Quantification of neurosteroids in rat plasma and brain following swim stress and allopregnanolone administration using negative chemical ionization gas chromatography/mass spectrometry
Analytical Biochemistry. 2000-12-01; 287(1): 153-166
DOI: 10.1006/abio.2000.4841
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Vallée M(1), Rivera JD, Koob GF, Purdy RH, Fitzgerald RL.
Author information:
(1)Department of Neuropharmacology, Scripps Research Institute, 10550 North
Torrey Pines Road, La Jolla, California 92037, USA.
A simplified method for the quantitative analysis of neurosteroids in rat plasma
and brain is described. The method uses negative chemical ionization gas
chromatography/mass spectrometry and involves the synthesis of
pentafluorobenzyloxime/trimethylsilyl ether derivatives with excellent
chromatographic and electron-capturing properties. Deuterium-labeled analogs of
the steroids of interest were synthesized and used as internal standards. The
steroids (allopregnanolone, epiallopregnanolone, pregnenolone, testosterone, and
dehydroepiandrosterone) were isolated from the plasma or brain matrix by a rapid
and straightforward solid-phase extraction procedure. The mass spectrometer was
operated in a selective ion monitoring mode, allowing for picograms of
neurosteroids to be quantified from biological extracts. The method was linear
(typical R(2) = 0.999) over the concentration range (100 to 8000 pg from 0.3 ml
plasma and 250 to 8000 pg from 100 mg brain tissue) with good precision and
accuracy. In experimental protocols, the procedure was suitable for measuring
concentrations of endogenous neurosteroids in rat plasma and brain. Significant
elevations (P < 0.001) were observed in the frontal cortex for allopregnanolone
and pregnenolone following a swim stress and for allopregnanolone and
epiallopregnanolone following allopregnanolone injection (8 mg/kg, sc). The
present method allows accurate determination of neurosteroids and will be
helpful in elucidating the role of neurosteroids in health and disease.
Copyright 2000 Academic Press.
DOI: 10.1006/abio.2000.4841
PMID: 11078595 [Indexed for MEDLINE]