PARTIAL-PURIFICATION OF A TOXIN FOUND IN HAMSTERS WITH ANTIBIOTIC-ASSOCIATED COLITIS - REVERSIBLE BINDING OF THE TOXIN BY CHOLESTYRAMINE

  • 1 January 1979
    • journal article
    • research article
    • Vol. 76  (3) , 468-476
Abstract
A toxin with cytotoxic and enterotoxic activities was isolated from cecal contents of hamsters receiving lincomycin. The toxin was partially purified by ultracentrifugation, ultrafiltration, (NH4)2SO4 precipitation and gel filtration. Cytotoxic activity, assayed on monolayers of HeLa cells, was restricted to material that eluted in the MW range of 107,000 .+-. 6000 daltons. Cytotoxicity of crude AAC [antibiotic-associated colitis] toxin could be demonstrated at concentrations as low as 0.04 .mu.g/ml. The toxin was heat labile (55.degree.-60.degree. C for 0.5 h) and sensitive to trypsinization, acidification at pH 3, or alkalinization at pH 9. Cytotoxic activity was inhibited by Clostridium sordellii antitoxin. Enterotoxic activity of the crude toxin and the cytotoxic fraction from gel filtration was demonstrated by fluid secretion in ligated rabbit ileal loops. Studies were done in vitro with cholestyramine resin, vancomycin or gentamicin to determine if the toxin was bound or denatured by these drugs. Cholestyramine apparently bound the toxin, significantly reducing its cytotoxicity. Reversible binding of the cytotoxic material was demonstrated by salt gradient elution. Neither vancomycin nor gentamicin had any effect on the in vitro cytotoxic activity of the toxin.

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