Free amino acid concentrations in vitreous humor and cerebrospinal fluid in relation to the cause of death and postmortem interval

We studied free amino acids in vitreous humor and cerebrospinal fluid from 58 cadavers in the course of routine medicolegal autopsies in the city of Granada. The main objective was to establish whether free amino acids contents in these fluids were related with the cause of death, postmortem interval, and severity of the classic signs of asphyxia. The amino acids (aspartic acid, glutamic acid, serine, glutamine, glycine/threonine/histidine, citruline, arginine, alanine, taurine, GABA, tirosine, valine, methionine, isoleucine, phenylalanine/tryptophan, leucine, and lysine) were quantified by high performance liquid chromatography. There were no statistically significant differences in amino acids concentrations in vitreous humor when the different causes of death were considered. Our results did not show any statistically significant relationship when asphyxial score was plotted against the vitreous content of each amino acid. A statistically significant increase with postmortem interval was observed in vitreous taurine (r = 0.3191, p = 0.01461), glutamate (r = 0.4323, p = 0.0007) and particularly in aspartate (r = 0.4508, p = 0.0003).     

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 (less than 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.     

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.     

Ethyl sulphate and ethyl glucuronide in vitreous humor as postmortem evidence marker for ethanol consumption prior to death

To clarify the circumstances of death, the degree of inebriation is of importance in many cases, but for several reasons the determination of the ethanol concentration in post-mortem samples can be challenging and the synopsis of ethanol and the direct consumption markers ethyl glucuronide (EtG) and ethyl sulphate (EtS) has proved to be useful. The use of a rather stable matrix like vitreous humor offers further advantages. The aim of this study was to determine the concentrations of ethanol and the biomarkers in the robust matrix of vitreous humor and to compare them with the respective levels in peripheral venous blood and urine. Samples of urine, blood from the femoral vein and vitreous humor were taken from 26 deceased with suspected ethanol consumption prior to death and analyzed for ethanol, EtS and EtG. In the urine samples creatinine was also determined. The personal data, the circumstances of death, the post-mortem interval and the information about ethanol consumption prior to death were recorded. EtG and EtS analysis in urine was performed by LC-ESI-MS/MS, creatinine concentration was determined using the Jaffé reaction and ethanol was detected by HS-GC-FID and by an ADH-based method. In general, the highest concentrations of the analytes were found in urine and showed statistical significance. The mean concentrations of EtG were 62.8mg/L (EtG100 206.5mg/L) in urine, 4.3mg/L in blood and 2.1mg/L in vitreous humor. EtS was found in the following mean concentrations: 54.6mg/L in urine (EtS100 123.1mg/L), 1.8mg/L in blood and 0.9mg/L in vitreous humor. Ethanol was detected in more vitreous humor samples (mean concentration 2.0g/kg) than in blood and urine (mean concentration 1.6g/kg and 2.1g/kg respectively). There was no correlation between the ethanol and the marker concentrations and no statistical conclusions could be drawn between the markers and matrices.     

家兔死后玻璃体液电导率变化与死亡时间的关系

目的探讨家兔死后玻璃体液电导率变化与死亡时间的相关性。方法采用电导分析法检测家兔30℃下死后48h内及20℃下死后120h内玻璃体液电导率变化。结果30℃和20℃下家兔死后玻璃体液电导率随死亡时间延长而持续升高。经统计分析得到抛物线型二项式回归方程,其相关系数分别为0.970和0.983,两式P〈0.01。结论家兔死后玻璃体液电导率升高程度可以作为推断死亡时间的参考指标。     

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.     

Determination of calcium and magnesium in postmortem human vitreous humor as a test to ascertain the cause and time of death

The concentrations of magnesium and total calcium human vitreous humors obtained postmortem was tested. In the groups of heart disease and asphyxia a correlation between the postmortem interval and calcium or magnesium concentrations was found. There was no correlation between postmortem interval and calcium respectively magnesium in the other causes of death. It seems possible that the analysis of vitreous humor may be useful in both diagnosis and determination of postmortem interval in the cases of heart death and asphyxia.     

Postmortem serum uric acid and creatinine levels in relation to the causes of death

Serum uric acid (UA) and creatinine (Cr) mainly derive from skeletal muscle tissues. Although, remarkable postmortem stability of the serum levels has been reported, there appears to be very poor knowledge of the diagnostic value in investigation of death, except for uremia. The aim of the present study was to evaluate postmortem serum UA and Cr levels using 395 forensic autopsy cases, in comparison with blood urea nitrogen (BUN), for investigation of the pathophysiology of death with special regard to the causes of death involving possible skeletal muscle damage, e.g. due to hypoxia, heat or agonal convulsions. Cr and BUN showed relatively good topographic stability in the cadaveric blood, whereas, UA was often much higher in the right heart blood than in the left heart and peripheral blood, independent of postmortem intervals. Moderate to marked elevation of Cr and BUN accompanied with hyperuricemia was observed in delayed death. In the acute death cases (survival time <30 min), UA, especially in the right heart blood, showed a considerable elevation in mechanical asphyxiation and drowning. The Cr level in fire victims with a lower carboxyhemoglobin (COHb) level (<60%) was significantly higher than in those with the possible fatal level (>60%). A similar elevation of Cr was observed in fatalities from heat stroke and methamphetamine (MA) poisoning. The observations suggested that hyperuricemia in acute death may be indicative of advanced hypoxia and that elevated Cr level may reflect the skeletal muscle damage, especially due to thermal influence.     

