Proteomics in Forensic Sciences: Identification of the Nature of the Last Meal at Autopsy

Abstract

A long-term psychiatric 40 years-old male patient was found dead at 9:00 a.m. in the clinic where he lived. Death was caused by traumatic injuries, which the sanitary staff imputed to a fall. Nurses declared that the patient refused having breakfast, whereas at autopsy the stomach contained 350 g of whitish semifluid material. Using both shotgun and gel-based proteomics, we demonstrated that the chyme contained partly digested milk- and bread-derived proteins, eaten during a recent breakfast. The conflict between evidence and assertions of the attending sanitary staff prompted the Legal Authority to undertake detailed investigations to ascertain facts and possible responsibilities. The herein characterization provides insights in the in vivo mechanisms of gastric breakdown of food proteins in a real meal. β-lactoglobulin was partially resistant to gastric digestion as confirmed by Western blot analysis, in contrast to caseins and wheat gluten proteins, which had been degraded by gastric fluids. In addition to a complex pattern of gastric proteins (e.g., mucin-5AC, pepsin A-3, pepsinogen C, gastric lipase, gastrokine-2, trefoil factors), chyme contained intact proteins and variably sized food-derived polypeptides arising from peptic and nonpeptic proteolytic cleavage as well as heterodimeric disulfide-cross-linked peptides. These findings suggest that the current analytical workflows offer only a partial picture of the real complexity of the human "digestome".

Use of micro, capillary, and nano liquid chromatography for forensic analysis

“…A Thermo Scientific Ultimate 3000 NF ultrahigh performance LC was used coupled to a Thermo Scientific Q-Exactive Orbitrap MS. Separations were performed employing reverse phase chromatography at a flow rate of 300 nL/min on an EASY-Spray PepMap C18 column 3 μm, 150 mm × 75 μm. In this case, nano-LC-MS/MS was instrumental in sequencing over 250 different peptides of the <6 kDa fraction obtained from SEC [24]. This methodology was used to confirm the last meal a patient had prior to decease, and in this case the results were contradicting of what was reported by the "sanitary staff" at the patient's clinic.…”

“…Detecting and characterizing proteins in hair is an excellent complement and/or alternative to mitochondrial DNA analysis for forensic casework, and methods applying micro-LC, CAPILLARY-LC, and nano-LC to hair protein analysis have been established [19][20][21][22][23][24]. In this regard, micro-LC, CAPILLARY-LC, and nano-LC has demonstrated the ability to provide excellent separation efficiency and low LODs, advantages that will be highly sought after in forensic studies.…”

“…In 2020, Pieri et al [24] investigated the nature of a decedent's last meal from gastric sample collected at autopsy using nano-LC. A Thermo Scientific Ultimate 3000 NF ultrahigh performance LC was used coupled to a Thermo Scientific Q-Exactive Orbitrap MS. Separations were performed employing reverse phase chromatography at a flow rate of 300 nL/min on an EASY-Spray PepMap C18 column 3 μm, 150 mm × 75 μm.…”

Flying blind, or just flying under the radar? The underappreciated power of de novo methods of mass spectrometric peptide identification

“…In addition to traditional (criminal justice) forensics, other fields also face the problem of characterizing a sample when little or nothing is known about its identity or origin, such as archaeology and cultural heritage. To cite just a few examples, proteomics has been used to characterize a victim's stomach contents upon autopsy, 60 identify trace evidence at a crime scene as vomit, 61 identify 3,500-year-old organic residue from tombs in northwestern China as a fermented dairy product, 62 and determine the host animal that was the source of a disease-vector tick's last blood meal. 63 All of the studies just mentioned used either database search or sequence-tag search to identify proteins.…”

Hidden “Digestome”: Current Analytical Approaches Provide Incomplete Peptide Inventories of Food Digests

“…The MS detection of food-derived large-sized polypeptides in a complex digest can be problematic as well, due to suppression ion effects, reduced sensitivity associated with wide charge distribution, and limited fragmentation efficiency . A number of structural features, such as low solubility, intrinsically scarce ionization efficiency, presence of post-translational modifications (e.g., multiple phosphorylation or glycosylation), incompleteness of the protein databases, variability of protein isoforms, and presence of hard to identify disulfide cross-linked peptide heterodimers, , further limits the comprehensive characterization of a food digestome. The complexity can even increase when foods undergo thermal processing as a result of variable protein modifications.…”