| Autor: |
Akira Honda, Kouwa Yamashita, Takashi Hara, Tadashi Ikegami, Teruo Miyazaki, Mutsumi Shirai, Guorong Xu, Mitsuteru Numazawa, Yasushi Matsuzaki |
| Jazyk: |
angličtina |
| Rok vydání: |
2009 |
| Předmět: |
|
| Zdroj: |
Journal of Lipid Research, Vol 50, Iss 2, Pp 350-357 (2009) |
| Druh dokumentu: |
article |
| ISSN: |
0022-2275 |
| DOI: |
10.1194/jlr.D800040-JLR200 |
| Popis: |
We describe a highly sensitive and specific method for the quantification of key regulatory oxysterols in biological samples. This method is based upon a stable isotope dilution technique by liquid chromatography-tandem mass spectrometry (LC-MS/MS). After alkaline hydrolysis of human serum (5 μl) or rat liver microsomes (1 mg protein), oxysterols were extracted, derivatized into picolinyl esters, and analyzed by LC-MS/MS using the electrospray ionization mode. The detection limits of the picolinyl esters of 4β-hydroxycholesterol, 7α-hydroxycholesterol, 22R-hydroxycholesterol, 24S-hydroxycholesterol, 25-hydroxycholesterol, 27-hydroxycholesterol, and 24S,25-epoxycholesterol were 2–10 fg (5–25 amol) on-column (signal-to-noise ratio = 3). Reproducibilities and recoveries of these oxysterols were validated according to one-way layout and polynomial equation, respectively. The variances between sample preparations and between measurements by this method were calculated to be 1.8% to 12.7% and 2.9% to 11.9%, respectively. The recovery experiments were performed using rat liver microsomes spiked with 0.05 ng to 12 ng of oxysterols, and recoveries of the oxysterols ranged from 86.7% to 107.3%, with a mean recovery of 100.6%. This method provides reproducible and reliable results for the quantification of oxysterols in small amounts of biological samples. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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