Purification and some physicochemical properties of slow-reacting substance of anaphylaxis (SRS-A) from sensitized guinea pig lung.

Abstract

AbstractThe slow‐reacting substance of anaphylaxis (SRS‐A) generated by antigen challenge of sensitized guinea pig lung fragments was partially purified and the physicochemical properties of this activity were studied. The SRS‐A recovered from antigen challenged lung preparations of 600 animals was used for the purification procedure. Treatment with organic solvents, extraction with 80% ethanol, Sephadex LH‐20 chromatography with 80% ethanol, and DEAE‐Sephadex A‐25 chromatography in 60% methanol eluted with 0.0 to 0.1 M NaCl in 60% methanol was the purification sequence finally adopted. Overall recovery of SRS‐A bioactivity was 60% with a specific activity of 2.52 units/ng of dry weight. This represented a 1.67 million‐fold purification over the starting material. The DEAE‐Sephadex A‐25 step alone provided a 7600‐fold purification. This highly purified SRS‐A had an apparent molecular weight of 380 to 400 daltons. The bioactivity was acid labile and alkaline stable and was blocked by low concentrations of the SRS‐A antagonist FPL 55712. The SRS‐A was thermostable in aqueous media and displayed enhanced bioactivity after heating at 60 C for 60 min. These results indicate that we have developed a highly efficient new approach to the isolation of guinea pig SRS‐A, which also may be useful in the study of SRS‐A from other tissues or species. The physicochemical properties of guinea pig SRS‐A appear to be very similar to those of SRS‐A from other species.