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Biomedical subjects

Frank Musshoff

Publications and source records attributed to Frank Musshoff.

At least 19 recordsLinked to original sources
PubMed2006

New trends in hair analysis and scientific demands on validation and technical notes.

This review focuses on basic aspects of method development and validation of hair testing procedures. Quality assurance is a major issue in drug testing in hair resulting in new recommendations, validation procedures and inter-laboratory comparisons. Furthermore recent trends in research concerning hair analysis are discussed, namely mechanisms of drug incorporation and retention, novel analytical procedures (especially ones using liquid chromatography-mass spectrometry (LC-MS) and alternative sample preparation techniques like solid-phase microextraction (SPME)), the determination of THC-COOH in hair samples, hair testing in drug-facilitated crimes, enantioselective hair testing procedures and the importance of hair analysis in clinical trials. Hair testing in analytical toxicology is still an area in need of further research.

Forensic Toxicology↗
PubMed2006

Tasks of research in forensic medicine - different study types in clinical research and forensic medicine.

In the last years the research output of forensic medicine has sometimes been regarded as insufficient and as of poor quality, especially when parameters as impact factors and external funding were taken into account. However, forensic medicine has different tasks compared to clinical medicine. The main difference between basic subjects, clinical and forensic medicine is not a lack of scientific efficiency in forensic medicine but is a result of the questions asked, the available methods and specific aims. In contrast to natural-scientific research, forensic science has furthermore important intersections with arts and socio-scientific disciplines. Etiologic and pathogenetic research is of only limited relevance in forensic medicine. Thus, forensic medicine is excluded from these research fields, which are mainly supported by external funding. In forensic medicine research mainly means applied research regarding findings, the probative value and reconstruction as well as examination at different points of intersection between medicine and law. Clinical types of research such as controlled randomised, prospective cross-sectional, cohort or case-control studies can only rarely be applied in forensic medicine due to the area specific research fields (e.g. thantatology, violent death, vitality, traffic medicine, analytical toxicology, hemogenetics and stain analysis). The types of studies which are successfully established in forensic medicine are comparison of methods, sensitivity studies, validation of methods, kinetic examinations etc. Tasks of research in forensic medicine and study types, which may be applied will be addressed.

Biomedical Research↗
PubMed2006

Validation of new methods.

Reliable analytical data are a prerequisite for correct interpretation of toxicological findings in the evaluation of scientific studies, as well as in daily routine work. Unreliable analytical data might not only be contested in court, but could also lead to unjustified legal consequences for the defendant or to wrong treatment of the patient. Therefore, new analytical methods to be used in forensic and/or clinical toxicology require careful method development and thorough validation of the final method. This is especially true in the context of quality management and accreditation, which have become matters of increasing relevance in analytical toxicology in recent years. In this paper, important considerations in analytical method validation will be discussed which may be used as guidance by scientists wishing to develop and validate analytical methods.

Data Interpretation, Statistical↗
PubMed2006

Postmortem biochemistry.

Postmortem biochemistry may provide significant information in determining the cause of death. Due to the rapid postmortem breakdown of metabolism and active membrane transport only analytes which are stable in blood can be determined on this fluid compartment, other parameters have to be analysed on other fluid compartments like vitreous humor (VH). However, using another fluid compartment as a mirror of blood at the moment of death involves severe methodical problems. The conceptual problems of reference values in vitreous humor as a mirror of blood are addressed. Additionally, the necessary steps to be taken before calculating the discriminating values between "normal" and "diseased" are described. For all chemical determinations, a clear definition of the site of sample acquisition is necessary. Up to now chemical determinations on alternative fluids have mainly been carried out using instruments calibrated for urine or serum. Developing calibrated methods for analysis of alternative fluids is a further task for the future.

Biomarkers↗
PubMed2006

Fatal Candida tropicalis infection in an 8-month-old infant with an aplasia of the thymus as a rare cause of death in infancy.

UNLABELLED: Anomalies of the thymus can lead to immunodeficiencies that are often associated with complex syndromes. The clinical relevance of those anomalies varies a lot. Regarding mortality and morbidity, opportunistic infections play a major role in immunocompromised individuals. CASE REPORT: An 8-month-old infant died of a sepsis with multi-organ failure after 32 days in intensive care. At autopsy, no thymus gland could be found; toxicological examinations yielded high morphine concentrations. Autopsy findings and the results of further examinations performed postmortem are discussed in view of the clinical findings, the classification and the cause of death.

