159 



(5) The effect of the nitrates used seemed to be due more to the 

 hydrolysis of the nitrates themselves rather than to reactions taking 

 place between them and the agar and gelatin. 



(6) Reaction of media should be adjusted by titrations made at the 

 temperature at which they are to be used. 



The results of this test lead one to presume that if proper care was 

 used in selecting the chemicals to be used in culture media, the acidity 

 of bacteriologic culture media would rarely have to be neutralized. 



Evidence Drawn From Literature tn Support of Contention That 

 Hydrolyzable Substances Should Be Avoided. 



Anthony and Ekroth (6) give a table which shows the reaction of 

 different peptones when titrated at room and boiling temperatures with 

 phenolphthalein as indicator. The results show that the variations in 

 acidity of the different peptones are large but that the peptone having 

 the lowest acidity at room temperature also has the lowest at boiling- 

 temperature. Witte's peptone has been almost universally agreed upon 

 as the best and is it not fair to suppose that this is due to its freedom 

 from hydrolyzable material? 



The same authors found that "Leibig's Extract of Beef" does not 

 undergo the hydrolysis that homemade extracts do. They say, "This 

 stability is due probably to very prolonged heating in the preparation 

 of the beef extract itself." In other words the more stable the extract 

 the more reason for its use. 



Itano (5) working with the hydrogen electrode finally, after experi- 

 mentation, decided on a medium containing both "Leibig's extract" and 

 Witte's peptone. He found that if these constituents were sterilized 

 before mixing, i. e., if they were stabilized, "the medium prepared from 

 them maintained the figured Pii fairly constantly." 



Fellers (17) finds that soil bacteria prefer a very slightly acid, a 

 neutral or just alkaline media. 



Summarizing the results obtained by these recent workers and 

 realizing that the standard method of titrating media (12) gives too 

 high titrations and thereby low acidity of adjusted media, it seems 

 probable that bacteriologic media in most cases should be very slightly 

 acid or neutral in reaction. 



The following procedure which is based on results reported in Tables 



