Endothelial dysfunction represents an early step in the development of atherosclerosis. The purpose of this study was to investigate the relationship between postprandial lipaemia and endothelial dysfunction in patients with overt hypothyroidism (oHT) and subclinical hypothyroidism (sHT). Female subjects with oHT and sHT, as well as female healthy subjects with euthyroid state were enrolled (10 cases in each group). The examination of flow-mediated dilation (FMD) was performed before and after an oral fat-loading by high resolution ultrasound. Endothelial dysfunction after an oral fat challenge was related to the extent of hypertriglyceridemia and free radicals. FMD decreased significantly at 4-h point in 3 groups, (p < 0.05) and then FMD in control and sHT restored to baseline at 8-h point, it was lower than baseline in sHT group at 6-h point (p = 0.042). However, FMD continued to decrease at 6-h point (p < 0.001), and then increased toward to baseline at 8-h point, which was still lower than baseline (p = 0.039) in oHT. Spearman's analysis showed a negative correlation between FMD and triglyceride, a negative correlation between FMD and thiobarbituric acid reactive substances (TBARS), and a positive correlation between triglyceride and TBARS levels during oral lipid-loading test in hypothyroid patients (p < 0.001) and controls (p < 0.05). In hypothyroid subjects including oHT and sHT, even in healthy individuals, FMD was impaired after an oral fat challenge. The endothelial dysfunction observed after an oral fat challenge was related to the extent of hypertriglyceridemia and oxygen-derived free radicals.