Analgesics, Opioid↗
PubMed2006

Enantiomeric determination of tramadol and O-desmethyltramadol by liquid chromatography- mass spectrometry and application to postoperative patients receiving tramadol.

A liquid chromatographic-mass spectrometric assay with atmospheric pressure chemical ionization is presented for quantification (selected-ion mode) of tramadol (T) and O-desmethyltramadol (ODT) in blood plasma after liquid-liquid extraction. The enantiomeric separation was achieved on a Chiralpak AD column containing amylose tris-(3,5-dimethylphenylcarbamate) as chiral selector. The validation data were within acceptable limits. The assay was successfully applied to authentic plasma samples allowing confirmation of diagnosis of overdose or intoxication as well as monitoring of patients' compliance. In postoperative patients receiving T, mean therapeutic plasma concentrations were determined as follows: 422.8 and 440.8 ng/mL for (+)-T and (-)-T, respectively, and 23.7 and 40.7 ng/mL for (+)-ODT and (-)-ODT, respectively. Quantitative results demonstrated a great interindividual variability in postoperative plasma analgesic drug concentrations and, therefore, a wide therapeutic concentration range. The 7.7% of the samples that revealed negative results for ODT are likely explained by genetic polymorphisms resulting in absent enzyme activity of CYP2D6.

Analgesics, Opioid↗
PubMed2006

The significance of putative urinary markers of illicit heroin use after consumption of poppy seed products.

After consumption of poppy seeds various substances were detected in urine or blood samples using an immunoassay and a sophisticated liquid chromatographic-tandem mass spectrometric procedure. These compounds are widely considered to be putative markers of heroin (HER) abuse whereas acetylcodeine was regarded as a marker for illicit preparations ("street HER"). Besides positive urinary opiate immunoassay results during a 48 hours monitoring period, peak concentrations of morphine (MOR), codeine and their glucuronides appeared 4 to 8 hours after ingestion of poppy seeds, and concentrations of total MOR higher than 10 microg/mL were observed. Also, in serum samples taken up to 6 hours after consumption, MOR glucuronides were found. Free MOR was only detected in traces (1 to 3 ng/mL) within 2 hours of consumption. In addition, 3 of 6 onsite opiate sweat tests revealed positive results 6.5 hours after ingestion. Furthermore, it was demonstrated that neither noscapine (NOS) nor papaverine (PAP) was detectable in urine or blood samples after the consumption of poppy seeds containing up to 94 microg NOS and up to 3.3 mug PAP. NOS and PAP were rapidly metabolized, whereas desmethylpapaverine and, especially, its glucuronide were found in urine samples of poppy seed consumers even 48 hours after consumption. According to these results PAP metabolites should not be regarded as markers of illicit HER abuse. In conclusion, only acetylcodeine can be regarded as a specific marker but has the problem of a short half-life. Therefore, we suggest that NOS and PAP, but not their metabolites, might be used cautiously as additional markers of illicit HER abuse as they have not been detected after oral intake of poppy seeds in normal doses. But it must be kept in mind that in some cases poppy seeds with an unusually high content of these alkaloids could be available, and that these substances are also agents in some pharmaceuticals.

Biomarkers↗
PubMed2006

Review of biologic matrices (urine, blood, hair) as indicators of recent or ongoing cannabis use.

Especially for cannabinoids, analytical procedures for the verification of recent use and generally for the assessment of the extent of drug abuse are of interest in clinical and forensic toxicology. For confirmation of abstinence, urine analysis seems to be a useful tool. Serial monitoring of THC-COOH to creatinine ratios can differentiate between recent drug use and residual THC-COOH excretion (THC-COOH/creatinine ratio > or = 0.5 compared with previous specimen ratio). For an assessment of the extent of cannabis use, the determination of free and bound THC-COOH and especially of THC and 11-OH-THC glucuronides are suggested as useful but need further confirmation. Blood analysis is preferred for the interpretation of acute effects after cannabis abuse. The cannabis influence factor (CIF) was demonstrated as a better tool to interpret the concentrations of THC and its metabolites in blood in forensic cases and therefore it was proposed to assume absolute driving inability because of cannabis intoxication from a CIF > or = 10. Additionally, a higher CIF is indicative of a recent cannabis abuse. Also discrimination between occasional use of cannabis and regular drug consumption is possible by analysis of THC-COOH in blood samples because of the long plasma half-life of THC-COOH and its accumulation in the blood of frequent cannabis consumers. In routine tests, blood samples have to be taken within a prescribed 8-day-period, and a THC-COOH concentration >75 ng/mL is assumed to be associated with regular consumption of cannabis products, whereas plasma THC-COOH concentrations <5 ng/mL are associated with occasional consumption. In contrast to other illicit drugs, hair analysis lacks the sensitivity to act as a detector for cannabinoids. THC and especially the main metabolite THC-COOH have a very low incorporation rate into hair and THC is not highly bound to melanin, resulting in much lower concentrations in hair compared with other drugs. Additionally, THC is present in cannabis smoke and also can be incorporated into the hair only by contamination. For the determination of the main metabolite THC-COOH in the picogram or femtogram per milligram range, which indicates an active consumption, special analytical procedures, such as GC/MS/MS techniques, are required.

