Postmortem biochemistry is a valuable tool in forensic investigations, providing insights into the tissue damage and organ dysfunction associated with death. This study aimed to identify biochemical markers that distinguish primary and secondary hypothermia. Twenty-one Wistar rats were allocated into three groups: the Control group (n = 7), which was exposed only to hypothermic conditions, the Alcohol + Hypothermia group (n = 7), and the Benzodiazepines + Hypothermia group (n = 7). The temperature metrics assessed included the normal core temperature, the post-ketamine (0.3 ml injection) core temperature, the immersion temperature, temperature at the onset of hypothermia, and temperature at death. Blood samples were collected from the thoracic aorta in EDTA vacuum tubes for biochemical analysis. The key biochemical parameters measured included the Total Protein (g/L), Albumin (g/L), Globulin (g/L), Albumin to Globulin Ratio, Alanine Aminotransferase (U/L), Alkaline Phosphatase (U/L), Cholesterol (mmol/L), Amylase (U/L), and Lipase (U/L), using an automated IDEXX (Netherlands) cell counter. Significant between-group differences were found for the total protein and globulin levels (p < 0.001 and p = 0.002, respectively), with post-hoc tests confirming differences between the alcohol and control, and benzodiazepine and control groups. The cholesterol levels were found to be significantly different through an omnibus test (p = 0.03), but post hoc tests did not confirm these differences on a statistically significant level. The amylase levels varied significantly across all groups (p < 0.001), with post hoc tests confirming significant differences among all pairs: alcohol vs. benzodiazepine (p = 0.002), alcohol vs. control (p = 0.003), and benzodiazepine vs. control (p < 0.001). The lipase levels showed significant differences in the omnibus test (p = 0.030), but there was no significance in the post hoc tests. Amylase emerged as the most significant parameter in our study, with reduced levels strongly associated with secondary hypothermia. These findings highlight the potential use of total protein, globulin, and amylase levels as biomarkers to differentiate between primary and secondary hypothermia in forensic contexts.

Microscopic signs indicative of drowning are not specific to drowning but also to any other form of suffocation where mechanical obstruction is involved. Our study aimed to evaluate both macroscopic and microscopic findings across different groups sharing a common mechanism of death but differing causes and to compare the diatom test with pathohistological examination.Twenty-nine adult Wistar rats, weighing within recommended ranges, were divided into four groups (L1-L4). The diatom test followed established guidelines for diatoms in water from the Bosna River. Microscopic examination revealed diatoms in the lungs of rats in L3 and L4 groups. Pathohistological findings showed varying degrees of changes including consolidation and inflammatory cell infiltration, dominated by lymphocytes and macrophages, with some samples also showing eosinophilic leukocytes.Significant differences were observed between animals whose cause of death was mechanical asphyxia (suffocatio) and those that were submersed for1 hour versus those that were submersed for 72 hours after death. Diatoms identified in group L4 samples 3, 4, and 5 included Navicula sp. (U3 and U6) and Ulnaria ulna (U4).Our findings suggest combining the diatom test with pathohistological analysis to support a drowning diagnosis. Further examination of other organs could enhance result reliability.

Background: There is no specified diagnostic procedure that can help in determining the cause of death and the diagnosis of drowning because the pathohistological signs are almost identical and non-specified. Aim: Our study aims to recognize and prove diatom appearance in organs from a forensic aspect in Bosnia and Herzegovina, and to examine which is the more specific method in the diagnosis of drowning, the diatom test or the pathohistological finding. Methods: Rats of the recommended body weight were divided into four groups: G1 (n = 8; mechanism of death—asphyxia; cause of death—suffocation, submerged 1 hour after death); G2 (n = 8: mechanism of death-asphyxia; cause of death-suffocation, immersed 72 hours after death); G3 (n = 8: mechanism of death-asphyxia; cause of death-drowning, autopsy immediately after death), and G4 (n = 8: mechanism of death-asphyxia; cause of death-drowning, post mortem 24 hours after death). Results: During the diatom analysis, four species of diatoms, Diatoma vulgaris, Melosira varians, Epithemia adnata, and Cymbella sp, were successfully recovered from the stomach. Microscopic analysis did not detect diatoms in the kidneys and brains of rats, while the pathohistological changes were relatively uniform. Conclusion: Our results propose that the diatom test is a sustainable tool for supporting the diagnosis of drowning in the forensic pathology analysis of the cause of death. This experimental study is a starting point toward the optimization of tests and sampling in cases of unexplained etiology.