Materials and methods

Study design and forensic setting

The present investigation was conceived as an observational, exploratory pilot study aimed at evaluating the feasibility and postmortem behavior of Bioelectrical Impedance Analysis (BIA) parameters in relation to the postmortem interval (PMI). The study was conducted between September and November 2024 at the Institute of Legal Medicine of the University Magna Graecia of Catanzaro, within the routine medico-legal activity of the institute. All measurements were performed in a routine mortuary environment, prior to body refrigeration whenever possible, and in accordance with standard forensic operating procedures. The study was designed as hypothesis-generating, without predefined cut-off values, in order to explore potential correlations between electrical parameters, PMI, and anthropometric variables. The study was intentionally designed as a cross-sectional exploratory analysis based on routine forensic casework rather than a longitudinal repeated-measurement protocol. Repeated serial measurements on the same cadaver were beyond the operational and medico-legal constraints of the present pilot investigation.

Case selection and inclusion criteria

A total of 29 deceased individuals were consecutively included in the study. Inclusion criteria were the availability of reliable information regarding the time of death, physical accessibility of the body for electrode placement, and the absence of advanced decomposition phenomena that could preclude meaningful bioelectrical measurements. Cases showing severe putrefactive changes, extensive skin loss, or major postmortem trauma interfering with standard electrode positioning were excluded. No restrictions were applied with respect to sex, age, cause of death, or presence of comorbidities, in order to preserve the heterogeneity typical of forensic casework and enhance the external validity of the findings.

Autopsy status and post-autopsy subgroup

Of the 29 included cases, 13 underwent full forensic autopsy according to national medico-legal standards, including systematic opening of the cranial, thoracic, and abdominal cavities. Sixteen cases did not undergo autopsy and were examined externally only. In a predefined subgroup of ten autopsied subjects, a second BIA measurement was performed immediately after completion of the autopsy procedure. This subgroup analysis was specifically designed to investigate the potential impact of internal anatomical disruption, organ manipulation, and fluid redistribution induced by autopsy on postmortem bioelectrical parameters (Fig. 1).

Fig. 1: Experimental study design

Demographic, anthropometric, and forensic variables

For each subject, demographic data including age and sex were recorded. Anthropometric variables consisted of body weight and height, measured or retrieved from medico-legal documentation, and used to calculate body mass index (BMI) as weight in kilograms divided by height in meters squared. Forensic variables included certified or reconstructed time of death, time of bioelectrical measurement, presumed cause of death, and relevant medical comorbidities when available. Ambient temperature was not systematically recorded during all measurements, although examinations were performed under routine mortuary conditions. The postmortem interval was calculated as the elapsed time between death and BIA assessment and was expressed in decimal hours to ensure analytical precision and consistency in statistical analysis.

Bioelectrical impedance analysis protocol

Bioelectrical measurements were performed using a Quantum V Segmental Analyzer (RJL Systems, Detroit, MI, USA), a single-frequency device widely validated for clinical body composition analysis. A tetrapolar electrode configuration was adopted, with surface electrodes placed on the right hand and right foot following standard manufacturer recommendations and previously described forensic applications. The bodies were positioned supine on the autopsy table, with limbs slightly abducted to minimize skin contact and electrical interference. A low-intensity alternating current was applied, and measurements were recorded once stable values were obtained. The electrical parameters analyzed included resistance (R, expressed in ohms), reactance (Xc, expressed in ohms), impedance (Z, expressed in ohms), and phase angle (expressed in degrees). All measurements were performed prior to any invasive procedure, except for the post-autopsy recordings, which were obtained using identical electrode placement and device settings.

Data handling and management

All collected data were entered into a dedicated Microsoft Excel database specifically created for the study. Missing or unavailable data were recorded as blank cells and were not imputed, in order to avoid artificial data distortion. Cases with partial missing data were retained for descriptive analyses but were excluded from specific correlation analyses when relevant variables were unavailable. Data integrity was verified through double-checking of entries against original forensic documentation and device readouts.

Statistical analysis

Statistical analysis was performed with an exploratory and hypothesis-generating approach. Continuous variables were summarized using descriptive statistics, including mean values and ranges, given the limited sample size and the heterogeneous distribution of the data. The postmortem interval (PMI) was expressed in decimal hours to improve temporal resolution and analytical consistency.

The relationships between PMI and bioelectrical parameters, including resistance, reactance, impedance, and phase angle, were evaluated using Pearson’s correlation coefficient in order to explore the strength and direction of linear associations. Correlation coefficients were interpreted according to conventional criteria, distinguishing weak, moderate, and strong associations. The relationship between PMI and body mass index (BMI) was also examined to assess potential confounding effects of anthropometric variability.

To further characterize the temporal behavior of phase angle, exploratory regression modeling was performed. A simple linear regression model was initially applied to describe the relationship between phase angle and PMI. Given the right-skewed distribution of PMI values and the visual inspection of data dispersion, a logarithmic transformation of PMI was subsequently applied, and a log-linear regression model was constructed with phase angle as the dependent variable and the natural logarithm of PMI as the independent variable. Model performance was evaluated using the coefficient of determination (R²), and residual variability was assessed through calculation of the root mean square error.

Comparative evaluation of bioelectrical measurements obtained before and after forensic autopsy was conducted descriptively in the subgroup of cases with paired measurements, without inferential testing, due to the limited sample size. No multivariate analyses were performed, as the study was not designed to develop predictive models but to explore potential time-dependent trends. Statistical significance was set at p < 0.05, and all analyses were conducted using standard spreadsheet and statistical software.

Given the non-normal distribution and right-skewed nature of PMI values, Spearman’s rank correlation coefficient was additionally calculated to assess monotonic associations independently of normality assumptions.