Steroid profiles of 49 patients having Cushing’s syndrome were investigated. Differential diagnostic criteria of Cushing’s syndrome with adrenocortical adenoma and adrenocarcinoma and features of steroid hormones metabolism were established by combination of classical tests and steroid profiles investigation using HPLC and gas chromatography - mass-spectrometry. The adrenocarcinoma was diagnosed for 22,4% of patients having Cushing’s syndrome and the main biochemical criteria were the increasing of 11-deoxycortisol in blood (>20 ng/ml), the excretion of 18-OH-corticosterone and metabolites of glucocorticoids and pregnenes, 8,2% of patients having Cushing’s syndrome had additionally increased androgens metabolites excretion.