A, B and H group specific substances in human vitreous humor

The presence of A, B and H group specific substances in vitreous humor taken from 105 human corpses was determined. Good agreement was obtained between these group substances and the ABO blood group. The relationship with the secretor type is discussed.     

家兔玻璃体液微量取样与一次取样的比较

目的比较微量取样与一次取样的优劣。方法36只家兔分为4组(每组9只),于死后12h取样:I组于一次取出所有VH;II组微量取样50μl,连续取15次;III组左眼一次取样。右眼微量重复取样。IV组双眼第1次微量取样,剩余一次取出。检测Ca2 、Cl-、K 、Na 和P等含量,数据统计分析。结果III组左右眼、(IV)组先后两次取样配对t检验无差异(P>0.05),各组微量取样样本的变异系数、双眼差较小。结论微量取样的操作影响较小、稳定性和重复性好,更适合法医学研究和应用。     

家兔玻璃体液微量取样与一次取样的比较

目的比较微量取样与一次取样的优劣。方法36只家兔分为4组（每组9只），于死后12h取样：Ⅰ组于一次取出所有VH；Ⅱ组微量取样50μl，连续取15次；Ⅲ组左眼一次取样。右眼微量重复取样。Ⅳ组双眼第1次微量取样．剩余一次取出。检测Ca^2＋、Cl^-、K^＋、Na^＋和P等含量，数据统计分析。结果Ⅲ组左右眼、（Ⅳ）组先后两次取样配对t检验无差异（P〉0．05）．各组微量取样样本的变异系数、双眼差较小。结论微量取样的操作影响较小、稳定性和重复性好．更适合法医学研究和应用。     

Immunohistochemistry of pulmonary surfactant apoprotein A in forensic autopsy: reassessment in relation to the causes of death

To reassess the immunohistochemical distribution of pulmonary surfactant apoprotein A (SP-A) in relation to the causes of death, 282 forensic autopsy cases were reviewed. The most intense and dense granular immunostaining of intra-alveolar SP-A was observed in the hyaline membrane syndrome from various traumas, protracted death from drowning, and perinatal aspiration of amniotic fluid. Similar granular staining pattern was found in fatal poisoning by a muscle relaxant and organophosphate pesticides. An evident increase of intra-alveolar granular staining was noted in most fatalities from mechanical asphyxia and drowning, and some cases of fire death. SP-A staining was usually very weak or sparse in alcohol intoxication, poisoning by hypnotics and also carbon monoxide poisoning. These findings suggest that the amount of intra-alveolar granular SP-A staining may be a possible indicator of severity and duration of respiratory distress (agony) from peripheral (non-central nervous system) origin and alveolar damage.     

家兔死后眼玻璃体液消光度与死亡时间关系研究

目的 为寻找一种精确推定死亡时间的方法。方法 应用 754分光光度计,在 420nm波长下,检测了 48只家兔死后不同时间的眼玻璃体液消光度,探讨其与死亡时间的关系。结果 眼玻璃体液消光度（ X）与死亡时间（ Y）呈线性正相关（ r=0.983 27, P< 0.05）,在死后 0～ 72小时,眼玻璃体液消光度与死亡时间存在回归关系（ F=116.54, P< 0.05）,直线回归方程为： Y = 453.30X＋ 0.75 [Y为死亡时间 (PMI),单位为小时; X为眼玻璃体液消光度 ]。结论眼玻璃体液消光度的测定可作为推断死后 72小时内死亡时间的参考指标之一。     

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。     

家兔玻璃体液间隔多次微量取样方法研究

目的探讨间隔多次微量取样玻璃体液是否可用于死亡时间推断。方法99只家兔分为2大组(I)大组45只,再分为A、B、C、D、E五个小组(每组9只),右眼(实验眼)于死后间隔12h多次取样直至24h(A组)、48h(B组)、72h(C组)、96h(D组)和120h(E组);各小组左眼(对照眼)仅分别于死后24h、48h、72h、96h和120h取样一次。检测Ca2+、Cl-、K+、Na+和P含量,同一取样时间的双眼做配对t检验。(II)大组54只,再分为2小组F组9只,每隔12h双眼交替多次取样直至120h;G组45只,每隔12h一次取样10眼(动物左右眼均编号后随机抽取每次10眼),直至120h;检测Ca2+、K+和Na+,两组差值做t检验及PMI回归方程比较。结果E组K+有差异(P<0.05),其余A~E组双眼物质含量配对t检验均无差异(P>0.05)。F与G组物质含量差值t检验无差异(P>0.05);K+、Na+的PMI线性回归方程的斜率、截距无差异(P>0.05),Ca2+与PMI无相关性。结论控制间隔时间和取样次数,多次微量取样对死后玻璃体液元素含量的规律性变化无显著影响,可以替代传统方法进行死亡时间研究。