Postmortem Measurement of Human Chorionic Gonadotropin in Vitreous Humor and Bile

Abstract: Postmortem human chorionic gonadotrophin (HCG) blood assay can confirm postmortem diagnosis of pregnancy or document situations in which HCG levels are elevated. In some cases, however, blood sampling is not possible at autopsy. In this study, HCG was quantified by enzyme‐linked fluorescent assay (ELFA) in the bile (n = 5), vitreous humor (n = 4), and postmortem blood (n = 4) of five pregnant women. There were no false negatives in the pregnant subjects (n = 5) or false positives in controls (n = 34), enabling this test to be recommended for routine use in forensic contexts in which the detection of elevated HCG levels could be of interest.     

Fatal Overdose of Gamma‐hydroxybutyrate Acid After Ingestion of 1,4‐Butanediol

We report a case of fatal intoxication from 1,4‐butanediol (1,4‐BD), which was ingested by a young and “naïve” gamma‐hydroxybutyrate (GHB) consumer during a party with the co‐ingestion of alcohol, cannabis, and methylene‐dioxy‐methamphetamine. The following drug concentrations were found using gas chromatography coupled with mass spectrometry on autopsy samples and on a cup and a glass found at the scene: 20,350 mg/L (bottle) for 1,4‐BD; 1020 mg/L (femoral blood), 3380 mg/L (cardiac blood), 47,280 mg/L (gastric content), and 570 mg/L (vitreous humor) for GHB. The concentration of GHB is difficult to interpret in forensic cases due to the possibility of an endogenous production of GHB. The variable tolerance of the user may also modify the peri‐ and postmortem GHB concentrations. This case underscores the need to have many different sources of toxicology samples analyzed to avoid the hypothesis of endogenous production of GHB.     

家兔急性胰腺炎死后不同时间眼玻璃体液淀粉酶变化

目的探讨急性胰腺炎死后不同时间眼玻璃体液淀粉酶变化。方法用1%牛胆酸钠诱导家兔急性胰腺炎动物模型建立,于死后不同时间和正常对照组,观察其淀粉酶变化。结果家兔急性胰腺炎死后72h眼玻璃体液淀粉酶含量(x)与死亡时间(y)存在相关关系,并导出其二项式回归方程为y=8.7420 0.7699x-0.0083x2(R2=92.62792,F=14.89734,P=0.001)。可作为推定早期损伤时间的参考指标。结论家兔急性胰腺炎死后不同时间眼玻璃体液淀粉酶改变为法医早期死亡时间推断提供了灵敏客观的实验依据。     

Stability of drugs in stored postmortem femoral blood and vitreous humor

The stability of 46 drugs in postmortem femoral blood stored for one year at -20 degrees C was investigated. The drugs included benzodiazepines, antidepressants, analgetics and hypnotics. For seven drugs we found a significant change in the concentration between the first and second analysis. Five substances; ethanol, desmethylmianserin, 7-amino-nitrazepam, THC and zopiclone showed a decrease in the concentration whereas the concentrations of two substances; ketobemidone and thioridazine increased. However, the changes observed were not of such an order that it would affect the interpretation in normal forensic casework. We also investigated the possible influence of potassium fluoride on the concentrations of the 46 drugs in vitreous humor after storage for one year. For two substances, ethanol and zopiclone, there were significantly lower concentrations in the samples without potassium fluoride. Furthermore, we also studied the correlation between the concentrations in femoral blood and vitreous humor. For 23 substances there was a significant difference between the concentrations in the vitreous humor and femoral blood. Significant correlations between the concentrations in these two specimens were found for 23 substances, indicating that vitreous humor can be an alternative specimen when blood samples are not available, provided that such correlation exists for the particular substance. Statistical analysis also revealed a correlation between the degree of protein binding of the different drugs and percentage of vitreous/femoral blood concentrations.     

