It has long been hypothesized that the oxidation of 7-dehydrocholesterol (7dC), made from dietary cholesterol (C), to 3-oxo-7dC (3-oxo-Δ5,7C) immediately precedes the unknown "Black Box" oxidations that lead to the formation of the first up-stream intermediate exhibiting the highly characteristic ecdysteroid structure of the steroid molting hormone of insects, crustaceans and some other arthropods. Perhaps rate-limiting and under the control of the prothoracicotropic hormone (PTTH), the biosynthesis of 3-oxo-7dC and its subsequent oxidative modifications have been difficult to study because of their apparent instability, i.e. no intermediates between 7dC and the diketol (3-oxo-25,22,2-trideoxyecdysone) have ever been observed or identified in insect prothoracic gland incubations with radiolabelled precursors. However, we show that 3-oxo-7dC can be converted into lipophilic, photosensitive, ketone-blocked (PSKB) ketal derivatives which will release 3-oxo-7dC when and where desired following brief irradiation with innocuous long-wave (365 nm) UV-light both in vivo and in vitro. In this manner, 3-oxo-7dC is quickly and efficiently incorporated into ecdysteroids by adult male and female Drosophila raised on a diet containing the PSKB ketals and in prothoracic glands of Manduca sexta incubated with the ketals emulsified into media. The instability of 3-oxo-7dC and its spontaneous transformation into extensively electron-delocalized intermediates will be discussed in relation to a possible mechanism of the Black Box oxidations eventually leading to the production of the active molting hormone 20-hydroxyecdysone (20E).
Bibliographical noteFunding Information:
We greatly appreciate the initial experimental suggestions of Dr. Bruce Hammock and Pavel Aronov (U. Cal. Davis) that ultimately led to the successful formulation of a photosensitive, ketone-protected precursor of the otherwise unstable 3-oxo-7dC molecule. Thanks are also given to Dr. Rene Lafont (Paris) and Dr. Huw Rees (Liverpool) for helpful readings of the manuscript. We also thank Susan Whitfield for graphics and Tory Sheets for technical assistance. This work was supported by a grant from the National Science Foundation which has helped us in our studies of ecdysteroid synthesis over many years (#IBN130825 to LIG and JTW).
- Black box
- Ketone-blocked ketal intermediates