84 V. H. YOUNG 



glucose-agar and were transferred regularly in order that fresh 

 spores might be available at all times during the course of the 

 experiments. 



All experiments were conducted in Jena glass containers and 

 all water, culture media, etc, were kept in Jena glass. In all 

 experiments, except one, Erlenmeyer flasks of 100 cg. capacity 

 were used for the cultures. In one experiment 250 cc. Erlen- 

 meyer flasks were used. All glassware was thoroughly cleansed 

 and rinsed several times in double distilled water. Great care 

 was taken to obtain distilled water free from all impurities. 



Inorganic salts used for culture media were those of Kahlbaum 

 and of Merck and were of a high degree of purity. Merck's 

 anhydrous glucose, maltose, soluble starch and inulin were em-' 

 ployed. In certain experiments the inulin used was prepared 

 according to methods described by Haas and Hill (1913, p. 114) 

 and appeared to be of a high degree of purity since it was homo- 

 geneous and gave almost no evidence of reducing power. Rock 

 candy crystals were used as a source of sucrose. 



The culture medium was that recommended by Kylin (1914) 

 using ammonium nitrate as a source of nitrogen as follows : 



• per cent 



Ammonium nitrate . 22 



Magnesium sulphate , . 06 



Potassium diphosphate 0.12 



Ferric chloride Trace 



For convenience in mixing these substances each salt was dis- 

 solved separately in a small amount of water. When needed for 

 use the required amounts were measured into a volumetric flask, 

 the carbohydrate added and the whole made up to volume. 



Culture flasks were then prepared by running in 25 cc. of 

 the medium from a burette and the solutions were sterilized 

 for three successive days in an Arnold steam sterilizer. This 

 method was substituted for sterilization with the autoclave in 

 order to prevent the hydrolysis of the inulin of the culture me- 

 dium. It was at first suspected that the use of acid phosphate in 

 the culture medium might result in the hydrolysis of the inulin 

 present but according to Dox (1910) this does not take place. 