Postmortem biochemical examination of synovial fluid--a preliminary study

Vitreous humor chemistry is used for postmortem analysis since serum values of many components are thought to be reflected in vitreous humor and to be stable for a prolonged postmortem interval (pmi). A similar isolated compartment to vitreous humor is synovial fluid which up to now was hardly used for postmortem chemistry.The aim of the present examination was to compare the values of various analytes in both fluid compartments to get first hints for the reliability of the examination of synovial fluid. Therefore, in 74 cases of sudden death both fluids were taken and analysed for natrium, potassium, calcium, chloride, urea, creatinine, and glucose. The results show that the examination of synovial fluid as compared to vitreous humor is a reliable method to get hints on the premortal metabolic status since most analytes are stable postmortem. The range of all values is comparable in both fluids. The time course of glucose concentrations and, much more important, potassium concentrations is nearly similar. If no vitreous humor is available synovial fluid may be used with all precautions in diagnosis known from vitreous humor.     

Glucose and lactate in vitreous humor compared with the determination of fructosamine for the postmortem diagnosis of diabetes mellitus

Diabetes mellitus is a chronic metabolic illness responsible for a great number of deaths. In postmortem diagnosis, because of the difficulty involved in interpreting blood glucose levels and relatively nonspecific pathologic features, biochemical markers in vitreous humor are useful. The aim of this study was to compare the results obtained for the combined determination of lactate and glucose with fructosamine levels recorded in the vitreous humor of two diagnostic groups (one diabetic and the other nondiabetic). The authors intended to ascertain the capacity of different markers measured in vitreous humor to diagnose diabetes mellitus. Fifty-one cadavers (mean age, 58.7 years; standard deviation, 17.09) were studied. The mean postmortem interval was 16.4 hours (standard deviation, 9.05). Cases were assigned to two diagnostic groups according to whether they were previously diagnosed as either diabetic or nondiabetic. Statistically significant differences for glucose, fructosamine, and the sum values of glucose and lactate were found between the two diagnostic groups. The highest levels were obtained in the group of cases with a previous diagnosis of diabetes mellitus. After the comparison of receiver operating characteristic curves, the sum values of glucose and lactate in vitreous humor is a better predictor of antemortem diabetes mellitus than the fructosamine.     

Vitreous humor carbohydrate-deficient transferrin concentrations in the postmortem diagnosis of alcoholism

Deaths from the effects of alcohol intoxication are encountered routinely in forensic practice. In an important number of cases difficulty may arise in interpreting the significance of results obtained in the autopsy. In clinical practice biochemical markers, particularly serum gamma-glutamyl-transpeptidase (GGT), alanine aminotransferase (ALT), aspartate transaminase (AST), carbohydrate-deficient transferrin (CDT), and erythrocyte mean corpuscular volume are used to diagnose heavy alcohol consumption. CDT is used as a reliable and specific marker. In postmortem diagnosis, because of the difficulty in interpreting blood alcohol levels and relatively non-specific pathological features, biochemical compounds have been studied for use as possible markers. The aim of this study was to evaluate the usefulness of the postmortem determination of CDT in vitreous humor as a confirmation of antemortem alcoholism. CDT levels were studied in 66 male cadavers with a mean age of 55.9 years (S.D. 17.0, range 22-87 years) with a mean postmortem interval of 17.9 h (S.D. 11.4, range 4-72 h). Cases were assigned to two diagnostic groups according to the antemortem diagnosis of alcoholism. Statistically significant differences were found for CDT and ALT concentrations between the two diagnostic groups. The highest vitreous humor levels of CDT and ALT were obtained in the group of cases with a previous diagnosis of alcoholism. Our results suggest that vitreous humor CDT levels are useful in cases where the postmortem diagnosis of alcoholism is hindered by the non-specificity of data.     

Bupropion Overdose Resulted in a Pharmacobezoar in a Fatal Bupropion (Wellbutrin®) Sustained‐release Overdose: Postmortem Distribution of Bupropion and its Major Metabolites

Bupropion (BUP) overdose commonly causes generalized seizures and central nervous system depression. The case of a 28‐year‐old woman who died from a massive lethal overdose with sustained‐release bupropion (Wellbutrin^® 300 mg) is herein presented. The autopsy revealed the presence of a pharmacobezoar consisting of at least 40 tablets in the stomach. Determination of bupropion and its active metabolites (hydroxybupropion, threobupropion, erythrobupropion) was achieved by a liquid chromatographic mass spectrometry (LC‐MS/MS) method. Postmortem concentrations for bupropion, hydroxybupropion, threobupropion, and erythrobupropion were obtained in intracranial blood, urine, bile, liver, kidney, and vitreous humor. In this case, intracranial blood level of the parent drug was 1.9 mg/L. Threobupropion was the most abundant metabolite in both blood and urine, 59.3 and 890.6 mg/L. Tissue distribution showed the highest concentration in the liver, 12.3 mg/kg. The 0.8 bupropion concentration ratio vitreous/blood suggested that vitreous could be a valuable specimen for toxicological analysis should postmortem blood be unavailable.     