Hair↗
PubMed2006

Accidental autoerotic death by volatile substance abuse or nonsexually motivated accidents?

In contrast to typical autoerotic fatalities, when death is due to asphyxia mostly by mechanical compression of the neck, atypical autoerotic accidental deaths (AADs) involve sexual self-stimulation by other means such as electrocution or inhalation of chemical agents. Especially in lethal cases of volatile substance abuse (VSA), a differentiation between suicide or sexually or nonsexually motivated accident is often complicated in practical casework. Considering the small number of AADs involving chemical substance abuse reported in the literature, the number of unreported cases seems to be very high. We report about 5 lethal cases of VSA; analysis was performed using headspace solid-phase microextraction (HS-SPME) and gas chromatography-mass spectrometry (GC/MS). When headspace sampling is not performed at autopsy, the analysis of volatile substances can be very complicated. In 2 cases, an AAD was diagnosed considering findings at the scene, reconstruction of the event, and discussion of the circumstances of the death. These findings demonstrate the importance of VSA in atypical autoerotic asphyxia. Therefore, in cases of suspected lethal inhalational intoxications, as a matter of principle, headspace asservation should be performed at autopsy and an autoerotic motivational background should be taken into consideration for differential diagnosis.

Accidents↗
PubMed2005

Determination of opiates and cocaine in hair using automated enzyme immunoassay screening methodologies followed by gas chromatographic-mass spectrometric (GC-MS) confirmation.

The objective of this study was to develop a two-step strategy for analysis of opiates and cocaine in hair samples involving an immunological screening procedure followed by confirmation of results using gas chromatography-mass spectrometry (GC-MS). A semi-quantitative automated competitive enzyme-linked immunosorbent assay (ELISA) methodology using Oral Fluid Micro-Plate Enzyme Immunoassays (Orasure Technologies, Inc.) was developed and validated. Applicability was proven by analysis of authentic head hair samples from drug users (n=103) and from opiate associated fatalities (n=21). The optimum cutoff values for the ELISA tests were 0.1 ng cocaine-equivalents/mg hair and 0.05 ng morphine-equivalents/mg hair using a 50 mg hair sample. Both ELISA tests had a sensitivity of 100%, the specificity was 66% for cocaine-equivalents and 42% for morphine-equivalents. The intraassay precision was 11% for the cocaine and 3% for the opiates ELISA, while interassay precision was 12% for the cocaine and 4% for the opiates ELISA test. The actual analyte concentrations in the hair samples were determined using GC-MS and were between 0.04 and 5.20 ng/mg for heroin (HER), between 0.04 and 30.01 ng/mg for 6-monoacetylmorphine (MAM), between 0.03 and 11.87 ng/mg for morphine (MOR), between 0.02 and 1.84 ng/mg for codeine (COD), between 0.02 and 2.48 ng/mg for acetylcodeine (AC), between 0.01 and 21.37 ng/mg for cocaine (COC), between 0.03 and 10.51 ng/mg for benzoylecgonine (BE) and between 0.05 and 1.26 ng/mg for cocaethylene (CE). The automated ELISA tests were proven to be valid screening procedures for the detection of cocaine and opiates in hair as confirmed by GC-MS. Screening methods provide rapid and inexpensive automated pre-test procedures to detect drugs in hair or other matrices. For forensic purposes screening therefore represents an ideal complement to routinely applied GC-MS procedures.

Case-Control Studies↗
PubMed2005

Systematic regional study of dopamine, norsalsolinol, and (R/S)-salsolinol levels in human brain areas of alcoholics.

