Cornelius Hess

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.

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.

Driving under the influence of synthetic phenethylamines: a case series

New psychoactive drugs, so-called legal highs, have gained more and more popularity during the last years. One of the most important groups of these legal high substances are the synthetic phenethylamines that share a common phenethylamine moiety. Based on certain structural characteristics, these synthetic phenethylamines can be divided into further subclasses, among which the synthetic cathinones (‘bath salts’) are particularly noteworthy. Synthetic cathinones are characterized by an additional carbonyl group attached at the beta position on the amino alkyl chain. Consumption of synthetic phenethylamines can lead to impairments similar to those observed after the use of, for instance, amphetamine or 3,4-methylenedioxy-N-methylamphetamine (MDMA, ‘ecstasy’). These impairments include diverse neurological and psychological symptoms which can affect a safe driving behaviour. Although several reports on clinical symptoms and poisonings due to these substances have been published, most of these publications do not contain any analytical data. Additionally, there is still a lack of information concerning pharmacological and toxicological effects of these rather new psychoactive substances. In particular, the knowledge of the impact on the ability to drive following consumption of synthetic phenethylamines is relevant for the police as well as for forensic toxicologists. In this publication, several cases of individuals driving under the influence (DUI) of synthetic phenethylamines (4-fluoroamphetamine, mephedrone (4-methylmethcathinone, 4-MMC), 2-DPMP (desoxypipradol), methylenedioxypyrovalerone (MDPV), benzedrone, N-ethylamphetamine (etilamfetamine), 3-methylmethcathinone (3-MMC)) are presented, focusing on analytical results and signs of impairment.

Pharmacological evaluation of synthetic cannabinoids identified as constituents of spice

In recent years, many synthetic cannabinoid (CB) receptor agonists have appeared on the market as constituents of herbal incense mixtures known as “spice”. Contrary to the declared use, they are perorally consumed as a replacement for marijuana to get “high”. In many cases, detailed information on the physicochemical and pharmacological properties of the synthetic compounds found in spice preparations is lacking. We have now evaluated a large series of heterocyclic compounds, 1,3-disubstituted indole and 2-azaindole derivatives known or assumed to be CB1 receptor agonists, many of which have previously been identified in forensic samples. The mainly observed structural variations to circumvent restriction by law were bioisosteric exchanges of functional groups in known CB1 agonists. We analyzed the structure-activity relationships of compounds at human CB1 and CB2 receptors based on affinities obtained in radioligand binding studies, and determined their efficacy in cAMP accumulation assays. Moreover, we investigated the activities of the compounds at the orphan G protein-coupled receptors GPR18 and GPR55 both of which are known to interact with cannabinoids. Most of the investigated compounds behaved as potent full agonists of CB1 and CB2 receptors with affinities in the low nanomolar to subnanomolar concentration range. Some compounds were moderately potent GPR55 antagonists, while none interacted with GPR18. Most derivatives were predicted to cross the blood–brain barrier as determined by bioinformatics tools. These data are useful for assessing synthetic cannabinoids and will be helpful for predicting pharmacological properties of novel compounds that appear on the illicit drug market.

Death due to diabetic ketoacidosis: Induction by the consumption of synthetic cannabinoids?

We present a case study on a man who suffered from diabetic ketoacidosis, probably following consumption of synthetic cannabinoids. In blood from a femoral vein AB-CHMINACA, AB-FUBINACA, AM-2201, 5F-AMB, 5F-APINACA, EAM-2201, JWH-018, JWH-122, MAM-2201, STS135 and THJ 2201 could be detected by LC-MS/MS. Diagnosis of ketoacidosis as cause of death was carried out using biochemical measurements of glucose and lactate concentrations in vitreous humour (sum formula: 463 mg/dl) and cerebrospinal fluid (sum formula: 506 mg/dl), of acetone (163 mg/l in femoral venous blood) and of HbA1c (98 mmol/mol). Death due to hyperglycaemia could have been induced by skipping of insulin doses due to his intoxicated state or by the cannabinoids which were described to be able to produce hyperglycaemia themselves.

Drug facilitated sexual assault with lethal outcome: GHB intoxication in a six-year-old girl.

A very serious case of DFSA (drug facilitated sexual assault) is presented, in which a six-year-old girl died following sedation with γ-hydroxybutyric acid (GHB). She had been sexually abused by a relative. Samples of cardiac blood, bile, vitreous humour, liver, kidney, brain tissues and hair were analysed by a LC-MS/MS method. The following GHB concentrations were determined: cardiac blood: 150 mg/l; bile: 292mg/l; vitreous humour: 58mg/l; liver: 100 mg/kg; kidney: 124.5 mg/kg, brain: 110 mg/kg. Very high GHB levels were found in the proximal part of the hair sample (about 40.9 ng/mg). In distal segments of hair - up to 12 cm distant from the hair scalp - GHB concentrations were higher than the overall found endogenous range of 2-3 ng/mg. Police investigations revealed that the uncle had also administered GHB to the older half-sister. Therefore, a sample of her hair was analysed accordingly, but unremarkable results were obtained. Comparing our toxicological results with police investigations and the offenders statements it can be assumed that the 6-year-old girl had ingested GHB. By exclusion of other causes of death a lethal intoxication with GHB could be confirmed.

