Rapid quantification of 11 prostanoids by combined capillary column gas chromatography and negative ion chemical ionization mass spectrometry: application to prostanoids released from normal human embryonic lung fibroblasts WI38 in a culture medium.
The rapid and simultaneous quantification of 11 prostanoids has been carried out with a short-capillary gas chromatograph and negative ion chemical ionization (ammonia) mass spectrometer. The methoxime-trimethylsilyl ether-pentafluorobenzyl esters (MO-TMS-PFB) of nine prostanoids, PGA1, PGA2, PGB1, PGB2, PGD2, PGE1, PGE2, 6-oxo-PGF1 alpha and TXB2 and the TMS-PFB of two prostanoids, PGF1 alpha and PGF2 alpha, were separated in less than 5.5 min on a bonded OV-1 capillary column 0.25 mm i.d. x 6 m (0.15 micron thickness) using hydrogen as a carrier gas. PGD2, PGE2, PGF2 alpha, 6-oxo-PGF1 alpha and TXB2 were quantified up to 2.5 fmol injected (0.1 pmol derivatized) and both PGA2 and PGB2 up to 25 fmol injected (1 pmol derivatized). In order to maintain the stability of the prostanoids containing a carbonyl group, such as TXB2 during the purification and derivatization steps of biological materials, methyl acetate was used in place of methyl formate as an eluant for Sep-Pak C18 purification. Normal human embryonic lung fibroblasts W138 (5.63 x 10(5) cells in a log phase) produced: PGA2 15.28, PGB2 13.48, PGD2 7.95, PGE1 2.62, PGE2 177.76, PGF2 alpha 25.14, 6-oxo-PGF1 alpha 27.33 and TXB2 61.00 pmol in 10 ml of Eagle minimal essential medium.
Shindo
N
N
Division of Biochemical Analysis, Juntendo University, School of Medicine, Tokyo, Japan.
Saito
T
T
Murayama
K
K
D002138:Calibration; D002460:Cell Line; D004622:Embryo, Mammalian; D005347:Fibroblasts / Q000378:metabolism; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods; D006801:Humans; D008168:Lung / Q000166:cytology / Q000378:metabolism*; D011453:Prostaglandins / Q000032:analysis* / Q000096:biosynthesis; D012997:Solvents; D013929:Thromboxane B2 / Q000032:analysis / Q000096:biosynthesis
D011453:Prostaglandins; D012997:Solvents; D013929:Thromboxane B2