Accidental death from hydromorphone ingestion

Abstract: A 15‐year‐old male orally consumed an unknown but fatal amount of sustained release hydromorphone. He was naïve to opioid use. No other drugs or alcohol were involved. The cause of death was acute aspiration‐related bronchopneumonia, secondary to hydromorphone ingestion; the manner of death was accidental. Hydromorphone and hydromorphone‐3‐glucuronide were quantified in postmortem fluids by tandem liquid chromatography–mass spectrometry. The hydromorphone concentrations in the peripheral blood, urine, and vitreous humor were 57, 4460, and 31 ng/mL, respectively. The hydromorphone‐3‐glucuronide concentrations in the corresponding three fluids were 459, 36,400, and 40 ng/mL. Hydromorphone‐3‐glucuronide accumulation probably did not contribute significantly to the opiate toxicity. The proposed minimum lethal hydromorphone blood concentration in the nontolerant user is in the vicinity of 60 ng/mL.     

家兔眼玻璃体液21例元素含量PMI关系的研究

目的寻找一种精确推定PMI的方法。方法应用电感耦合等离子体质谱(ICP-MS),系统研究了家兔死后96h内眼玻璃体液21种元素含量与PMI的相关性。结果眼玻璃体液Co、Mo、Cd、Sb、I、Pb、Bi、Li共8种元素与PMI无关。K、Mg、Fe、P、Na、Al、Ca、Ti、Ni、Cu、Zn、Sr、Ba共13种元素含量与PMI有关。结论眼玻璃体液K、Mg、Fe、P、Na、Al、Ca、Ti、Ni、Cu、Zn、Sr、Ba共13种元素与PMI有关。为研究人尸体眼玻璃体液元素与PMI的关系奠定了选择指标的基础。     

Vitreous humor biochemical constituents: evaluation of between-eye differences

The present study was conducted to investigate the differences in the vitreous humor biochemical concentrations for vitreous electrolytes and calcium in the same pair of eyes at identical postmortem interval (PMI). The vitreous humor samples were collected independently in both eyes from 48 autopsies (PMI range, 4.5-84.3 hours) with documented time of death. The samples were analyzed for potassium, sodium, chloride, and calcium using a Beckman Coulter LX20 Automated Analyzer based on ion-selective electrode methodology. There were no statistically significant between-eye differences at identical postmortem interval. A significantly high correlation was observed between paired potassium concentrations of both the eyes. A highly significant linear correlation was observed between the individual eye and mean potassium concentrations of both the eyes with postmortem interval. The observed differences were not significantly correlated with postmortem interval. The results demonstrated that the between-eye differences for vitreous electrolytes and calcium are insignificant. Therefore, the utility of vitreous biochemistry, particularly potassium in postmortem interval estimation and other forensic applications, cannot be questioned solely on the basis of these differences.     

Research Progress on Postmortem Interval Estimation by Vitreous HumorCSCD

Postmortem interval (PMI)estimation is one of the most challenging problems in the field of forensic science. Vitreous humor is a hotspot which has been used for PMI estimation and postmortem chemical analysis in forensic pathology. In order to provide novel perspectives for the future research of PMI estimation using vitreous humor, the comparison between vitreous humor with other common body fluids, the effect of temperature on vitreous humor, vitreous humor detection method and data fitting method have been reviewed in this paper. © 2018 by the Editorial Department of Journal of Forensic Medicine.     

