New ecdysteroid conjugate: Isolation and identification of 26-hydroxyecdysone 26-phosphate from eggs of the tobacco hornworm,Manduca sexta (L.)

The major ecdysteroid conjugate present in eggs (48–64 h old) of the tobacco hornworm has been purified by XAD-2 chromatography, C18 SEP-PAK separations, and ion suppression reversed-phase high-performance liquid chromatography. Enzymatic hydrolysis of the conjugate with acid phosphatase from human seminal fluid gave 26-hydroxyecdysone. The conjugate was identified as 26-hydroxyecdysone 26-phosphate by nuclear magnetic resonance and fast atom bombardment mass spectrometry. This compound is also the major conjugate of newly laid eggs (0–1 h old) of the tobacco hornworm. The role for ecdysteroid conjugates is discussed.