Arpad A. Vass

Decomposition Chemistry of Human Remains: A New Methodology for Determining the Postmortem Interval

This study was conducted to characterize the chemistry associated with the decomposition of human remains with the objective of identifying time-dependent biomarkers of decomposition. The purpose of this work was to develop an accurate and precise method for measuring the postmortem interval (PMI) of human remains. Eighteen subjects were placed within a decay research facility throughout a four-year time period and allowed to decompose naturally. Field autopsies were performed and tissue samples were regularly collected until the tissues decomposed to the point where they were no longer recognizable (encompassing a cumulative degree hour (CDH) range of approximately 1000 (approximately 3 weeks)). Analysis of the biomarkers (amino acids, neurotransmitters, and decompositional by-products) in various organs (liver, kidney, heart, brain, muscle) revealed distinct patterns useful for determining the PMI when based on CDHs. Proper use of the methods described herein allow for PMIs so accurate that the estimate is limited by the ability to obtain correct temperature data at a crime scene rather than sample variability.

The elusive universal post-mortem interval formula

The following manuscript details our initial attempt at developing universal post-mortem interval formulas describing human decomposition. These formulas are empirically derived from data collected over the last 20 years from the University of Tennessees Anthropology Research Facility, in Knoxville, Tennessee, USA. Two formulas were developed (surface decomposition and burial decomposition) based on temperature, moisture, and the partial pressure of oxygen, as being three of the four primary drivers for human decomposition. It is hoped that worldwide application of these formulas to environments and situations not readily studied in Tennessee will result in interdisciplinary cooperation between scientists and law enforcement personnel that will allow for future refinements of these models leading to increased accuracy.

Surface-enhanced Raman scattering detection of chemical and biological agent simulants

Surface-enhanced Raman scattering spectra of chemical and biological agent simulants, such as dimethyl methylphonate, pinacolyl methylphosphonate, diethyl phosphoramidate, 2-chloroethyl ethylsulfide, bacillus globigii, erwinia herbicola, and bacillus thuringiensis were obtained from silver-oxide film-deposited substrates. Thin AgO films ranging in thickness from 50 to 250 nm were produced by chemical bath deposition onto glass slides. Further Raman intensity enhancements were noticed in UV irradiated surfaces due to photo-induced Ag nanocluster formation, which may provide a possible route to producing highly useful plasmonic sensors for the detection of chemical and biological agents upon visible-light illumination.

Microbial Community Analysis of Human Decomposition on Soil

Human decomposition is a complex process, involving a multitude of microbial species. Currently, little is known about the microbial species and processes that occur as cadavers decompose, particularly in outdoor environments. With the development of molecular ecology tools that allow the study of complex microbial communities, the ‘black box’ of the microbiology associated with decomposition is being opened. A preliminary study was performed to evaluate the changes in the soil bacterial and fungal communities that occur in response to human cadaver decomposition, and to assess the potential for using such changes in either of these two microbial communities for forensic time-since-death estimation. Terminal restriction fragment length polymorphism (T-RFLP) and phospholipid fatty acid (PLFA) analyses were performed on soil samples collected from beneath decomposing human cadavers at the University of Tennessees Forensic Anthropology Centre. The soil fungal and bacterial communities showed significant changes in response to decomposition events, with some succession patterns emerging as decomposition progressed. Evidence of sequential changes occurring in the soil microbial community provides support for the idea that specific microbes could be used as time-since-death biomarkers, although major differences in the way replicate cadavers decomposed in this study suggested that both the bacterial and fungal communities may need to be considered to ensure such a method is forensically useful. Some difficulties were also encountered with the polymerase chain reaction (PCR) step in the T-RFLP method being inhibited by soil or decomposition compounds, which suggest that while DNA-based ‘profiling’ methods may be useful for studying the effects of decomposition on soil, other more specific techniques are likely to be most useful in a forensic post-mortem interval context.

Advanced Scientific Methods and Procedures in the Forensic Investigation of Clandestine Graves

Our goal is to discuss the new technologies and procedures that we have developed for the discovery and recovery of buried victims. We argue that forensic investigations of clandestine graves must be grounded in the most advanced scientific methods and evidence-collection techniques available. A structured program that includes an interdisciplinary team of forensic scientists and law enforcement experts is proposed to facilitate all aspects of the investigative and legal process. Such issues are of great relevance because most legal jurisdictions have a number of cases each year and present operating procedures are not standardized. There is a clear need for national dialog to improve our investigative efforts and insure best practices in forensic science across legal jurisdictions and law enforcement agencies.

Antitumor effect of valorphin in vitro and in vivo: combined action with cytostatic drugs.

