88 MANUAL OF MICROBIOLOGICAL METHODS 



quently used for this purpose because the first appearance of its pink 

 color, at about pH 8.5, is a convenient and usually sufficiently accurate 

 indication of the end point of such a titration. 



In fact, except for refinements that may be neglected for ordinary pur- 

 poses, pH 8.5, detectable by means of phenolphthalein, is a fairly satis- 

 factory end point for the titration of strong acids and of all weak acids 

 with pK' values of less than 6.0. In the case of acids with pK' values 

 greater than 6.0, it is necessary, by application of Eq. (5), to calculate the 

 pH of the equivalence point, and to refine the method of end-point 

 determination. For a discussion of the elementary theory of acid-base 

 titration, see Clark (1928). 



Titratable acidity of a culture. The titration of an acid (or a base) to 

 an equivalence point, as discussed above, is a rational application of 

 simple acid-base theory. On the other hand, in the titration of complex 

 mixtures such as milk, tissue extract, or culture media, an equivalence 

 point has no precise meaning. In such a case, the selection of an end- 

 point pH is arbitrary, and fixed by custom (e.g., pH 8.5 with phenol- 

 phthalein) or by some special requirement. 



In bacteriology, there is frequent need for determining the so-called 

 titratable acidity produced during the growth of a culture in a fluid 

 medium. To do this, it is necessary first to select a base line — that is, a 

 pH number which is to be used as an end point in the titration and for the 

 selection of an appropriate acid-base indicator. In the absence of special 

 criteria, it is reasonable to choose as a base line the pH of the uninocu- 

 lated medium. The selection of pH 7 as a base line may be acceptable, 

 because many bacteria grow optimally in this region, not necessarily 

 because it represents the pH of theoretical ''neutrality." Other base 

 lines may be chosen in accordance with the special requirements for which 

 the titration is to be made. 



The titratable acidity of the culture can be measured by titration of a 

 known volume of the fluid with O.XN NaOH to the predetermined end 

 point as shown by a standardized glass electrode or by the color of a suit- 

 able indicator. In the latter case, it is necessary to prepare for com- 

 parison an appropriate color standard representing the pH of the chosen 

 end point (see earlier discussion of the essential requirements for adequate 

 color comparison) . If the end-point pH is other than that of the uninocu- 

 lated control, a titration is made of the latter and its titration value is 

 subtracted algebraically as a correction, or ''blank," from that of the 

 culture. The result is usually recorded as millihters of 0.1 normal acid 

 per 100 ml of the culture fluid. If the culture produces an alkaline 

 reaction, the titration is performed with 0.1 A^HCl and recorded after 

 correction, if any, in the same way but as a minus quantity of titratable 

 acid. Special precautions are necessary if the titratable acidity is to 



