Development and validation of an ultraperformance liquid chromatography-tandem mass spectrometry method for quantitation of total 3,3',5-triiodo-L-thyronine and 3,3',5,5'-tetraiodo-L-thyronine in rodent serum

Abstract

Many environmental chemicals are known to disrupt thyroid function. Measurement of thyroid hormones in animal studies provides useful information to understand the effects of environmental chemicals on thyroid hormone metabolism. We report an efficient method, utilizing a protein precipitation followed by ultraperformance liquid chromatography-tandem mass spectrometry analysis, to quantitate total 3,3',5-triiodo-L-thyronine (triiodothyronine, T3) and total 3,3',5,5'-tetraiodo-L-thyronine (thyroxine, T4) in rodent serum. The use of synthetic serum for calibration standards eliminated the interferences from endogenous total T3 and T4 and allowed the experimental lower limits of quantitation (LOQ) to be set at the required concentration (T3, 20 ng/dL; T4, 0.5 mu g/dL) to allow quantitation of endogenous concentrations. The method was linear (r > 0.99; range 20.0-600 ng/dL T3, 0.500-15 mu g/dL T4) with good assay recoveries (90.4-107%) for both analytes. Intra- and inter-day accuracy, estimated as percent relative error, were