Determination of chloral hydrate and its metabolites in blood plasma by capillary gas chromatography with electron capture detection

A sensitive, accurate, and reliable method is described for the quantitative determination of chloral hydrate (CH) and its metabolites in blood plasma of mice and rats. Metabolites of CH include trichloroacetic acid (TCA), trichloroethanol (TCE), and trichloroethanol glucuronide (TCE-Glu). This new method uses capillary gas chromatography with electron-capture detection (GC/ECD). Procedures for improving sample stability and quality assurance are also described that were not mentioned in previous literature. Rat or mouse plasma (50 μl) is acidified (or treated enzymatically for TCE-Glu determination) and extracted with peroxide free methyl t-butyl ether. Distilled diazomethane (CH2N2) is added to derivatize TCA to its methyl ester. Detection limits were estimated at 0.2 μg/ml for CH and TCE, and 0.1 μg/ml for TCA. Detector response to TCA and TCE were shown to be linear in the range of 3.125–200 μg/ml (r≥0.9996). For CH, the response fits a second-order equation in this same range (r=0.99994)