[Vol. 6 



34 ANNALS OF THE MISSOURI BOTANICAL GARDEN 



of potassium as .2 gni. K2HPO4. The following medium was 

 therefore prepared: 



Dextrose 



20 gms. 



Calcium glycerophosphate 0.2 gm. 



Magnesium sulphate 0.2 gm. 



Sodium chloride 0.2 gm. 



Potassium sulphate 0.2 gm. 



Distilled water 1000 cc. 



10% ferric chloride 3 drops 



No precipitate formed on heating the medium. It was fil- 

 tered, however, to remove the few particles of foreign mate- 

 rial which most likely were introduced in the dextrose. The 

 perfectly clear medium then was found to give a P H value of 

 about 6.7 as measured with the aid of standards and buffer 

 solutions recommended by Clark and Lubs ('17). Duplicate 

 5-cc. portions were titrated with N/50 alkali and phenolphtha- 

 lein and the computed amount (9.00 cc.) of N/50 alkali neces- 

 sary to bring an aliquot of the medium to P H 8. After the ad- 

 dition of this amount of alkali the P H value was found to be a 

 trifle less than 8. 



Fifty-cc. portions of this adjusted medium were then pi- 

 petted into each of twelve 700-cc. "Nonsol" Erlenmeyers. To 

 6 of these was then added a pinch of calcium carbonate. 

 After sterilization by the intermittent method the cultures 

 were incubated for 7 days at 28-30° C. The plan of the ex- 

 periment is shown in table xi. The inoculum was from a 24- 

 hour 10-cc. mannite shaker culture prepared by heavy inocula- 

 tion from F17, 7 days old. One spiral of this culture was used 

 in seeding each flask. The results appear in table xi. 



The results indicate that the phosphorus in glycerophos- 

 phates is to some extent available for the growth of Azoto- 

 bacter, although it is barely possible that the glycerophos- 

 phate may have hydrolyzed in the faintly alkaline solutions 

 during sterilization. The medium, although adjusted in re- 

 action to practically P H 8, was not suitable for growth unless 

 CaC03 was added, practically no growth taking place in the 

 absence of CaCOa. The rotator proved beneficial in the pres- 

 ence of calcium carbonate. 



