Spectrophotometric monitoring of lipoxygenase and cyclooxygenase pathway activity using ionophore stimulated whole blood.
We have employed clotting human blood stimulated with ionophore to develop a system for measuring cyclooxygenase, 5-lipoxygenase, and 12-lipoxygenase pathway products released into the serum fraction. In a single chromatographic run, 5-HETE, 12-HETE, 12-OH-heptadecatrienoic acid (HHT), LTB4, 20-OH-LTB4, and 20-COOH-LTB4 are quantitated by UV monitoring after separation by HPLC. The kinetics of product formation/release of all fatty acid products into the serum show an apparent lag of approximately 2 min, after which time the amounts of HHT, 5-HETE, and LTB4, respectively, plateau at 10 min while 12-HETE increases over a 60 min period. The system is responsive to standard cyclooxygenase and lipoxygenase inhibitors, and is of value of evaluating prospective blockers of AA metabolism in a whole blood setting.
D000001:Calcimycin / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D005231:Fatty Acids, Unsaturated / Q000097:blood; D006801:Humans; D066298:In Vitro Techniques; D008084:Lipoxygenase / Q000097:blood*; D011451:Prostaglandin-Endoperoxide Synthases / Q000097:blood*; D013056:Spectrophotometry, Ultraviolet
D005231:Fatty Acids, Unsaturated; D000001:Calcimycin; D008084:Lipoxygenase; D011451:Prostaglandin-Endoperoxide Synthases