Girard derivatization for LC-MS/MS profiling of endogenous ecdysteroids in Drosophila.

Abstract:

:Ecdysteroids are potent developmental regulators that control molting, reproduction, and stress response in arthropods. In developing larvae, picogram quantities of individual ecdysteroids and their conjugated forms are present along with milligrams of structural and energy storage lipids. To enhance the specificity and sensitivity of ecdysteroid detection, we targeted the 6-ketone group, which is common to all ecdysteroids, with Girard reagents. Unlike other ketosteroids, during the reaction, Girard hydrazones of ecdysteroids eliminated the C14-hydroxyl group, creating an additional C14-C15 double bond. Dehydrated hydrazones of endogenous ecdysteroids were detected by LC-MS/MS in the multiple reaction monitoring (MRM) mode using two mass transitions: one relied upon neutral loss of a quaternary amine from the Girard T moiety; another complementary transition followed neutral loss of the hydrocarbon chain upon C20-C27 cleavage. We further demonstrated that a combination of Girard derivatization and LC-MS/MS enabled unequivocal detection of three major endogenous hormones at the picogram level in an extract from a single Drosophila pupa.

journal_name

J Lipid Res

authors

Lavrynenko O,Nedielkov R,Möller HM,Shevchenko A

doi

10.1194/jlr.D035949

subject

Has Abstract

pub_date

2013-08-01 00:00:00

pages

2265-72

issue

8

eissn

0022-2275

issn

1539-7262

pii

jlr.D035949

journal_volume

54

pub_type

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