Frank Musshoff

Concentrations of Tramadol and O‐desmethyltramadol Enantiomers in Different CYP2D6 Genotypes

The influence of CYP2D6 genotype and CYP2D6 inhibitors on enantiomeric plasma levels of tramadol and O‐desmethyltramadol as well as response to tramadol was investigated. One hundred and seventy‐four patients received one hundred intravenous tramadol 3 mg/kg for postoperative analgesia. Blood samples drawn 30, 90, and 180 min after administration were analyzed for plasma concentrations of the enantiomers (+)‐, (−)tramadol and (+)‐, (−)O‐desmethyltramadol by liquid chromatography‐tandem mass spectrometry. Different CYP2D6 genotypes displaying zero (poor metabolizer (PM)), one (heterozygous individual (HZ)/intermediate metabolizer (IM)), two extensive metabolizer (EM), and three (ultra rapid metabolizer (UM)) active genes were compared. Concentrations of O‐desmethyltramadol differed in the four genotype groups. Median (1/3 quartile) area under the concentration–time curves for (+)O‐desmethyltramadol were 0 (0/11.4), 38.6 (15.9/75.3), 66.5 (17.1/118.4), and 149.7 (35.4/235.4) ng·h/ml for PMs, HZ/IMs, EMs, and UMs (P<0.001). Comedication with CYP2D6 inhibitors decreased (+) O‐desmethyltramadol concentrations (P<0.01). In PMs, non‐response rates to tramadol treatment increased fourfold compared with the other genotypes (P<0.001). In conclusion, CYP2D6 genotype determined concentrations of O‐desmethyltramadol enantiomers and influenced efficacy of tramadol treatment.

Automated headspace solid-phase dynamic extraction for the determination of amphetamines and synthetic designer drugs in hair samples

The technique of automated headspace solid-phase dynamic extraction (SPDE) coupled with gas chromatography-mass spectrometry was evaluated for the determination of amphetamines and synthetic designer drugs in hair samples. Headspace SPDE is a novel method for the solventless extraction of organic compounds in aqueous samples. In a so-called inside needle capillary absorption trap a hollow needle with an internal coating of polydimethylsiloxane is used as extraction and preconcentration medium. Sampling is performed on the solution headspace by passing the gas through the device actively by a syringe. Analytes present in the sample are sorbed onto the deposited stationary phase. The syringe needle is placed into the injection port of a GC and rapid heating of the metal needle induces the desorption of analytes. For the determination of amphetamine, methamphetamine, 3,4-methylendioxyamphetamine (MDA), 3,4-methylendioxymethamphetamine, 3,4-methylendioxyethylamphetamine (MDEA), 3,4-methylendioxyphenyl-2-butanamine and N-methyl-1-(3,4-methylendioxyphenyl)-2-butanamine in human hair samples, 10 mg of hair were hydrolysed with sodium hydroxide. After absorption of analytes for an on-coating derivatization procedure the SPDE needle was directly placed into the headspace of a second vial containing N-methyl-bis(trifluoroacetamide). A validation procedure revealed absolute analyte recoveries between 10.2 and 16.7%. Linearity was obtained from 0.1 to 20 ng/mg with coefficients of correlation between 0.992 and 0.999. Intra- and inter-day precision were determined at two different concentrations and resulted in ranges between 1.4 and 4.1% (intra-day) and 4.2-14.6% (inter-day). Limits of detection between 0.03 ng/mg (MDA) and 0.19 ng/mg (MDEA) were achieved. Results indicated that SPDE is a rapid and sensitive method for the analysis of biological samples. Compared to solid-phase microextraction we found a higher extraction rate coupled with a faster automated operation.

Automated headspace solid-phase dynamic extraction for the determination of cannabinoids in hair samples.

