Flurbiprofen-sepharose chromatography of the prostaglandin synthetase from bovine seminal vesicles.
Flurbiprofen-Sepharose and Acetyl-Sepharose have been prepared by coupling dl-2-(2-fluoro-4-biphenylyl)propionic acid [Flurbiprofen] and acetic acid, respectively, to 3-(N-[3-aminopropyl)aminopropyl Sepharose 4B using a water soluble carbodiimide. The arachidonic acid oxygenase activity of solubilized bovine seminal vesicle microsomes is retarded during chromatography on Flurbiprofen-Sepharose but not Acetyl-Sepharose. Thus binding of the oxygenase to Flurbiprofen-Sepharose results from interaction with the immobilized inhibitor. However, the impure oxygenase is either not bound and/or not eluted in a biospecific manner since the abilities of flufenamic acid, R(+) and S(-)-5-cyclohexylindan-1-carboxylic acid, and R and S-Naproxen to remove the enzyme from Flurbiprofen-Sepharose do not parallel the relative efficacies of these compounds as prostaglandin synthesis inhibitors. Nevertheless, gradient elution of arachidonic acid oxygenase activity from Flurbiprofen-Sepharose with flufenamic acid provides a 15 fold enrichment of the enzyme from solubilized bovine seminal vesicle microsomes in 80% yield indicating that this chromatographic reagent can be a powerful tool for use in purification of the prostaglandin synthetase.
D000818:Animals; D001095:Arachidonic Acids / Q000378:metabolism; D002417:Cattle; D002845:Chromatography / Q000379:methods*; D005480:Flurbiprofen; D008297:Male; D008861:Microsomes / Q000201:enzymology; D006899:Mixed Function Oxygenases / Q000032:analysis*; D010105:Oxygenases / Q000378:metabolism; D011451:Prostaglandin-Endoperoxide Synthases / Q000032:analysis*; D012669:Seminal Vesicles / Q000201:enzymology*; D012685:Sepharose
D001095:Arachidonic Acids; D005480:Flurbiprofen; D012685:Sepharose; D006899:Mixed Function Oxygenases; D010105:Oxygenases; D011451:Prostaglandin-Endoperoxide Synthases