More often than not, crime scene investigation is a complex analytical process involving a variety of different tools and techniques. Many of them are oriented towards estimation of the actual time of the crime, an extremely helpful parameter in crime scene reconstruction. In this context, bloodstains are one of the most important biological traces that may provide information on the timing and sequence of events during the commitment of a violent crime. Physicochemical analysis and optical techniques, frequently used for forensic dating of blood traces, are usually reliable only up to 2 months after the event. In contrast, the analysis of bloodstains with Electron Paramagnetic Resonance (EPR) spectroscopy can provide valuable information on the time of the crime even several years after the crime was committed.
In a feasibility study, we followed the time evolution of the EPR spectra of human bloodstains deposited on filter paper. The typical EPR spectra acquired for bloodstains after 1 day (Day 1st) and 76 days (Day 76th) following the sample collection are shown below (left panel). Relevantly, although the major spectral features are conserved, the ratios of their amplitudes (denoted as A and B) markedly evolve as a function of aging time. Specifically, the significant increase of the ratio B/A (right panel) stems from the slow oxidation of iron in bloodstains under ambient conditions. We followed the change of B/A for almost 3 months and found that this change is a converging function that makes it possible to determine the time since deposition of bloodstains.