This article describes a fully automated procedure for detecting cannabinoids in human hair samples. The procedure uses alkaline hydrolysis and headspace solid-phase dynamic extraction (HS-SPDE), followed by on-coating derivatization and gas chromatography-mass spectrometry (GC-MS). SPDE is a further development of solid-phase microextraction (SPME), based on an inside needle capillary absorption trap. It uses a hollow needle with an internal coating of polydimethylsiloxane as extraction and pre-concentration medium. Ten mg of hair were washed with deionised water, petroleum ether and dichloromethane. After adding deuterated internal standards, the sample was hydrolyzed with sodium hydroxide and directly submitted to HS-SPDE. After absorption of analytes for an on-coating derivatization procedure, the SPDE-needle was directly placed into the headspace of a second vial containing N-methyl-N-trimethylsilyl-trifluoroacetamide before GC-MS analysis. The limit of detection was 0.14 ng/mg for Delta(9)-tetrahydrocannabinol, 0.09 ng/mg for cannabidiol, and 0.12ng/mg for cannabinol. Absolute recoveries were in the range of 0.6 to 8.4%. Linearity was verified over a range from 0.2 to 20 ng/mg, with coefficients of correlation between 0.998 and 0.999. Intra- and inter-day precision were determined at two different concentrations and resulted in ranges between 2.3 and 6.0% (intra-day) and 3.3 and 7.6% (inter-day). Compared with conventional methods of hair analysis, this automated HS-SPDE-GC-MS procedure is substantially faster. It is easy to perform without using solvents and with minimal sample quantities, and it yields the same sensitivity and reproducibility. Compared to SPME, we found a higher extraction rate, coupled with a faster automated operation and greater stability of the device.

Driving under the influence of synthetic cannabinoids (“Spice”): a case series

Recreational use of synthetic cannabinoid receptor agonists—so-called “Spice” products—became very popular during the last few years. Several reports on clinical symptoms and poisonings were published. Unfortunately, most of these reports do not contain any analytical data on synthetic cannabinoids in body fluids, and no or only a limited number of cases were reported concerning driving under the influence (DUI) of this kind of drugs. In this article, several cases of DUI of synthetic cannabinoids (AM-2201, JWH-018, JWH-019, JWH-122, JWH-210, JWH-307, MAM-2201 (JWH-122 5-fluoropentyl derivative), and UR-144) are presented, focusing on analytical results and signs of impairment documented by the police or the physicians who had taken the blood sample from the suspects. Consumption of synthetic cannabinoids can lead to impairment similar to typical performance deficits caused by cannabis use which are not compatible with safe driving. These deficits include centrally sedating effects and impairment of fine motor skills necessary for keeping the vehicle on track. Police as well as forensic toxicologists and other groups should become familiar with the effects of synthetic cannabinoid use, and be aware of the fact that drug users may shift to these “legal” alternatives due to their nondetectability by commonly used drug screening tests based on antibodies. Sophisticated screening procedures covering the complete range of available compounds or their metabolites have to be developed for both blood/serum and urine testing.

Post-mortem biochemical investigations of vitreous humor

For several decades vitreous humor has been used for post-mortem biochemical investigations with the objective of a post-mortem diagnosis of pre-existing diseases and the clarification of forensic issues, in particular the determination of the post-mortem interval. For the determination of measured concentrations in vitreous humor pre-analytic factors as well as analytical and instrumental variations have to be taken into consideration. The aim of this study was a methodical investigation of two methods of sample pre-treatment as influencing variables. The compared methods were centrifugation and treatment in the ultrasonic bath. The determined parameters were sodium, potassium, chloride, calcium, lactate, urea, glucose and creatinine. Analyses were performed photometrically or by an ion-selective electrode. For some of the analytes a dilution was necessary before analysing. Regarding to the two pre-treatment methods, significant differences in the measured concentrations were not found. The precision proved to be mostly unsatisfying and was clearly better in diluted samples than in undiluted aliquots. A comparison of the vitreous humor of the two ocular bulbs did not lead to significant differences.

