ANTI-ENZYME IMMUNITY 239 



exercises: (a) a powerful hydrolytic action on cartilage, chondroitin 

 sulfuric acid, polysaccharide containing sulfuric acid and pig-skin poly- 

 saccharide containing sulfur whose hydrolysis parallels that exercised 

 against hyaluronic acid which is free from the sulfuric acid group, and 

 (b) no action on heparin and mucoitin sulfuric acid, indicates that the 

 specificity of this enzyme is directly related to the specific structure of 

 the polysaccharide moiety of the above mentioned active substrates of 

 dissimilar composition. For a review on the chemical composition of 

 these substrates the reader is referred to an article by Meyer (1947). 



a. Quantitative Methods of Measuring Hyaluronidase Activity. 

 For the measurement of the activity of hyaluronidases several methods 

 have been employed : 



Mucin Clot Prevention Method. Seastone (1939) found that strep- 

 tococcal capsular polysaccharide when mixed with serum protein, gives 

 a typical mucin clot in the presence of acetic acid. After the treatment 

 of polysaccharide with hyaluronidase (mucinase), however, this reac- 

 tion no longer takes place (Robertson, Ropes and Bauer, 1940). In an 

 attempt to correlate hyaluronidase activity quantitatively with diffusing 

 activity, McClean (1943) described a method which depends upon 

 destruction by the enzyme of the power of a substrate-protein complex 

 to form a typical fibrinous clot on the addition of acetic acid. After 

 incubation with the specific enzyme the quantity of the clot is reduced 

 and the character of the precipitate changes from a fibrous to a floc- 

 culent precipitate, yielding finally a clear solution. McClean reported 

 that the destruction of the clotting power of the substrate appears to 

 develop in an early stage of its degradation, this loss being detectable 

 before any appreciable fall in viscosity occurs. The test is positive only 

 with crude hyaluronate and when carried out in the native fluid or 

 with the isolated and redissolved protein salt. Pure hyaluronate pre- 

 cipitates protein in acidic solution in flocculent form as contrasted with 

 the fibrous clot obtained with crude hyaluronate-protein complex. 

 Meyer (1947) failed to find a correlation between mucin clot preven- 

 tion units and viscosity reducing units, or a constant trend associated 

 with the potency or the source of the enzyme. 



Turhidimetric Method. Kass and Seastone (1944), in their study of 

 the relation of hyaluronidase activity and virulence of hemolytic strep- 

 tococcus, developed a turhidimetric method of determinina the activity 

 and the concentration of the enzyme of bacterial origin. The method 