Clinical and forensic examinations of glycemic marker 1,5-anhydroglucitol by means of high performance liquid chromatography tandem mass spectrometry.

Postmortem diagnosis of diabetes and a diabetic coma can be difficult because of the lack of characteristic morphological findings. 1,5-Anhydroglucitol (1,5-AG), the 1-deoxy form of glucose, competes with glucose for reabsorption in the kidneys. Therefore, diabetics with a permanent hyperglycemia show significantly lower serum concentrations of 1,5-AG than non-diabetics. A liquid chromatography-mass spectrometric method for the determination of 1,5-AG in serum and postmortem blood was developed and validated according to international guidelines. Linearity was given between 1 μg/ml and 50 μg/ml. Recovery rates ranged between 70.8% and 89.8%, the limit of quantification of the procedure was 0.20 μg/ml, limit of quantification was 0.55 μg/ml. Serum of 199 diabetics and 116 non-diabetics and femoral blood of 31 diabetic and 27 non-diabetic deceased was measured. Average concentrations were significantly (p<0.001) higher in non-diabetics compared to diabetics ante and postmortem. Seven of the diabetics may have died because of a hyperglycemic coma indicated by a sum formula of Traub>450 mg/dl. 1,5-AG average concentrations in these deceased were not significantly different to diabetics which did not die because of a diabetic coma. Concentrations of 1,5-AG give a hint for not well controlled diabetes antemortem and postmortem and can be assumed as an additional and alternative information postmortem to the measurement of HbA1c or fructosamine.

Simultaneous detection of 93 synthetic cannabinoids by liquid chromatography-tandem mass spectrometry and retrospective application to real forensic samples

The Internet is flooded with steadily changing synthetic cannabinoids in `Spice` products. In routine forensic work, it is difficult to keep the analytical methods for the detection of these analytes up to date. We describe a liquid chromatography-tandem mass spectrometry method after liquid-liquid extraction for the detection of 93 synthetic cannabinoids in human serum. The method was validated for selectivity and specificity, matrix effects, and analytical limits (<1 ng/mL for 81 substances) for qualitative analysis. A short quantitative validation regarding linearity and precision data was also conducted. The method was applied to 189 serum samples provided by police authorities. Sixty-four samples (33.8%) were found positive for at least one synthetic cannabinoid, whereby MDMB-CHMICA, AB-CHMINACA, and 5 F-PB-22 were the substances most frequently detected. Consumption of these substances and plasma concentrations are linked to symptoms documented by the police. Six case reports are presented. Copyright

Determination of levamisole, aminorex, and pemoline in plasma by means of liquid chromatography-mass spectrometry and application to a pharmacokinetic study of levamisole

Levamisole is an anti-helminthic drug and gained forensic interest after it was found that it was used as a cocaine adulterant. A liquid chromatography-mass spectrometry (LC-MS) method for the determination of levamisole and its metabolite aminorex in human plasma is described. Selectivity is given; calibration curves were linear within a calibration range of 1 ng/mL-500 ng/mL. Limits of detection and quantification (LODs, LOQs) were 0.85 ng/mL for levamisole and 0.09 ng/mL, and 0.34 ng/mL for aminorex, respectively. Precision data was in accordance with the GTFCh guidelines. The validated method was successfully applied to study the pharmacokinetics of levamisole after administration of 100 mg of levamisole orally. Levamisole could be detected up to 36 h after ingestion in serum, while aminorex never exceeded the LOQ. A one-compartment model best described levamisole pharmacokinetics. The following parameters were calculated: ka = 1.2 [1/h], CL/F = 52 l/h, V/F = 347 l, f (renal) = 0.0005, t ½ = 2.0 h, AUC = 1923 ng/mL*h, cmax = 214 ng/mL, tmax = 1.98 h. Levamisole could be quantified in 42.5% of cocaine--positive plasma samples (2.2 to 224 ng/mL). Aminorex was positive in only 11.3% of the cases; however, it was never found higher than the LOQ. Pemoline, another stimulant detected in horse urine samples after administration of levamisole, was not found either in serum or in urine of this pharmacokinetic study. In post-mortem cases, levamisole and aminorex could be detected in femoral blood and the urine of cocaine users. Pemoline was not detected.

Determination of hypoglycaemia induced by insulin or its synthetic analogues post mortem

The determination of human insulin or its synthetic analogues in post-mortem specimens represents a challenge for forensic toxicologists due to its proven instability in post-mortem blood. We present two cases of an insulin-induced hypoglycaemia. In the first case, ante-mortem material was available for the detection of an injection with human insulin. Human insulin was detected by immunopurification with magnetic beads and liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses at a concentration of 5180 μU/ml. The molar ratio human insulin:C-peptide was 111. The second case describes a suicide by self-injection of Insulin lispro and determination of the drug after pre-extraction with methanol and immunopurification by LC-MS/MS at the injection site, in vitreous humour and organs. Apart from the well-known matrices--femoral blood and urine--the specimen vitreous humour and the injection site promise the best possibilities for a proof of insulin at autopsy. In addition to insulin analyses, the parameters C-peptide, proinsulin, glucose, lactate, and sulfonylureas should be measured in case of suspected fatal hypoglycaemia.