Disorders of glucose metabolism–post mortem analyses in forensic cases: part I

In developed countries, diabetes is one of the ten most common causes of death. Post mortem diagnosis of glucose metabolism disorders can be difficult and vague because of the lack of characteristic morphological findings. Reviews of the literature are presented concerning biochemical problems in cases of unclear hyper- or hypoglycemia. After repetition of causes, frequency, and mortality of diabetic metabolism disorders, we give hints for the detection of diabetic ketoacidosis, hyperosmolar coma, insulinoma, and insulin- or oral diabetic-induced hypoglycemia. The first part discusses the analytes glucose and lactate, glycated proteins and oral antidiabetics, with special regard to their matrices post mortem, to reference concentrations, stability data and to analytic procedures that should be used in clinical or toxicological laboratories to detect diabetic metabolism disorders after death.

An fMRI study on the acute effects of exercise on pain processing in trained athletes.

Summary Endurance exercise modulates affective pain ratings and pain‐evoked responses in distinct areas of the human pain matrix, presumably via central opioidergic modulation. Abstract Endurance exercise is known to promote sustained antinociceptive effects, and there is evidence that the reduction of pain perception mediated by exercise is driven by central opioidergic neurotransmission. To directly investigate the involved brain areas and the underlying neural mechanisms in humans, thermal heat‐pain challenges were applied to 20 athletes during 4 separate functional magnetic resonance imaging (fMRI) scans, i.e., before and after 2 hours of running (exercise condition) and walking (control condition), respectively. Imaging revealed a reproducible pattern of distributed pain‐related activation in all 4 conditions, including the mesial and lateral pain systems, and the periaqueductal gray (PAG) as a key region of the descending antinociceptive pathway. At the behavioral level, running as compared with walking decreased affective pain ratings. The influence of exercise on pain‐related activation was reflected in a significant time × treatment interaction in the PAG, along with similar trends in the pregenual anterior cingulate cortex and the middle insular cortex, where pain‐induced activation levels were elevated after walking, but decreased or unchanged after running. Our findings indicate that enhanced reactive recruitment of endogenous antinociceptive mechanisms after aversive repeated pain exposure is attenuated by exercise. The fact that running, but not walking, reproducibly elevated β‐endorphin levels in plasma indicates involvement of the opioidergic system in exercise. This may argue for an elevated opioidergic tone in the brain of athletes, mediating antinociceptive mechanisms. Our findings provide the first evidence using functional imaging to support the role of endurance exercise in pain modulation.

Disorders of glucose metabolism: post mortem analyses in forensic cases–part II

In continuation to part I, a literature review is presented concerning biochemical problems of forensic post mortem cases of unclear hyperglycaemia or hypoglycaemia. Clinical parameters for this purpose were recently reviewed. Particular attention was paid to the detection of diabetic ketoacidosis, of hyperosmolar coma, insulinoma, insulin-induced or oral diabetic-induced hypoglycaemia. The second part of the review discusses the analytes ketone bodies, synthetic insulins, human insulin, C-peptide, proinsulin and insulin antibodies. Special interest is given to post mortem matrices for those analytes to reference concentrations, stability data, analytic interferences and analytical procedures which should be used in toxicological laboratories willing to detect diabetic metabolism disorders after death.

Molecular pathology in forensic medicine —Introduction

Techniques of molecular biology have improved diagnostic sensitivity, accuracy and validity in forensic medicine very much, especially in the field of identification (paternity testing, stain analysis). Since more than 10 years these techniques - meanwhile well established in clinical disciplines - are used also for other applications in forensic medicine: determination of cause and manner of death, tissue identification by mRNA and microRNA, examination of gene expression levels (survival time, time since death, cause of death), toxicogenetics.

A gas chromatographic analysis of phosphine in biological material in a case of suicide

In a suicide committed using aluminium phosphide (AlP) the liberated toxic phosphine gas was detected in post-mortem specimens using a headspace gas chromatographic procedure with a nitrogen-phosphorous detector (HS-GC/NPD). At autopsy a direct sampling into airtight headspace vials for a later analysis is recommended. AlP has to be considered a potent pesticide and its use and availability should be restricted as much as possible.