Aims: To develop a non-invasive method to demonstrate the presence of haemoglobin and its degradation products in bruises in live human subjects for the purposes of objectively assisting in the determination of the age of a bruise. Methods: The cuvette holder unit of a Cary 100 Bio UV-Visible Spectrophotometer was replaced with the manufacture's fibre optic cable and optical reflectance probe. The probe was placed on the skin surface. The absorption spectrum from 780 to 380 nm was collected and transformed into the first derivative. Calculation of the first derivative permits absorption attributed to haemoglobin degradation (primarily to bilirubin, but also haemosiderin) to be separated from absorption by haemoglobin. First derivative and colorimetry values, expressed as CIEL*a*b data, were derived from scans of 50 bruises. Results: The fibre optic cable and probe allowed the spectrophotometer to collect reproducible absorption spectra of bruises in the skin of living subjects. A bruise at three days has greater negative first derivative values at 480 and 490 nm than does a fresh bruise, indicating the local degradation of haemoglobin. Correlation between the first derivative and the CIEL*a*b ''b'' values in a series of bruises indicates that the yellow colour in a bruise is proportional to the amount of local haemoglobin breakdown. Conclusion: The ability to demonstrate the presence of haemoglobin and measure its degradation in bruises in living human subjects by a non-invasive method has not been described previously, and may be of use in the objective ageing of bruises for forensic purposes. B y visual inspection it may be possible to suggest that a bruise is either recent (fresh) or old (more than 18 hours), based on the presence or absence of yellow colour.

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Bruises can have medicolegal significance such that the age of a bruise may be an important issue. This study sought to determine if colorimetry or reflectance spectrophotometry could be employed to objectively estimate the age of bruises. Based on a previously described method, reflectance spectrophotometric scans were obtained from bruises using a Cary 100 Bio spectrophotometer fitted with a fibre-optic reflectance probe. Measurements were taken from the bruise and a control area. Software was used to calculate the first derivative at 490 and 480 nm; the proportion of oxygenated hemoglobin was calculated using an isobestic point method and a software application converted the scan data into colorimetry data. In addition, data on factors that might be associated with the determination of the age of a bruise: subject age, subject sex, degree of trauma, bruise size, skin color, body build, and depth of bruise were recorded. From 147 subjects, 233 reflectance spectrophotometry scans were obtained for analysis. The age of the bruises ranged from 0.5 to 231.5 h. A General Linear Model analysis method was used. This revealed that colorimetric measurement of the yellowness of a bruise accounted for 13% of the bruise age. By incorporation of the other recorded data (as above), yellowness could predict up to 32% of the age of a bruise-implying that 68% of the variation was dependent on other factors. However, critical appraisal of the model revealed that the colorimetry method of determining the age of a bruise was affected by skin tone and required a measure of the proportion of oxygenated hemoglobin, which is obtained by spectrophotometric methods. Using spectrophotometry, the first derivative at 490 nm alone accounted for 18% of the bruise age estimate. When additional factors (subject sex, bruise depth and oxygenation of hemoglobin) were included in the General Linear Model this increased to 31%-implying that 69% of the variation was dependent on other factors. This indicates that spectrophotometry would be of more use that colorimetry for assessing the age of bruises, but the spectrophotometric method used needs to be refined to provide useful data regarding the estimated age of a bruise. Such refinements might include the use of multiple readings or utilizing a comprehensive mathematical model of the optics of skin.

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