Profiling of ecdysteroids in complex biological samples using liquid chromatography/ion trap mass spectrometry.
A sensitive method using high-performance liquid chromatography coupled to a mass spectrometer with electrospray ionization source (HPLC/ESI-MS) was developed for detection of ecdysteroids in biological samples. We report here for the first time that ecdysteroids can be classified into three groups based on ESI full-scan mass spectra: group 1 (ecdysone (E), 2-deoxyecdysone (2dE), 2,22-dideoxyecdysone (3beta5beta-KT), and 3alpha5alpha[H]-dihydroxycholest-7-en-6-one (3alpha5alpha-KD)), in which loss of one molecule of water from the protonated molecular ion ([M+H](+)) represents the dominant ion; group 2 (20-hydroxyecdysone (20E), makisterone A (MakA), 3beta5beta-KD, and 3beta5alpha-KD), in which [M+H](+) is a major ion but some water loss is observed; and group 3 (muristerone A (MurA) and ponasterone A (PonA)), in which [M+H](+) is the dominant ion with no water loss observed. Based on the analytical procedure in combination with structural information from the group classification and with the application of source-induced dissociation, we identified free ecdysteroids in biological samples: 20,26-dihydroxyecdysone and ecdysonic acid in the larval hemolymph, and the progressive metabolism of 26-hydroxyecdysone (26E) to 3alpha-26E from day-1 to day-3 embryos of the tobacco hornworm Manduca sexta.
Li
Yutai
Y
Department of Environmental Sciences and Engineering, School of Public Health, The University of North Carolina at Chapel Hill, NC 27599, USA. yutai_li@merck.com
Warren
James T
JT
Boysen
Gunnar
G
Gilbert
Lawrence I
LI
Gold
Avram
A
Sangaiah
Ramiah
R
Ball
Louise M
LM
Swenberg
James A
JA
D000818:Animals; D002853:Chromatography, Liquid / Q000379:methods*; D026461:Ecdysteroids / Q000032:analysis* / Q000378:metabolism*; D020869:Gene Expression Profiling; D006458:Hemolymph / Q000378:metabolism*; D007814:Larva / Q000378:metabolism; D018402:Manduca / Q000196:embryology* / Q000378:metabolism*; D008832:Microchemistry / Q000379:methods; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
D026461:Ecdysteroids