The Action of Urea and Some of its Derivatives on Bacteria
Authors: Weinstein Louis , McDonald Alice 
Published: January 2023

Summary and Discussion
The combination of non-bacteriostatic amounts of sulfonamide with quantities of either urea or urethane which had no antibacterial action resulted in a mixture which possessed a high degree of antimicrobial activity. It was impossible to ascertain from the data whether this phenomenon was additive or synergistic, but the latter appeared to be the most likely possibility because of the marked bacteriostasis produced by the mixing of concentrations which, separately, exerted no inhibitory effect on growth. Since the tests were carried out in infusion broth containing serum, the possibility arose that the carbamates potentiated the sulfonamides by antagonizing the inhibitors of the latter drugs present in the medium. Such an explanation did not seem to account for all the facts, since, as was pointed out in the studies on the inhibition of para-aminobenzoic acid by the carbamates, this effect was slight in degree and persisted for a short time only. The size of the inoculum was found to bear a direct relationship to the degree of bacteriostasis produced.
The addition of sulfonamide to a highly lethal concentration of urethane was found to accentuate somewhat the bactericidal effect of the carbamate on gram-negative bacteria although it was difficult to estimate the actual degree of increased effectiveness since the time required to kill these organisms with 10 per cent ethyl carbamate alone was very short. The time of exposure to 10 per cent urethane required to produce death of S. aureus was fairly long, 3 to 4 hours; the addition of sulfonamide shortened the bactericidal period appreciably. The mechanism of the increased lethal action exhibited by combinations of carbamate and sulfonamide was probably the same as that of the enhancement of the bacteriostatic effect by sulfanilamide. The stimulating action of sulfonamide, however, appeared to be more marked in arresting growth than in causing the death of cells.
The data obtained in the work reported here confirmed the observations of Tsuchiya and his co-workers and of Lee, Epstein, and Foley on the anti-para-aminobenzoic-acid effect of carbamates. This phenomenon, however, was limited sharply by the relative concentrations of PABA and urea or urethane. It was found that the inactivation of the sulfonamide-inhibitor by urea or its ethyl derivative occurred only with in a narrow range of concentration of para-amino-benzoic acid and persisted for only a relatively short time. There was a direct relationship between the amount of carbamate employed and the quantity of PABA which it antagonized. Urethane appeared to be about four times more potent than urea in this respect.
The experiments of Kirby which did not confirm those of Tsuchiya et al and of Lee, Epstein, and Foley were not comparable for several reasons. In the first place, as pointed out by Lee et al, Kirby used a different medium than that employed by Tsuchiya and his group. An even more important reason for the inability of Kirby to demonstrate inactivation of para-aminobenzoic acid by urea would appear to be the fact that the quantity of PABA used was too large for the concentration of carbamide with which it was combined. Since a direct quantitative relationship between carbamate and para-amino-benzoic acid must be maintained if inhibition of the latter is to occur, as shown by the data presented in this paper, the reason for the discrepancy of Kirby's results becomes apparent.
It is doubtful whether the inhibition of para-aminobenzoic acid by urea or urethane was very important in the observed potentiation of sulfonamide because the amount of PABA inactivated was very small and the duration of the effect short. For this reason, the good clinical results obtained with combinations of carbamide and sulfonamide probably are not completely explainable on the basis of PABA neutralization alone. Since large amounts of urea and urethane, 100 per cent and 10 per cent concentrations, respectively, have been used clinically, large amounts of para-aminobenzoic acid are probably inhibited, but the antimicrobial activity of these concentrations of the carbamates is so great that it is impossible to ascertain just what role PABA inactivation plays in the good results obtained in the treatment of infections with combinations of sulfonamide and urea or urethane. For that matter, the question may also be raised as to whether or not the addition of sulfonamide to urea or urethane is of great value in the treatment of clinical infections since the concentrations of the carbamates used are usually sufficiently high to kill most species of gram-negative organisms and to inhibit growth and, in some instances, to produce death of S. aureus.
The effect of urea in increasing the solubility of sulfonamide was pointed out by Holder and MacKay and by Sober. Studies of urethane showed that a 10 per cent of this drug increased the solubility of sulfanilamide at least two and of sulfathiazole three times. The increase in solubility of the sulfonamides would probably have been even greater in solutions containing larger quantities of ethyl carbamate.