Determination of nitrate in blood by gas chromatography and gas chromatography–mass spectrometry

We devised a sensitive and simple method for determining nitrate in whole blood, using an extractive alkylation technique. Nitrate in whole blood was reduced to nitrite by hydrazine sulfate in the presence of Cu2+ and Zn2+ as catalysts, and alkylated with pentafluorobenzyl bromide using tetradecyldimethylbenzylammonium chloride as the phase-transfer catalyst. The obtained derivative was analyzed qualitatively by gas chromatography–mass spectrometry and quantitatively by gas chromatography with electron-capture detection. The detection limit of nitrate in whole blood was 0.01 mM. The calibration curve was linear over the concentration range from 0.02 to 1.0 mM for nitrate in whole blood. The accuracy and precision of the method were evaluated and the relative standard deviations were found to be within 10%. Using this method, the blood nitrate levels of two victims who committed suicide by inhaling automobile exhaust gas were determined.