Journal de recherche médico-légale

Quantitative Determination of Hemoglobin in Tooth and Bone by Uv/Vis Spectroscopy and Cyclic Voltammetry

Abstract

Stephanie E Hooper Marosek, L Kayci Miller Gowins and Alice H Suroviec

Detection and quantification of hemoglobin in skeletal or dried remains could potentially have versatile utilization with the forensic field. However, the majority of studies within this area focus on the identification of hemoglobin and its derivatives rather than determining the relative amount of protein present. This research focuses on the use of leuco crystal violet with Ultraviolet/Visible spectroscopy and applying cyclic voltammetry for the determination and quantification of hemoglobin in a tooth and femur sample. Using leuco crystal violet, possible interferences were also investigated, and it was determined that ferrous iron samples appeared to actually mask the overall reaction, while glycine had no obvious effect. Albumin showed no apparent interference, while myoglobin produced a very faint green color that absorbed at a different wavelength, so this species produced minimal interference. Extractions of hemoglobin from a femur bone using 0.5 M hydrochloric acid and from a tooth using 6.0 M urea all yielded the violet product when reacted with leuco crystal violet. Using the measured absorbance of each positive extraction, the corresponding amount of hemoglobin in each sample was determined. Hemoglobin standards and extraction samples from the tooth and femur bone were also analyzed by the electrochemical method, cyclic voltammetry. Hemoglobin levels in the tooth and femur were found to be in the micromolar range in both the spectroscopic and electrochemical analysis, and these levels are consistent with a decrease in hemoglobin concentration postextraction and post-mortem