Rapid Determination of Carboxyhemoglobin in Postmortem Blood using Fully-Automated Headspace Gas Chromatography with Methaniser and FID

ABSTRACT

A new method, combining fully-automated headspace gas chromatography with methaniser and flame ionisation detector (FID) is introduced to determine carboxyhemoglobin (COHb) in postmortem blood samples. This highly automated procedure utilises a robot-like autosampler for reproducible mixing and thermostating (30 min at 50°C) during carbon monoxide (CO) liberation of COHb. Apart from transferring the blood sample and CO liberating solution (saponin (15 g/L) in 1 M sulphuric acid), all steps are performed without manual intervention. After headspace injection and gas chromatographic separation, the CO is reduced by a nickel catalyst to methane, which is then detected by using FID. The COHb saturation of the sample is calculated as percentage of a 100% carboxylated sample as follows: COHb [%] = Area (Original Sample) * 100 / Area (100% carboxylated sample).

The method was shown to be precise with coefficients of variation between 1.2 and 5.0%. Linearity was obtained from 0.5-100% COHb with excellent correlation (R=0.998). The applicability of the procedure was proven by analysis of postmortem blood samples and the results were compared to those of the standard photometric procedure. The method is especially applicable when postmortem blood samples have decomposed or their hemoglobin composition has been changed by thermal stress.

Keywords:

Carboxyhemoglobin (COHb), postmortem blood, headspace gas chromatography, Alamar blue assay, flame ionisation detector.