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München, Germany

Thieme U.,Ludwig Maximilians University of Munich | Schelling G.,Ludwig Maximilians University of Munich | Hauer D.,Ludwig Maximilians University of Munich | Greif R.,University of Bern | And 7 more authors.
Drug Testing and Analysis | Year: 2014

The effects of tetrahydrocannabinol (THC) and endogenous cannabinoids (endocannabinoids, ECs) are both mediated by activation of the cannabinoid receptors CB1 and CB2. Exogenous activation of these receptors by THC could therefore alter EC levels. We tested this hypothesis in healthy volunteers (n=25) who received a large intravenous dose of THC (0.10 mg/kg). Effects on the EC system were quantified by serial measurements of plasma ECs after THC administration. Eleven blood samples were drawn during the first 5 h after THC administration and two more samples after 24 and 48 h. THC, its metabolites THC-OH (biologically active) and THC-COOH (non-active), and the ECs anandamide and 2-arachidonoylglycerol (2-AG) were quantified by liquid chromatography-mass spectrometry. EC-plasma levels showed a biphasic response after THC injection reaching maximal values at 30 min. Anandamide increased slightly from 0.58±0.21 ng/ml at baseline to 0.64±0.24 ng/ml (p<0.05) and 2-AG from 7.60±4.30 ng/ml to 9.50±5.90 ng/ml (p<0.05). After reaching maximal concentrations, EC plasma levels decreased markedly to a nadir of 300 min after THC administration (to 0.32±0.15 ng/ml for anandamide and to 5.50±3.01 ng/ml for 2-AG, p<0.05). EC plasma concentrations returned to near baseline levels until 48 h after the experiment. THC (0.76±0.16 ng/ml) and THC-OH (0.36±0.17 ng/ml) were still measurable at 24 h and remained detectible until 48 h after THC administration. Although the underlying mechanism is not clear, high doses of intravenous THC appear to influence endogenous cannabinoid concentrations and presumably EC-signalling. © 2013 John Wiley & Sons, Ltd. Source

Thieme D.,Institute of Doping Analysis | Sachs H.,Forensic Toxicological Center | Uhl M.,Bavarian State Criminal Police Office
Drug Testing and Analysis | Year: 2014

The identification of 11-nor-Δ9-tetrahydrocannabinol-9-carboxylic acid (THCCOOH) in hair represents an exceptional forensic analytical challenge due to low target concentrations in a complex matrix. Several dedicated techniques [gas chromatography - negative chemical ionization- tandem mass spectrometry (GC-NCI-MS/MS) or GC-GC-MS couplings] were specifically introduced into forensic toxicology aiming to a selective and sensitive identification of THCCOOH in hair. The combination of liquid-chromatography (LC) and MS/MS gained an outstanding relevance in forensic toxicology (including the detection of cannabinoids). However, its application to hair matrix is characterized by a lack of specificity which is due to the unspecific decarboxylation as most abundant fragmentation reaction. Therefore, various chemical modifications of the carboxyl and/or phenolic hydroxyl groups were examined to improve the selectivity. The selective methylation of the 9-carboxyl-group proved to be the most efficient derivatization procedure. Hair extracts were redissolved in acetonitrile and after addition of few milligrams of solid sodium carbonate derivatized with 25μL methyl iodide. The resulting THC-9-carboxymethylester was separated by conventional reverse phase LC and selectively detected using negative electrospray ionization by recording the fragmentation reactions 357→325 and 357→297. Resulting limits of quantification were below 100fg/mg. A further significant improvement was achieved by application of the multistage MS3 fragmentation 357→325→297. To verify the validity of this procedure, a systematic quantitative comparison of THCCOOH concentrations in hair with data from a well established GC-NCI-MS/MS technique was performed. Both techniques proved to be in good accordance (R2 =0.647, p=<0.001) and equally suitable for hair testing of THCCOOH. © 2013 John Wiley & Sons, Ltd. Source

Pragst F.,University Medicine Charite | Broecker S.,University Medicine Charite | Hastedt M.,University Medicine Charite | Herre S.,University Medicine Charite | And 3 more authors.
Therapeutic Drug Monitoring | Year: 2013

BACKGROUND:: Children living in homes with drug-addicted parents are in a steady danger of poisoning and may suffer from neglect, maltreatment, and lagging behind in development. Hair analysis could be a suitable way to examine this endangering exposure to drugs. METHODS:: Hair samples from 149 children (aged 1-14 years) living with parents substituted by methadone and/or suspected for abuse of illegal drugs, and from 124 of the parents in a German community were investigated by liquid chromatography-hybrid quadrupole time-of flight mass spectrometry and by headspace solid phase microextraction gas chromatography-mass spectrometry for methadone, heroin, cocaine, amphetamines, ecstasy, cannabinoids and benzodiazepines and their metabolites or degradation products (32 compounds). RESULTS:: From the children's hair, only in 35 samples, no drugs were detected. Cannabinoids were found in 56 samples, in 20 of them as the only drug. In the remaining 95 samples, methadone was identified 35 times with additional use of illegal drugs in 28 cases. Drug use in the children's environment was obvious for heroin in 44 cases, cocaine in 73 cases, amphetamine or ecstasy in 6 cases, and diazepam in 8 cases. The concentrations varied from limit of quantification to 2.16 ng/mg of methadone, 11.1 ng/mg of 6-acetylmorphine, 17.8 ng/mg of cocaine, 3.29 ng/mg of amphetamine, and 0.72 ng/mg of Δ-tetrahydrocannabinol. In general, hair from younger children contained higher concentrations than from their elder siblings. Systemic incorporation of methadone, cocaine, or cannabinoids appeared likely from detection of the nonhydrolytic metabolites 2-ethylidene-1,5-dimethyl-3,3- diphenylpyrrolidine in 11 cases, norcocaine in 16 cases, and 11-nor-9-carboxy-Δ-tetrahydrocannabinol in 9 cases. Within the families, hair samples of children and parents provided often the same drug pattern. External deposition from smoke and by contact with contaminated surfaces or parent's hands and systemic deposition after passive smoking, administration, or oral intake by hand-to-mouth transfer were discussed as alternative incorporation mechanisms into hair. CONCLUSIONS:: Altogether, investigation of children's hair proved to be a useful way to detect endangering drug use in their environment and lead to a more thorough inspection and measures to improve their situation in many of the cases. Copyright © 2013 by Lippincott Williams & Wilkins. Source