The action of the cytostatic drugs (epirubicin and vincristine) in combination with the endogenous antiproliferative £]-hemoglobin fragment (33-39), valorphin, was studied in tumor (L929 and A549) cell cultures, primary culture of murine bone marrow cells and in murine model of breast carcinoma in vivo. Simultaneous application of 1 µM valorphin and 1 µM epirubicin, in vitro, did not result in an additive suppressive effect on cell culture growth. Additive effects were achieved with alternating applications of the peptide and the drugs, namely, 0.5 µM (but not 1 µM) epirubicin added 24 h prior to 1 µM valorphin; 1 µM valorphin added 48 h prior to 0.1 µM epirubicin, or 0.1 µM vincristine, or 0.05 µM vincristine, which resulted in 100% cell death in the both series with vincristine and up to 78% cell biomass reduction in the experiments with epirubicin. In the in vivo model (female BLRB mice with subcutaneously inoculated syngeneic mammary carcinoma), simultaneous treatment with 25 mg/m2 epirubicin and 1 mg/kg valorphin resulted in 42% of tumor growth inhibition, as compared with the negative control group and 22% inhibition as compared with the epirubcin-treated group (at 20th day of treatment). Survival was significantly improved (69% compared to 39% in the group treated with epirubicin only) at day 26 after the treatment beginning.

Identification of a disinterred grave by molecular and stable isotope analysis

Confirmation of a potential disinterred grave was sought by GC and GC/MS analyses of lipid extracts of whole soils and white particulate matter. Fatty acid profiles and concentrations determined for three of the soils correlated with the deposition of a large amount of exogenous organic matter, most likely adipocere and/or decomposed body fat. Determination of C16:0 and C18:0 fatty acid delta13C values by GC/C/IRMS revealed the input to be isotopically distinct from common British domesticated animals, plotting closely to values determined for adipose fat obtained from of a murder victim. By considering the difference between delta13C values (delta13C18:0-16:0) a potential isotopic proxy for identifying the source of adipocere (human) and adipose tissue was proposed.

Surface-enhanced Raman scattering (SERS) detection for chemical and biological agents

Surface-enhanced Raman scattering (SERS) spectra of chemical agent simulants such as dimethyl methylphonate (DMMP), pinacolyl methylphosphonate (PMP), diethyl phosphoramidate (DEPA), and 2-chloroethyl ethylsulfide (CEES), and biological agent simulants such as bacillus globigii (BG), erwinia herbicola (EH), and bacillus thuringiensis (BT) were obtained from silver oxide film-deposited substrates. Thin AgO films ranging in thickness from 50 nm to 250 nm were produced by chemical bath deposition onto glass slides. Further Raman intensity enhancements were noticed in UV irradiated surfaces due to photo-induced Ag nanocluster formation, which may provide a possible route to producing highly useful plasmonic sensors for the detection of chemical and biological agents upon visible light illumination.

Mutation analyses of a series of TNT-related compounds using the CHO-hprt assay.

Trinitrotoluene (TNT) and related compounds were tested for induction of mutation in the CHO‐hprt mutation assay. The parent compound, TNT, was consistently found to be mutagenic at concentrations above 40 μg ml−1, whether or not S9 activating enzymes were added. Five TNT metabolites gave statistically significant but small increases in mutation frequency over solvent controls: 4‐amino‐2,6 dinitrotoluene, 2,4′,6,6′‐tetranitro‐2′,4‐azoxytoluene, 2,2′,6,6′‐tetranitro‐4,4′‐azoxytoluene, 2′,4,6,6′‐tetranitro‐2,4′‐azoxytoluene and triaminotoluene. Clear dose–response relationships could not be established for the mutagenic response of these compounds. They are considered as very weak mutagens in this mammalian test system. Five compounds did not produce statistically significant mutation frequencies at the levels tested: 2‐amino‐4,6‐dinitrotoluene, 2,4‐diamino‐6‐nitrotoluene, 1,3,5‐trinitrobenzene, 2,6‐diamino‐4‐nitrotoluene and 4,4′,6,6′‐tetranitro‐2,2′‐azoxytoluene. The results indicate that none of the TNT metabolites tested pose a significant mutational health risk, at least as judged by the CHO‐hprt assay. Copyright

Endogenous fragment of hemoglobin, neokyotorphin, as cell growth factor.

It is shown that neokyotorphin (the alpha-globin fragment 137-141) stimulates proliferation of normal cells (murine embryonic fibroblasts, red bone marrow and spleen cells) and tumor cells (murine melanoma and transformed fibroblasts L929) in the absence or in the presence of fetal bovine serum. In contrast to serum deprivation conditions, the ability to potentiate L929 cell growth in the presence of fetal serum is strongly cell density dependent. The peptide also enhances the viability of L929 cells, murine embryonic fibroblasts and of the primary cultures of murine red bone marrow cells and splenocytes under serum-deprivation conditions for at least 72 h. The results of flow cytometry analysis suggest that the effect of neokyotorphin on survival of L929 cells in serum-free culture medium is due to maintenance of cell proliferation in the absence of growth factors. Along with cell cycle progression the peptide induces reversible reduction of L929 cell size.