BACKGROUND: Dopamine (DA)-derived tetrahydroisoquinolines (TIQs) are discussed as neurochemical factors of addiction processes in alcoholism. In a prospective study, the regional distribution of DA, (R)-salsolinol (SAL), and (S)-SAL, as well as norsalsolinol (NorSAL) was examined systematically in a large collective of human brain samples obtained by autopsy. METHODS: The material comprises 44 brains of alcoholics and 47 controls with 6 standardized specimens in each case. The analytes were determined after solid-phase extraction and enantioselective derivatization using gas chromatography-mass spectrometry. RESULTS: Levels of DA, (R/S)-SAL, and NorSAL in alcoholics did not differ significantly from those of the control group. A relationship between alcohol consumption and SAL formation could not be proved. Topical differences and no ubiquitous occurrence were encountered. Significant amounts of (R)-SAL and (S)-SAL as well as NorSAL only were found in DA-rich areas of the basal ganglia, whereas in other regions of the brain, no TIQs were detected. Especially in the nucleus caudatus, the concentrations of DA, SAL, and NorSAL decreased significantly with rising age. CONCLUSION: These findings do not support the hypothesis that one of the SAL enantiomers or NorSAL is involved in the genesis of alcoholism. However, they suggest that the concentration of the substrate DA may determine the alkaloid level during in vivo formation. The revealed data can serve as reference for other studies in humans concerning the cause of alcoholism or other neurodegenerative diseases with the involvement of TIQs.

Adult↗
PubMed2004

Sevoflurane inhibits phorbol-myristate-acetate-induced activator protein-1 activation in human T lymphocytes in vitro: potential role of the p38-stress kinase pathway.

BACKGROUND: Modulation of immune defense mechanisms by volatile anesthetics during general anesthesia may compromise postoperative immune competence and healing reactions and affect the infection rate and the rate of tumor metastases disseminated during surgery. Several mechanisms have been suggested to account for these effects. The current study was undertaken to examine the molecular mechanisms underlying these observations. METHODS: Effects of sevoflurane, isoflurane, and desflurane were studied in vitro in primary human CD3 T-lymphocytes. DNA-binding activity of the transcription factor activator protein-1 (AP-1) was assessed using an electrophoretic mobility shift assay. Phorbol-myristate-acetate-dependent effects of sevoflurane on the phosphorylation of the mitogen-activated protein kinases were studied using Western blots, the trans-activating potency of AP-1 was determined using reporter gene assays, and the cytokine release was measured using enzyme-linked immunosorbent assays. RESULTS: Sevoflurane inhibited activation of the transcription factor AP-1. This effect was specific, as the activity of nuclear factor kappabeta, nuclear factor of activated T cells, and specific protein-1 was not altered and several other volatile anesthetics studied did not affect AP-1 activation. Sevoflurane-mediated suppression of AP-1 could be observed in primary CD3 lymphocytes from healthy volunteers, was time-dependent and concentration-dependent, and occurred at concentrations that are clinically achieved. It resulted in an inhibition of AP-1-driven reporter gene activity and of the expression of the AP-1 target gene interleukin-3. Suppression of AP-1 was associated with altered phosphorylation of p38 mitogen-activated protein kinases. CONCLUSION: The data demonstrate that sevoflurane is a specific inhibitor of AP-1 and may thus provide a molecular mechanism for the antiinflammatory effects associated with sevoflurane administration.

Anesthetics, Inhalation↗
PubMed2003

Determination of cannabinoids in hemp food products by use of headspace solid-phase microextraction and gas chromatography-mass spectrometry.

A fully automated procedure using alkaline hydrolysis and headspace solid-phase microextraction (HS-SPME), followed by on-fiber derivatization and gas chromatographic-mass spectrometric (GC-MS) detection has been developed for determination of cannabinoids in hemp food samples. After addition of a deuterated internal standard, the sample was hydrolyzed with sodium hydroxide and submitted to direct HS-SPME. After absorption of analytes for on-fiber derivatization, the fiber was placed directly into the headspace of a second vial containing N-methyl- N-trimethylsilyltrifluoroacetamide (MSTFA), before GC-MS analysis. Linearity was good for Delta(9)-tetrahydrocannabinol (THC), cannabidiol, and cannabinol; regression coefficients were greater than 0.99. Depending on the characteristics of the matrix the detection limits obtained ranged between 0.01 and 0.17 mg kg(-1) and the precision between 0.4 and 11.8%. In comparison with conventional liquid-liquid extraction this automated HS-SPME-GC-MS procedure is substantially faster. It is easy to perform, solvent-free, and sample quantities are minimal, yet it maintains the same sensitivity and reproducibility. The applicability was demonstrated by analysis of 30 hemp food samples. Cannabinoids were detected in all of the samples and it was possible to differentiate between drug-type and fiber-type Cannabis sativa L. In comparison with other studies relatively low THC concentrations between 0.01 and 15.53 mg kg(-1) were determined.

