forensic purposes is still limited. Several problems have been demonstrated: (a) differences in the collection protocol produce variable results and often, e.g., during roadside testing, only very small volumes of saliva are obtained; (b) the salivary concentrations are much lower than in urine; (c) saliva principally contains the parent drug and until now, no suitable immunoassays have been commercialized. Although salivary drug concentrations are well correlated with pharmacological effects for some drugs, e.g., cocaine, further studies have to prove whether saliva is a suitable matrix to demonstrate "driving under the influence" of psychoactive drugs. Furthermore, an on-site screening assay for drugs of abuse in saliva and the establishment of appropriate cutoff levels should facilitate the use of saliva during roadside testing.

Hepatotoxicity of N, N-dimethylformamide (DMF) in acute poisoning with the veterinary euthanasia drug T-61.

1. We report on a patient who was resuscitated after a suicide attempt with the veterinary euthanasia product T-61 and treated with N-acetylcysteine (NAC) to prevent hepatotoxicity from N,N-dimethylformamide (DMF), the solvent of T-61. 2. Serum concentrations of DMF were high as compared with values published on occupational exposure. 3. The patient showed only a transient increase in liver enzymes with eventually a full recovery. 4. The hepatoprotective effect of NAC was studied in a rat model using the rise in serum sorbitol dehydrogenase (SDH) as a marker for DMF-induced hepatotoxicity. 5. Four series of randomized, controlled and double-blind experiments were carried out and consistently showed a lower increase in SDH in NAC-treated animals in each series. The difference was statistically significant only when the data of the 4 series were pooled. This is probably due to the large interindividual variations in the effect of DMF. 6. We hypothesize that in the rat NAC may have a protective effect. Whether NAC is also protective in patients, in which it is administered after exposure to DMF, cannot be concluded from the present experiments.

Transferability of lipase titrimetric assays: deductions from an interlaboratory study.

Following the selection of the most appropriate method for emulsification and the optimization of the reaction medium, interlaboratory studies were conducted to check the effect of preparing substrates and measuring the catalytic concentration of lipase at different sites as well as the effect of transport on emulsion. The determinations of lipase activity in an abnormal chemistry control against emulsions prepared by two laboratories (and used by both laboratories) and, also, against five separate emulsions prepared by one laboratory (and used by five different laboratories) resulted in average enzyme activity values (2234 +/- 125 and 2263 +/- 204 U/l respectively) which are not statistically different. Standard preparations of lipase, control sera and reference materials can therefore be titrated according to the procedure followed by at least two laboratories for at least 3 days against two separate emulsions.