Fatality from olanzapine induced hyperglycemia

A case history of a 31-year-old male schizophrenic patient is presented. The man was treated with olanzapine for three weeks before he died. After one week on a 10 mg daily dose of olanzapine, his fasting blood glucose was elevated to 11.3 mmol/L (203 mg/dL). In order to treat more aggressively his psychosis, the olanzapine dose was raised to 20 mg daily resulting in his fasting blood glucose climbing to 15.8 mmol/l (284 mg/dL). On the days preceding his death, he became progressively weaker, and developed polydipsia with polyuria. He had no personal or family history of diabetes mellitus and he was on no other medication at the time of his death. Postmortem blood, vitreous humor, and urine glucose concentrations were 53 mmol/L (954 mg/dL), 49 mmol/L (882 mg/dL), and 329 mmol/L (5922 mg/dL), respectively. Drug screen on urine and blood indicated only a small amount or olanzapine and no alcohols. Peripheral blood olanzapine concentration was within therapeutic limits, 45 ng/mL. Analysis of vitreous humor and urine revealed severe dehydration with small amounts of ketones. Death was attributed to hyperosmolar nonketotic diabetic coma, and olanzapine was felt most likely to be the cause. Another atypical neuroleptic, clozapine, has also been associated with the development and exacerbation of diabetes mellitus or diabetic ketoacidosis. We recommend including vitreous glucose and beta-hydroxybutyrate analysis as part of postmortem toxicology work up when the drug screen reveals the presence of either olanzapine or clozapine.     

The use of vitreous humor as an alternative to whole blood for the analysis of benzodiazepines

In postmortem drug analysis, the most commonly used sample matrix is whole blood. However, postmortem changes can denature this matrix, resulting in a loss or degradation of drugs, thus biasing analytical findings. Vitreous humor is thought to be less affected by these changes and should, therefore, have the potential to provide a more reliable estimation of antemortem drug concentrations. To assess the usefulness of vitreous humor for the analysis of benzodiazepine drugs, vitreous humor and whole blood were obtained postmortem in 27 cases. Three benzodiazepine drugs were investigated-temazepam, diazepam, and desmethyldiazepam. For temazepam and diazepam, some correlation was found between the matrices (R2 = 0.789 and 0.724, respectively). However, for desmethyldiazepam, no correlation was observed (R2 = 0.068). Regression analysis on plots of vitreous humor versus blood concentrations produced gradients of less than 1.0 showing that, in general, levels in blood are higher than the corresponding levels in vitreous humor.     

A Study on the Estimation of Postmortem Interval Based on Environmental Temperature and Concentrations of Substance in Vitreous Humor

A method to determine postmortem interval (PMI) based on environmental temperature and the concentrations of vitreous humor (VH) molecules were explored. Rabbit carcasses were placed in a chamber at 5, 15, 25, or 35°C, and 80–100 μL of VH was collected with the double‐eye alternating micro‐sampling method every 12 h. A Roche DPPI biochemical analyzer was used to measure the concentrations of six substances in VH samples. The interpolation function model and mixed‐effect model were employed for data fitting to establish equations for PMI estimation. The concentrations of K⁺, P, Mg²⁺, creatinine (CRE), and urea nitrogen (UN) exhibited an upward trend with increasing PMI in all temperature groups, while the concentration of Ca²⁺ showed a downward trend. Validation results using K⁺ and Mg²⁺ ions revealed that the mixed‐effect model provided a better estimation than the interpolation function model using the data from our experiment. However, both models were able to estimate PMI using temperature and VH molecule concentrations.     

Postmortem vitreous humor beta-hydroxybutyrate: its utility for the postmortem interpretation of diabetes mellitus

Ketoacidotic coma is one of the most serious complications arising from diabetes mellitus, especially type I, and may be the cause of sudden death especially in diabetes type I. Since beta-hydroxybutyrate (beta-OHB) serum concentrations might provide more information on the severity of ketoacidosis, the aim of this study was to evaluate the concentrations of beta-OHB in vitreous humor and its correlation with other biochemical parameters during postmortem examination. We intended to ascertain the sensitivity and the specificity of these markers for diagnosing diabetes mellitus and the presence of ketoacidosis. This study involved 453 cadavers with a mean age of 57.6 years (S.D. 20.7) and a mean postmortem interval of 17.8 h (S.D. 9.6, range 2-61 h). Cases were assigned to two diagnostic groups according to the antemortem diagnosis of diabetes mellitus, based on the patients' medical records. In vitreous humor statistically significant differences were found in biochemical marker concentrations between the two diagnostic groups, the highest values being obtained in the group of subjects with a previous diagnosis of diabetes mellitus. The measurement of beta-OHB in vitreous humor may be a useful alternative to using blood during postmortem analysis. The presence of high levels of beta-OHB may help interpret the cause of death in diabetics when the autopsy result is negative.     

