502 THE BIOCHEMISTRY OF B VITAMINS 



from 1 at a pH range of 9.2-7.55 to 1,200 at pH 3.65, but then decreases 

 to 380 at pH 2.25. These values corrected for the relative activity of 

 sulfanilamide give 1, 245 and 76, respectively, for the relative activities 

 of p-aminobenzoic acid. Since the point of maximum activity corresponds 

 closely to the isoelectric point of p-aminobenzoic acid, it was proposed 

 that the undissociated molecules penetrate into the cell at a greater rate 

 than the ions. 



For Escherichia coli and Aspergillus niger, sulfanilamide was less 

 effective in acidic than in neutral media; however, the variation with 

 Escherichia coli was very slight and only fivefold for Aspergillus niger. 198 

 Consequently, any theory must account for such small variations. 



Some degree of correlation of the inhibitory activity of sulfonamides 

 with their ability to dissociate to sulfonamide ions was subsequently 

 noted. 199 - 200 The ratios of the concentration of sulfanilamide, sulfa- 

 pyridine, sulfathiazole or sulfadiazine to that of p-aminobenzoic acid at 

 which inhibition of growth of Escherichia coli becomes apparent are ap- 

 proximately 500, 40, 8 and 8, respectively, at pH 7. However, these ratios 

 calculated on the basis of the ratio of sulfonamide anion to p-amino- 

 benzoate, are 1.4, 1.4, 4.9 and 6.4, respectively. 199 Approximately eight 

 times as much sulfanilamide is required to inhibit the growth of Esche- 

 richia coli at pH 6.8 than at pH 7.8. 199 At pH 9, there is comparatively 

 little difference in the activity of sulfathiazole and sulfanilamide. 200 Since 

 an increase in the pH of the culture medium favors ionization of the 

 sulfonamides, the increased effectiveness of sulfonamides with increased 

 pH of the medium and the similar activities of certain sulfonamide ions 

 led to the consideration that the anion is the active form of the ionizable 

 sulfonamides. 199 ' 200 



Subsequently, it was observed that the activity of the sulfonamides 

 increased with pH of the culture medium only up to the point where 

 ionization is approximately half complete. 124 - 201 Actual decreases in 

 bacteriostatic activity are observed with highly ionized sulfonamides 

 such as sulfadiazine, when the pH is increased above the point of 50 per 

 cent ionization. Thus, when the pH of the medium exceeds the pK a of 

 the sulfonamide, the activity begins to decline. For sulfadiazine, an 

 increase of approximately 8.8 fold in concentration is required for bac- 

 teriostasis when the pH is increased from 6.5, where the sulfonamide is 

 approximately 50 per cent ionized, to 8.9 where the sulfadiazine is ap- 

 proximately 99.6 per cent ionized. 



These results led to the suggestion that only the molecular form pene- 

 trated the cell wall of bacteria while only the ionic form combines with 

 the enzyme which is inhibited. 201 



The decreasing ability of p-aminobenzoic acid (pK a 4.68) to prevent 



