IDENTIFICATION OF INTERMEDIATES AFTER INHIBITION OF CHOLESTEROL-SYNTHESIS BY AMINOTRIAZOLE TREATMENT INVIVO

Cholesterol synthesis from mevalonate is inhibited by aminotriazole treatment in vivo. We tried to identify intermediates accumulated in liver of aminotriazole-treated rats. At 6 h after the aminotriazole treatment, the liver was excised. Sterols were extracted from it, and subjected to capillary gas-liquid chromatography, high-performance liquid chromatography, gas-liquid chromatography linked to mass spectrometry and gas-liquid chromatography linked to Fourier-transform infrared spectrometry. It was found that 4-alpha-methyl-5-alpha-cholest-7-en-3-beta-ol and 4,4-dimethyl-5-alpha-cholest-8-en-3-beta-ol were accumulated in the liver, mainly as the free forms. The contents of the former and the latter were increased to 25- and 64-times the control values, respectively. In another experiment, [2-C-13]mevalonate was injected at 2 h after aminotriazole treatment, and 4 h later the liver was excised. The sterols extracted from the liver were subjected to gas-liquid chromatography linked to mass spectrometry. Specific fragment ions reflecting the incorporation of [C-13] mevalonate were detected in the mass spectra of the intermediate sterols. Accumulation of 4-alpha-methyl-5-alpha-cholest-7-en-3-beta-ol and 4,4-dimethyl-5-alpha-cholest-8-en-3-beta-ol after aminotriazole treatment suggests that elimination of the 4-alpha-methyl group from 4-methyl intermediate sterols is inhibited by aminotriazole.