HYDROGEN-ION CONCENTRATION OF BROTH MEDIA 149 



Colorimetric determination of hydrogen-ion concentration 



In properly buffered solutions it is possible partially to elimi- 

 nate such factors as color and turbidity by diluting the test fluid 

 with water. Preliminary tests showed that with broth and cul- 

 tures it was possible to dilute Ice. of the material with 4 cc. of 

 distilled water without altering the hydrogen-ion concentration. 

 Accordingly this technic was employed in all the determinations. 

 Freshly boiled and cooled distilled water was used for diluting 

 as preliminary tests had shown that unboiled water gave slightly 

 lower Ph readings. The Ph of the water itself was usually found 

 to rise from 4.8 to 6.8 upon boiling, probably due to liberation of 

 carbon dioxide. To eliminate factors of color and turbidity more 

 completely Walpole's (1911) method of superposition was used by 

 employing the comparator block described by Dernby and Avery 

 (1918). All determinations were carried out at room tempera- 

 ture. The limit of error in the readings was 0.1 Ph. 



The adjustment of broth media 



One cubic centimeter of the broth was diluted with distilled 

 water (freshly boiled and cooled). Two acid solutions and two 

 basic solutions were kept on hand. They were n/1 HCl and an 

 3xact 1:10 dilution of the same; n/1 NaOH and an exact 1:10 

 dilution. A specially made micro burette, of 1 cc. capacity and 

 graduated to 0.01, contained the diluted acid or base. This 

 was added to the tube containing the medium, water, and 0.3 

 cc. of the proper indicator solution until the color produced therein 

 exactly matched that of the color standard of desired Ph. The 

 reading on the micro burette was then taken and by calculation 

 the amount of stock acid or base needed to adjust the total 

 amount of broth was determined. Following the addition of the 

 acid or base to the entire lot of medium a check determination 

 was always carried out. The broth was autoclaved at 15 pounds 

 for twenty minutes. In case this caused the formation of a pre- 

 cipitate the medium was filtered and subjected to a second auto- 

 claving for twenty to thirty minutes at 10 pounds pressure. The 

 low pressure prevents a second precipitation of the medium. The 



