Formation of 6 beta-hydroperoxyprogesterone in rat liver microsomes

To verify the concept that molecular oxygen can be enzymically introduced as such into the steroid nucleus, radioactive progesterone and also 5-pregnene-3,20-dione were incubated with microsomal preparations of rat liver. In both cases, a significant amount of radioactive 6 beta-hydroperoxyprogesterone was isolated, together with 6 beta-hydroxyprogesterone and 6-oxoprogesterone. Addition of p-hydroxymercuribenzoate and mercurichloride significantly inhibited the formation of the hydroperoxide, whereas potassium cyanide and carbon monoxide were only partially inhibitory. Addition of 6 beta-hydroperoxyprogesterone to cytochrome P-450 containing fractions of hepatic microsomes induced a Type I difference spectrum characterized by an absorption maximum at 392 nm, a minimum at 420 nm, and an isosbestic point at 407 nm. At 4 degrees C, its apparent dissociation constant was found to be in the same order of magnitude as that of 6 beta-hydroxyprogesterone, namely 1.33 microM. A new pathway in rat liver for the formation of both 6 beta-hydroxyprogesterone and 6-oxoprogesterone via the 6 beta-hydroperoxide as a common precursor, is proposed.