Cross Validation of HS-GC/MS to Quantify Total Carbon Dioxide in Horse Plasma

The use of alkinizing agents by trainers to counteract the accumulation of lactic acid in racehorses has been well documented. A by-product of this administration is elevated total carbon dioxide (tCO2) concentrations. Most regulatory authorities have set the threshold for tCO2 in plasma at 37 mM. The quantification of tCO2 often presents a challenge to laboratories due to the instrumentation required, which can be expensive to use and maintain. The Beckman DxC 600 (Brea, CA) is currently used in our laboratory for tCO2 quantification. The goal of this research was to determine if another analytical method could be used for this procedure. We report the use of headspace gas chromatography-mass spectrometry (HS-GC/MS) as an acceptable alternative to the use of the Beckman DxC 600. A six-point calibration curve ranging from 10 to 60 mM was analyzed along with controls at 15, 25 and 45 mM. Imprecision was found to be <3% at all concentrations. Inaccuracy was measured at <4% at all concentrations. Measurement of uncertainty was determined using the Simplified GUM approach and was found to be 3% at 99.7% confidence level with eight measurements at a tCO2 concentration of 45 mM. Furthermore, the HS-GC/MS was cross-validated according to international protocols with all parameters being met. During cross validation, a standard at a known concentration was analyzed by both instruments. The average difference using 25 replicates in calculated concentrations was <0.1 mM. Also, plasma samples from four untreated horses were analyzed by both instruments. The difference in calculated concentrations between the two instruments was <0.6 mM for all horses. The results show that the use of HS-GC/MS is an acceptable alternative to the use of the Beckman DxC 600 for the quantification of tCO2 in horse plasma.