348 IMMUNO-CATALYSIS 



Py 8, 6 and c-toxins were found to exercise no effect upon human 

 serum or lecitho-vitellin, and antitoxins to group A, B, C and D, if 

 containing insufficient antitoxin to neutralize the a-toxin as judged 

 by the hemolytic or lethal testing, failed to prevent the reaction of 

 toxin with serum and with lecitho-vitellin. It thus appears that the 

 active principle in toxins causing the phenomenon of opalescence, etc., 

 in human sera, is contained in a-toxin and is neutralizable only by 

 a-anti toxin. 



Potassium oxalate, disodium phosphate and sodium citrate pre- 

 vented both hemolysis and opalescence, suggesting that ionized cal- 

 cium, which was found by MacFarlane and Knight (1941) to be an 

 accelerating factor of lecithinase activity, might play an essential 

 part in the reaction. The restoration of calcium in the system removed 

 by the above salts, led to the restitution of all of the in vitro activities 

 of the toxin. Thus it is clearly shown that hemolysis by a-toxin and 

 its reaction with serum and lecitho-vitellin, as described above, occur 

 only in the presence of ionized calcium. 



The above discussed phenomenon of opalescence produced by the 

 action of the a-toxin of CI. welchii on sera and lecitho-vitellin appears 

 to be identical with that observed in the experiments of MacFarlane 

 and Knight (1941) in which the action of a-toxin on clear egg-yolk 

 produced a turbidity which increased as the hydrolysis of lecithin pro- 

 gressed. Both studies show clearly that this toxin exercises a powerful 

 hydrolytic action on lecithin and lecitho-vitellin that not only splits 

 lecithin into a diglyceride and phosphocholine, but also causes the 

 splitting of lecithin from lecitho-vitellin and the "denaturation" of the 

 protein component. The splitting of lecithin from its conjugation with 

 the protein (serum or egg-yolk) appears to be due to a primary effect 

 of "denaturation" or "proteolysis" on the protein component by toxin. 

 Such an effect by toxin will naturally result in the rupture of the bonds 

 between lecithin and the protein in a manner similar to the splitting 

 of the prosthetic groups of conjugated proteins by denaturation or 

 proteolysis of the protein components (Sevag and Smolens, 1941). The 

 hydrolysis of lecithin by the lipase activity of toxin would represent, 

 therefore, a second stage in the occurrence of the phenomenon of 

 opalescence. The above assumption that toxin exercises a primary 

 effect of "denaturation" or proteolysis on the conjugated lipo-protein 

 or lecitho-vitellin of serum would seem to gain a certain degree of 



