Scaled-up production and purification of Clostridium perfringens type A enterotoxin.
Methods for small-scale production of Clostridium perfringens type A enterotoxin were unsuitable for large-scale culture of this organism. Rapid, efficient harvesting of 40 1 batch culture of Cl. perfringens was achieved by tangential flow micro-filtration with the Millipore Pellicon cassette system. Enterotoxin-containing extracts were prepared by passing concentrated suspensions of the harvested cells through a French pressure cell. The overall yield of purified enterotoxin was 38.8%. The toxin gave a single band on native polyacrylamide gels but formed high molecular weight aggregates in the presence of sodium dodecyl sulphate. These aggregates frequently occurred during storage of non-sterile enterotoxin preparations but could be separated from the monomer toxin by gel filtration on Sephadex G-100. Purified monomer enterotoxin had biological activities of 119.3 micrograms/kg mouse lethal dose when injected intraperitoneally and 3333 capillary permeability increasing units/mg protein in guinea pig skin. Thirty micrograms of the enterotoxin caused fluid accumulation in ligated rabbit ileal loops. Aggregated enterotoxin had no demonstrable biological or immunological activity.
D000818:Animals; D002850:Chromatography, Gel; D003016:Clostridium perfringens / Q000378:metabolism* / Q000502:physiology; D004591:Electrophoresis, Polyacrylamide Gel; D004768:Enterotoxins / Q000096:biosynthesis / Q000302:isolation & purification* / Q000633:toxicity; D006168:Guinea Pigs; D051379:Mice; D008807:Mice, Inbred BALB C; D011817:Rabbits; D013171:Spores, Bacterial
D004768:Enterotoxins; C035598:enterotoxin, Clostridium