Improved methods for purification of an enterotoxin produced by Bacillus cereus

An enterotoxin (ET) produced by Bacillus cereus was purified by ammonium sulfate precipitation followed by chromatography on SP-Sephadex C-25, chromatofocusing, and gel filtration on Sephacryl S-300. Purified ET was electrophoretically homogeneous with a molecular mass of 45,000 and with an isoelectric point of 5.5. It showed vascular permeability activity in rabbits, was lethal to mice, and caused fluid accumulation in mouse ligated intestinal loops. It showed cytotoxicity to Vero cells, but did not show any hemolytic or phospholipase C activities. These biological activities were found to be easily inactivated by heating at 56 degrees C for 5 min, by digestion with trypsin and pepsin, and by exposure to pH below 3.0 or higher than 11.