NIR Raman spectra of whole human blood: effects of laser-induced and in vitro hemoglobin denaturation

摘要

Care must be exercised in the use of Raman spectroscopy for the identification of blood in forensic applications. The Raman spectra of dried whole human blood excited at 785 nm are shown to be exclusively due to oxyhemoglobin or related hemoglobin denaturation products. Raman spectra of whole blood are reported as a function of the incident 785-nm-laser power, and features attributable to heme aggregates are observed for fluences on the order of 10~4 W/cm~2 and signal collection times of 20 s. In particular, the formation of this local-heating-induced heme aggregate product is indicated by a redshifting of several heme porphyrin ring vibrational bands, the appearance of a large broad band at 1,248 cm~(-1), the disappearance of the Fe–O2 stretching and bending bands, and the observation of a large overlapping fluorescence band. This denaturation product is also observed in the low-power-excitation Raman spectrum of older ambient-air-exposed bloodstains (2 weeks or more). The Raman spectrum of methemoglobin whole blood excited at 785 nm is reported, and increasing amounts of this natural denaturation product can also be identified in Raman spectra of dried whole blood particularly when the blood has been stored prior to drying. These results indicate that to use 785- nm-excitation Raman spectra as an identification method for forensic applications to maximum effect, incident laser powers need to be kept low to eliminate variable amounts of heme aggregate spectral components contributing to the signal and the natural aging process of hemoglobin denaturation needs to be accounted for. This also suggests that there is a potential opportunity for 785-nm-excitation Raman spectra to be a sensitive indicator of the age of dried bloodstains at crime scenes.