Development of a Raman microspectrophotometric method to quantitate carboxyhemoglobin in blood

Abstract

Carboxyhemoglobin (COHb) quantitation is a routine toxicological analysis performed in cases of suspected acute carbon monoxide poisoning. Current methods available for COHb quantitation include spectrophotometric methods and gas chromatographic methods, both of which have advantages and disadvantages. The goal of the present study was to develop a Raman microspectrophotometric COHb quantitation method and assess its precision and accuracy. A strong linear correlation (average R2 = 0.98) was found to exist between COHb concentration and the ratio of the Raman intensities at approximately 1,552 and 1,580 cm−1. The limits of detection and quantitation were found to be 4.4% and 13.2% COHb, respectively. Intraday and interday studies of three calibrators indicated high precision, with the coefficient of variation ranging from 0.85% to 3.4%. The accuracy of this method was evaluated through the quantitation of blind samples, first using the proposed Raman microspectrophotometric method and then by a published UV–visible spectrophotometric method. For the proposed Raman method, 87.5% of the samples were quantitated with less than 20% error. The average variances of the two methods were found to be statistically different using a two‐sample F test, with the Raman microspectrophotometric method having a lower average variance. This study demonstrates that Raman microspectrophotometry may be used to quantitate the COHb concentration of blood samples. Furthermore, this method is comparable with established spectrophotometric methods while eliminating the need for time‐consuming sample preparation and limiting the use of consumables and the generation of waste.