HYDROGEN ION CONCENTRATION AND PROTEOLYSIS. 155 



Thus, the proteolysis can be followed from time to time. 



In applying this method in the present investigation it may be clearly 

 seen from the preceding discussions that the proteolysis can be followed 

 accurately by determining the difference between the amount of amino 

 acid present in the original culture medium and after the growth of an 

 organism in it. It is, however, necessary to exercise special care in regard 

 to the presence of ammonia and carbon dioxide, produced in the course 

 of deamidation, which interferes with the determination. To avoid these 

 sources of error ammonia and carbon dioxide-free air was passed through 

 the media, which were made alkaline by the addition of n/1 NaOH pre- 

 vious to determhiation. 



Preparation of Media. 

 Steps in Selection oj Medium. 

 Common experiences dictate that the properties of media are influenced 

 by constituents and technic of preparation. To obtain stable and com- 

 parable data is essential in these investigations from the standpoint of 

 biology, chemistry and physics because a very slight variation will fur- 

 nish unsatisfactory results. Several media were carefully tested. 



Medium I.: — 

 1,000 grams chopped lean beef. 

 1 ,000 c. c. distilled water. 



Medium II.: — 

 1 per cent. Liebig's meat extract. 

 1,000 CO. distilled water. 



Medium III.: — 

 1 per cent. Witte's peptone. 

 0.5 per cent. NaCl. 

 1,000 c. c. distilled water. 



Medium IV. : — 

 1 per cent. Liebig's meat extract. 

 1 per cent. Witte's peptone. 

 0.5 per cent. NaCl. 

 1,000 c. c. distilled water. 



The method of preparation and testing is outlined below. 



Medium I. — Five hundred grams of chopped lean beef were obtained 

 from a local meat market and immediately well mixed. Ten lots of 30 

 grams each were weighed out and each lot was transferred into a 250 

 cubic centimeter Erlenmeyer flask. These flasks were marked a, b, c, d, 

 e, /, g, h, i,j, k and I, and divided into groups, one a to^, inclusive, marked 

 A, and the other, g to /, inclusive, marked B. Group A was treated directly, 

 while group B was placed in an incubator at 37° C. for twenty-four hours. 

 After an addition of 300 cubic centimeters of distilled water to the former 

 the flasks were shaken for thirty minutes. At the end of this period the 