Cannabinoids↗
PubMed2003

Area-specific increased density of mu-opioid receptor immunoreactive neurons in the cerebral cortex of drug-related fatalities.

In animal experiments and in cell culture, chronic morphine treatment has been followed by "up-regulation" as well as "down-regulation" of the mu-opioid receptor (OR) number. The present postmortem morphometric study of morphine-related fatalities of drug-addicts (n=13, 20-35 years old, with blood unconjugated morphine levels from 27.1 ng/ml to 458 ng/ml, m.v. 198.5 ng/ml) versus a non-addicted control group (n=13, 10-44 years old) was intended to examine, whether chronic opiate exposure affects the numerical density of mu-OR expressing neurons in the human neocortex (areas 11, 24 and 25 according to Brodmann). For the immunohistochemical procedure, vibratome sections (100 microm) were incubated with a monoclonal antibody against the mu-OR, diluted 1:100, and immunolabelled sites were visualized using an immunoperoxidase protocol. The numerical densities of OR immunoreactive neuronal profiles and Nissl-stained central profiles were assessed morphometrically (camera lucida-drawings). In both groups, the anti-mu-OR-immunoreactivity was mainly localized in pyramidal neurons of layers (L) II/III and V and in multiform neurons of L VI. In the areas 24 and 25, the density of the immunoreactive neuronal profiles did not display a significant difference between the two examined groups. In the area 11, however, the number of immunolabelled neuronal profiles amounted to 2777+/-206 mm(3) in the drug-related fatalities and to 2320+/-124 mm(3) in the control group and thus was significantly increased.

Adolescent↗
PubMed2003

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.

Cannabinoids↗
PubMed2003

Methadone substitution: medicolegal problems in Germany.

In Germany, the substitution of methadone for heroin abusers has arisen recently and has resulted in various medicolegal problems. Normally, these concern methadone-associated deaths, or the prescription of methadone resulting in criminal prosecutions concerning physicians. Not to be forgotten is the problem of driving while taking methadone. In the years 1997-2001, we detected methadone in the blood of 398 cases that were analysed by the Institute of Legal Medicine, Bonn. Methadone was the only drug in only 18 cases. In most of the cases, up to five additional drugs were also being taken: benzodiazepines (61%), ethanol (40%), morphine (39%), cannabinoids (35%), cocaine (28%), anti-depressants (3%), and amphetamines (2%).

Adolescent↗
PubMed2003

Application of tandem mass spectrometry combined with gas chromatography and headspace solid-phase dynamic extraction for the determination of drugs of abuse in hair samples.

A new method combination, headspace solid-phase dynamic extraction coupled with gas chromatography/tandem mass spectrometry (HS-SPDE/GC/MS/MS), is introduced to determine drugs of abuse in hair samples. This highly automated procedure utilizes SPDE for pre-concentration and on-coating derivatization as well as GC and triple quadrupole MS/MS for selective and sensitive detection. All these steps, apart from washing and cutting of the hair samples, are performed without manual intervention on a robot-like autosampler.SPDE is a solventless extraction technique related to solid-phase microextraction (SPME). The analytes are absorbed from the sample headspace directly into a hollow needle with an internal coating of polydimethylsiloxane by repeated aspirate/dispense cycles.The HS-SPDE/GC/MS/MS procedure was applied to the analysis of methadone, the trimethylsilyl derivatives of cannabinoids and the trifluoroacetyl derivatives of amphetamines and designer drugs. The method was shown to be sensitive with detection limits between 6 and 52 pg/mg hair matrix and precision between 0.4 and 7.8% by the use of an internal standard technique. Linearity was obtained from 0.1-20 ng/mg with coefficients of correlation between 0.995 and 0.999. Compared with conventional methods of hair analysis, HS-SPDE/GC/MS/MS is easier to use, substantially faster, with the degree of sensitivity and reproducibility demanded in clinical and forensic toxicology. The main advantage of the SPDE technique in relation to SPME is the robustness of the capillary.

Gas Chromatography-Mass Spectrometry↗