The Stability of 4-Chloromethcathinone in Blood and Vitreous Humor

We present results of our study on the stability of 4-chloromethcathinone (4-CMC) in authentic postmortem peripheral blood and vitreous humor samples. The stability of 4-CMC was determined in postmortem blood samples (for a period of 90 days) and vitreous humor (30 days) at three different temperatures: −15°C, +4°C, and + 23°C. The analyses were carried out using ultra-high-performance liquid chromatography coupled with triple-quadrupole tandem mass spectrometry (UHPLC-QqQ-MS/MS). In both materials, the lowest 4-CMC stability was demonstrated at room temperature. The blood samples stored in a freezer (−15°C) showed stability for the entire study period (90 days), while in the case of the vitreous humor sample stored at the same temperature the concentration of the substance decreased by 53% after 30 days. The study carried out in authentic postmortem blood and vitreous humor samples confirms the previous reports of 4-CMC instability in biological material. Authors suggest that the biological material should be stored frozen until analyses are carried out as soon as possible after collection of the material.     

Postmortem Distribution of 3‐Beta‐Hydroxybutyrate

The concentrations of 3‐beta‐hydroxybutyrate (3HB) in femoral blood, urine, vitreous humor as well as pericardial and cerebrospinal fluids were retrospectively examined in a series of medico‐legal autopsies, which included cases of diabetic ketoacidosis, hypothermia fatalities without ethanol in blood, bodies presenting mild decompositional changes, and sudden deaths in chronic alcoholics. Similar increases in 3HB concentrations were observed in blood, vitreous, and pericardial fluid, irrespective of the cause of death, suggesting that pericardial fluid and vitreous can both be used as alternatives to blood for postmortem 3HB determination. Urine 3HB levels were higher than blood values in most cases. Cerebrospinal fluid 3HB levels were generally lower than concentrations in blood and proved to be diagnostic of underlying metabolic disturbances only when significant increases occurred.     

Screening for barbiturates in vitreous humor by the EMIT-st serum enzyme immunoassay

In 16 medical examiner's cases, which were found to be barbiturate-positive by thin-layer chromatographic screening of the liver, blood barbiturate concentrations were determined by gas chromatography. The corresponding vitreous humor samples were screened by the enzyme multiplied immunoassay technique, the EMIT-st serum barbiturate assay. By using the recommended dilution for detecting serum barbiturates, it was possible to detect barbiturates in vitreous humor at a toxic concentration. By using one fourth the amount of diluent, the barbiturates could be detected also at a therapeutic concentration. The EMIT-st assay proved to be useful in the screening for barbiturates in vitreous humor, a material that is readily available in forensic toxicology.     

References for determining the time of death by potassium in vitreous humor

The different statements concerning the slope and intercept of the regression line and the 95% limits of confidence are the reason that potassium in vitreous humor is not used (at least in Germany) as an aid in estimating the time of death. The relationship between the concentration of potassium and the time of death is mainly influenced by antemortem electrolyte imbalances caused by disease and/or duration of terminal episode. The influence of terminal episode is best identified by its duration (Adelson et al., J. Forensic Sci., 8 (1963) 503–514). In order to have a method suitable for every case and to be as precise as possible we looked therefore for parameters in vitreous humor which were stable postmortem and indicating antemortem electrolyte imbalance. Urea is such a parameter, being stable postmortem (Coe, Am. J. Clin. Pathol., 51 (1969) 741–750) and useful as a marker of antemortem electrolyte imbalance. Our investigations on potassium in vitreous humor, including sudden and hospital deaths after chronic lingering disease, revealed 95% limits of confidence of ±34 h up to 120 h postmortem. Reviewing only cases with urea less than 100 mg/dl the 95% limits of confidence could be reduced to ±22 h. Considering the duration of terminal episode (<6 h) the precision was ±20 h. In this way our modified procedure is suitable for every case with the resulting precision of estimation being determined only by the duration of the terminal episode and urea concentration.