MATERIALS. 



39 



Normal 



b. Adjust the acidity to 1.5 per cent, as follows: 



Measure out 10 c.c. of the mixture, placing it in an evaporating dish 

 with ten times its bulk of water. Bring to a boil and add a few drops of 

 phenolphthalein solution.* This solution is colorless so long as it is acid, 

 but turns red when alkaline. The agar medium remains red after the 

 addition of phenolphthalein, showing it to be acid. 



Place the evaporating dish under a 

 burette containing i/io normal NaOH 

 (one part normal NaOH diluted with 

 nine parts of distilled water) (Fig. 57). 

 Take the reading on the burette. Allow 

 the solution to fall drop by drop into the 

 evaporating dish, stirring between each 

 drop. As long as the material remains 

 acid no color will appear. Continue 

 adding the NaOH until a faint red color 

 appears and does not disappear upon 

 stirring. When this point is reached 

 the contents of the evaporating dish 

 are neutral. 



Take the reading upon the burette 

 and determine the difference between 

 the first and second reading. The 

 difference gives the number of cubic 

 centimeters of i/io normal NaOH 

 needed to neutralize 10 c.c. of the cul- 

 ture medium. Divide by 10 to de- 

 termine the amount necessary to neu- 

 tralize one c.c., and multiply by 990 to 

 determine the amount necessary to 

 neutralize the rest of the agar medium. 



Add to the agar medium sufficient 

 NaOH to neutralize it. Instead of 

 adding i /io normal, add normal NaOH, 

 using of course only i/io as many 

 cubic centimeters as the above calcula- 

 tion shows would be needed of i/io 

 normal NaOH. This will bring the 

 whole to the neutral point. 



Add normal HC1 to the amount of 

 15 c.c. per liter. In this case, there being only 990 c.c. left, the amount 

 added should be 14.9 c.c. This makes an acidity of 1.5 per cent., the best 

 reaction for most bacteria. 



c. Boil vigorously for fifteen minutes and then restore the original 

 weight by adding water. Test the reaction again as before to see if it is 



FIG. 57 Two burettes arranged for 

 neutralizing culture media. 



* Eight per cent, phenolphthalein in 50 per cent, alcohol. 



