Untargeted Metabolomics Profiling for Determination of the Time since Deposition of Biofluids in a Forensic Context: A Proof-of-Concept for Urine, Saliva, and Semen in Addition to Blood

In a criminal trial, the reconstruction of a crime is one of the fundamental steps of the prosecution process. Common questions, such as what happened, where and how it happened, and who made it happen, need to be solved. Biological evidence at crime scenes can be crucial in the determination of these fundamental questions. One of the more challenging riddles to solve is the when? A trace left at a crime scene can prove a person’s presence at the crime scene. Knowledge about when it was deposited there, the time since deposition (TsD), would allow linking the person in space and time to the site. This could fortify allegations against a suspect or discharge accusations if proven to be outside of the temporal boundaries where a suspected crime had occurred. Determining the TsD has yet to become routine forensic casework, despite recent research efforts, especially for blood traces. However, next to blood, other biological traces are also commonly encountered in crime scenes. We here present a study to profile the metabolomes of artificially aged dried body fluid spots of blood, semen, saliva, and urine over 4 weeks by liquid chromatography high-resolution mass spectrometry and data-dependent acquisition. All four body fluids (BFs) exhibited diverse time-dependent changes, and a large number of molecular features (MF) were associated with TsD. Still, significant differences between the BFs were observed, limiting universal interpretability independent of the BF and facilitating a need to further study time-dependent changes of different BFs individually toward the goal of TsD estimation.