Thin-layer chromatographic in situ analysis of insect ecdysones via fluorescence-quenching

The possibility of quantitating insect ecdysones in situ on thin-layer chromatographic plates was examined. Two approaches were evaluated: 1) the induction of ecdysone fluorescence by sulfuric acid treatments and 2) the fluorescence-quenching of fluorescent thin-layer chromatographic plates by ecdysones. The fluorescence-quenching method was found to be most suitable and had a linear response range from 0.5 to 3 microgram for alpha-ecdysone and 20-hydroxyecdysone. Fluorescence-quenching and high pressure liquid chromatographic analyses obtained from extracts of alpha-ecdysone 20-hydroxylase incubations gave similar results. New data concerning the acid-induced fluorescence of ecdysones showed alpha-ecdysone to be twice as fluorescent as 20-hydroxyecdysone.