Thieme D.,Institute of Doping Analysis | Sachs U.,Albert Ludwigs University of Freiburg | Sachs H.,Forensic Toxicological Center | Moore C.,Immunalysis Corporation
Drug Testing and Analysis | Year: 2015

Formation of picolinic acid esters of hydroxylated drugs or their biotransformation products is a promising tool to improve their mass spectrometric ionization efficiency, alter their fragmentation behaviour and enhance sensitivity and specificity of their detection. The procedure was optimized and tested for the detection of cannabinoids, which proved to be most challenging when dealing with alternative specimens, for example hair and oral fluid. In particular, the detection of the THC metabolites hydroxyl-THC and carboxy-THC requires ultimate sensitivity because of their poor incorporation into hair or saliva. Both biotransformation products are widely accepted as incorporation markers to distinguish drug consumption from passive contamination. The derivatization procedure was carried out by adding a mixture of picolinic acid, 4-(dimethylamino)pyridine and 2-methyl-6-nitrobenzoic anhydride in tetrahydrofuran/triethylamine to the dry extraction residues. Resulting derivatives were found to be very stable and could be reconstituted in aqueous or organic buffers and subsequently analyzed by liquid chromatography-mass spectrometry (LC-MS). Owing to the complex consecutive fragmentation patterns, the application of multistage MS3 proved to be extremely useful for a sensitive identification of doubly picolinated hydroxy-THC in complex matrices. The detection limits - estimated by comparison of corresponding signal-to-noise ratios - increased by a factor of 100 following picolination. All other species examined, like cannabinol, THC, cannabidiol, and carboxy-THC, could also be derivatized exhibiting only moderate sensitivity improvements. The assay was systematically tested using hair samples and exemplarily applied to oral fluid. Concentrations of OH-THC identified in THC-positive hair samples ranged from 0.02 to 0.29pg/mg. © 2014 John Wiley & Sons, Ltd. Source

Suesse S.,DC Drogencheck | Pragst F.,Charite - Medical University of Berlin | Mieczkowski T.,University of South Florida | Selavka C.M.,Trimega Laboratories | And 5 more authors.
Forensic Science International | Year: 2012

This article presents results from 1872 hair samples, which were analyzed for fatty acid ethyl esters (FAEEs) and ethyl glucuronide (EtG). The results were evaluated in the context of self-reported drinking behavior, the use of hair cosmetics, the gender of the sample donors and hair sample length. For comparison, CDT and GGT in serum were available in 477 and 454 cases, respectively. A number of alcohol abstainers or low moderate drinkers and excessive drinkers were selected for assessment of cut-offs for FAEEs in the proximal 6. cm hair segments and for EtG in the proximal 3. cm hair segments. Cut-off values were assessed by ROC analysis. It was found that the cut-offs of 1.0. ng/mg FAEE and 30. pg/mg EtG presently used for excessive drinking lead to a low portion of false positives (4% and 3% respectively) but to a higher portion of false negatives (23% and 25% respectively). Comparison of the mean and medium concentrations in samples without any reported hair cosmetics (N= 1079) and in samples with reported use of hair spray (N= 79) showed an increase by the factor of about two for FAEE but no significant difference for EtG. Mean values of EtG were decreased by 80% in bleached samples (N= 164) and by 63% in dyed samples (N= 96). There was no significant effect of bleaching and dyeing on FAEE. Hair gel and hair wax, oil or grease showed no significant effect on both FAEE and EtG. With respect to gender and investigated hair length ambiguous results were obtained because of major differences in the compared subpopulations of male with higher alcohol consumption and mainly shorter hair, and less drinking female with longer hair. For excessive drinkers FAEEs in the 0-6. cm hair segment and EtG in the 0-3. cm segment decreased with increasing time of reported abstinence before sample collection. These drinkers attain the level of teetotalers only after more than 10 months of abstinence. In comparison to scalp hair, FAEEs recovered from armpit hair and leg hair were lower and from chest hair were higher. EtG in armpit hair was lower and in leg hair higher than in scalp hair. It is concluded that the combined use of FAEE and EtG essentially increases the accuracy of interpretation since both markers complement each other by a different sensitivity to sources of error. © 2011 Elsevier Ireland Ltd. Source

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