CHAPTER LV 



THE PREPARATION AND PURIFICATION OF 

 TOXINS, TOXOIDS, AND ANTITOXINS 



EDWIN J. BANZHAF 



Bureau of Laboratories, New York City Department of Health 



SOME PROPERTIES OF BACTERIAL TOXINS 



Bacterial antigens are classified as extracellular and endocellular. Among the 

 , extracellular are the exotoxoids. The classical exotoxins are primarily toxic and re- 

 gardless of the amount injected are not immediately fatal but require an incubation 

 period before fatality ensues. The extracellular toxins produced by the pathogenic 

 anaerobes such as B. ivelchii {Clostridium wclchii) and B. oedematis nialigni (Clostri- 

 dium oedcmatis) would not under this classification be classed with these exotoxins 

 in that they produce almost immediate symptoms even though a small dose be in- 

 jected intravenously. They are, however, toxins, and the older definition should be 

 revised to the simple one that toxins are antigenic poisons (Coca); i.e., toxins possess 

 the property of stimulating the production of antitoxins which are specific and which 

 will neutralize the toxins in vitro and in vivo. The endocellular substances possess 

 the property of stimulating the production of agglutinins, precipitins, opsonins, or 

 bacteriotropins and bacteriolytic and bactericidal antibodies. The actual existence 

 of specific endotoxins which are antigenic is still debatable. The best-known exo- 

 toxins are those produced by B. diphtheriae {Corynebacterium diphtheriae), B. tetani 

 {Clostridium tetani), B. botidinus {Clostridium botulimim), B. welchii {Clostridium 

 welchii) , B. anthracis symptomatici {Clostridium chauvei) , B. dysenteriae Shiga {Eberthella 

 dysenteriae), Streptococcus scarlatinae, and Streptococcus erysipelatus. 



Toxins are non-crystallizable and have not been obtained in a pure form. They 

 are precipitated by salts of the heavy metals, ammonium sulphate at saturation, 

 alcohol, nucleic acid, and acetic acid. They are considered by Oppenheim as large 

 molecular complexes, probably related to the proteins and classed as "enzymes." 

 They are electropositive, and are not readily dialyzable except through thin mem- 

 branes. They are readily absorbed by animal charcoal and are destroyed by pro- 

 teolytic ferments. Sodium chloride at saturation destroys the toxins of B. diphtheriae 

 and B. tetani. Freezing also destroys about half of these toxins. Toxins are sensitive 

 to oxidation, light, and Roentgen rays. Heating B. diphtheriae and B. tetani toxins 

 to 50° C. for twelve hours destroys about half of the acutely toxic fraction (toxophore, 

 Ehrlich's hypothesis) and modifies the rest into toxoid fraction (haptophore, Ehrhch). 

 The toxoids are capable of neutralizing antitoxins and are antigenic. Heating to 

 56° C. for two hours and to 58° C. for one-half hour produces practically the same 

 results as heating to 50° C. for twelve hours. A notable exception to the lability of 

 the toxins to high dilution and heat is the streptococcus scarlatinal toxin. This toxin 

 is practically stable for years, and even in high dilutions holds its value for at least 



745 



