[Determination of unconjugated neurosteroids in rat brain regions by liquid chromatography-negative atmospheric pressure ionization mass spectrometry].
OBJECTIVE
To simultaneously determine three unconjugated neurosteroids, dehydroepiandrosterone (DHEA) , pregnenolone (PREG), allopregnenolone (AP), from several brain regions of the rat.


METHODS
Neurosteroids were isolated separately in a two steps procedure by using ethyl acetate-n-hexane (90:10) as the first step to extract the unconjugated steroids, then the steroid fractions were further purified by SPE. All steroids were derivatized with 2-nitro-4-trifluoromethylphenylhydrazine (2-NFPH) and analyzed by HPLC-MS ( APCI) using selected-ion monitoring. Methyltestosterone was chosen as the internal standard. Results The linear calibration curve of DHEA was obtained in the concentration range of 0.030-2.00 microg x L(-1). The linear calibration curves of PREG and AP were obtained in the concentration range of 0.025-2.00 microg x L(-1). The concentrations of DHEA, PREG and AP in male rat brain regions were (0.70 +/- 0.23), (4.8 +/- 1.9), (1.1 +/- 0.6) ng x g(-1) for frontal cortex, (0.57 +/- 0.28), (6 +/- 3), (0.5 +/- 0.3) ng x g(-1) for hippocampus, (1.5 +/- 1.0), (9 +/- 5), (1.4 +/- 0.9) ng x g(-1) for amygdale, (0.52 +/- 0.14), (7.7 +/- 2.8), (0.5 +/- 0.6) ng x g(-1) for striatum, (2.9 +/- 1.6), (18 +/- 9), (1.6 +/- 1.3) ng x g(-1) for nucleus accumbens, (4.0 +/- 2.0), (27 +/- 12), (0.8 +/- 0.5) ng x g(-1) for pituitary gland, (1.7 +/- 1.2), ( 16 +/- 10), and (0. 8 +/- 0.7) ng x g(-1) for hypothalamus, respectively.


CONCLUSIONS
Good linearity and accuracy were observed for each steroid. The procedure was suitable for measuring concentrations of the unconjugated steroids in rat brain simultaneously.
Ren
Jin-min
JM
Bathune International Peace Hospital, Shijiazhuang 050082, China.
Hou
Yan-ning
YN
D000679:Amygdala / Q000737:chemistry; D000818:Animals; D001923:Brain Chemistry; D002851:Chromatography, High Pressure Liquid; D003342:Corpus Striatum / Q000737:chemistry; D003687:Dehydroepiandrosterone / Q000032:analysis*; D006624:Hippocampus / Q000737:chemistry; D007031:Hypothalamus / Q000737:chemistry; D008297:Male; D013058:Mass Spectrometry / Q000379:methods; D009714:Nucleus Accumbens / Q000737:chemistry; D017397:Prefrontal Cortex / Q000737:chemistry; D011284:Pregnenolone / Q000031:analogs & derivatives* / Q000032:analysis*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012680:Sensitivity and Specificity
D003687:Dehydroepiandrosterone; D011284:Pregnenolone