Qualitative and quantitative analyses of juvenile hormone and ecdysteroids from the egg to the pupal molt in Trichoplusia ni.

A method was developed to determine in the same extract juvenile hormone and various types of ecdysteroids in precisely staged eggs and larvae of Trichoplusia ni. Ecdysteroids were tentatively identified on the basis of their retention time in ion suppression reversed-phase HPLC and their cross-reactivity with two relatively non-specific, complimentary antibodies, whereas juvenile hormone was identified using reversed-phase HPLC combined with Galleria bioassay. Freshly laid eggs contained low levels of immunoreactive ecdysteroids. Mid-polar ecdysteroids increased in the phase of segmentation (14-18 h) and 1st larval cuticle formation (36-44 h), when 20-hydroxyecdysone and 20,26-dihydroxyecdysone were found to be predominant. Only traces of ecdysone and 26-hydroxyecdysone were seen. Toward hatching ecdysteroids decreased and represented mainly compounds more polar than 20,26-dihydroxyecdysone. In larval development ecdysteroids were low at the beginning of the feeding phases, increased toward cessation of feeding, and reached highest levels 12-15 h before ecdysis. In feeding stages ecdysone and 20-hydroxyecdysone were predominant, whereas in molting stages they were seen together with 20,26-dihydroxyecdysone and 20-hydroxyecdysonoic acid. The juvenile hormone titer was very low in freshly laid eggs and was high (approximately 25 ng/g) in embryos at the stage of 1st larval cuticle formation and eye pigmentation. In eggs we tentatively identified juvenile hormones I and II, whereas in larval stages juvenile hormone II appeared to be the predominant or exclusive juvenile hormone. Its titer fluctuated rapidly and was high in early 1st-instar larvae and again before the molts into the 3rd, 4th, and 5th instar. Highest titers were reached concomitant with the peak in 20-hydroxyecdysone 12-15 h before ecdysis.