A precursor role of arachidonic acid for slow reacting substance of anaphylaxis (SRS-A) in the guinea pig

Abstract

A possible precursor role of arachidonic acid for slow reacting substance of anaphylaxis (SRS-A) was examined. SRS-A formed from sensitized guinea pig lung fragments stimulated with specific antigen under the coexistence of indomethacin and 14C-radiolabeled arachidonic acid, was sequentially purified by 80% ethanol extraction, adsorption chromatography of Sephadex LH-20, acidic organic solvent extraction, ion exchange chromatography of DEAE-Sephadex A 25, acidic organic solvent extraction and partition chromatography of Sephadex LH-20. SRS-A bioactive peak from the final partition chromatography step closely corresponded to one of the peaks of radioactivity. When this radiolabeled, highly purified SRS-A was applied to high performance liquid chromatography (HPLC), bio-and radioactivities were completely cochromatographed. Furthermore, the very close correspondence of bioactivity and radioactivity on HPLC of SRS-A partially inactivated by heating in acidic condition was also observed, indicating the evidence that arachidonic acid is a metabolic precursor of SRS-A. The chemical analysis of highly purified SRS-A including chemical degeradation, amino acid analysis and inactivation by soybean lipoxygenase showed that the SRS-A structure is C20 fatty acid oxygenated at the 5 position, having double bond at the 7, 9, 11 and 14 positions and probably containing cysteinyl-glycine at the b position. The marked chemical similarities and the very similar chromatographic behavior of SRS-A to one of the forms of SRS from rat basophilic leukemia (RBL-1) cells stimulated with Ca ionophore A 23187 suggested that both species have the identical structure.