Biosynthesis of sterols and ecdysteroids in Ajuga hairy roots

Hairy roofs of Ajuga reptans var, atropurpurea produce clerosterol, 22-dehy droclerosterol, and cholesterol as sterol constituents, and 20-hydroxyecdys one, cyasterone, isocyasterone, and 29-norcyasterone as ecdysteroid constit uents. To better understand the biosynthesis of these steroidal compounds, we carried out feeding studies of variously H-2- and C-13-labeled sterol su bstrates with Ajuga hairy roots. In this article, we review our studies in this field. Feeding of labeled desmosterols, 24-methylenecholesterol, and C -13(2)-acetate established the mechanism of the biosynthesis of the two C-2 9-sterols and a newly accumulated codisterol, including the metabolic corre lation of C-26 and C-27 methyl groups. in Ajuga hairy roots, 3 alpha-, 4 al pha-, and 4 beta-hydrogens of cholesterol were all retained at their origin al positions after conversion into 20-hydroxyecdysone, in contrast to the o bservations in a fern and an insect. Furthermore, the origin of 5 beta-H of 20-hydroxyecdysone was found to be C-6 hydrogen of cholesterol exclusively , which is inconsistent with the results in the fern and the insect. These data strongly support the intermediacy of 7-dehydrocholesterol 5 alpha,6 al pha-epoxide. Moreover, 7-dehydrocholesterol, 3 beta-hydroxy-5 beta-cholest- 7-en-6-one (5 beta-ketol), and 3 beta,14 alpha-dihydroxy-5 beta-cholest-7-e n-6-one (5 beta-ketodiol) were converted into 20-hydroxyecdysone. Thus, the pathway cholesterol --> 7-dehydrocholesterol --> 7-dehydrocholesterol 5 al pha,6 alpha-epoxide --> 5 beta-ketol --> 5 beta-ketodiol is proposed for th e early stages of 20-hydroxyecdysone biosynthesis. 3 beta-Hydroxy-5 beta-ch olestan-6-one was also incorporated into 20-hydroxyecdysone, suggesting tha t the introduction of a 7-ene function is not necessarily next to cholester ol. C-25 Hydroxylation during 20-hydroxyecdysone biosynthesis was found to proceed with ca. 70% retention and 30% inversion. Finally, clerosterol was shown to be a precursor of cyasterone and isocyasterone.