268 JOURNAL OF THE SOCIETY OF cOSMETIC CHEMISTS The search for enzyme inhibitors appeals to investigators because it offers a means of preventing acid formation without necessarily disturbing the bacterial population. The numbers and types of organisms in saliva do not differ greatly When small or large amounts of sugar are being consumed by a patient. When sugar is not being consumed, very little acid is formed by the dental plaque. This reasoning suggests that acid formation is not essential to the vitality of the oral micro6rganisms. If it were possible to find a substance which would be adsorbed by one of the enzymes necessary for acid production and would serve to inactivate the enzyme, perhaps the metabolism of micro/3rganisms would proceed just as if sugar were not present. Because there are several enzymes which might be blocked by in- hibition, we have chosen to be empirical in our search for inhibitors. The empirical approach has been used successfully in other fields, such as for study of insecticides, antimalarial and antituberculosis drugs, and for cancer therapy. Our evaluation makes use of three criteria and eliminates or fails any compound which does not pass all three. The first requirement is that the substance be able to diffuse into a material which resembles dental plaque in some of its properties. The material employed is the sediment obtained by centrifuging saliva. The next requirement is that the substance be able to raise the pH of a synthetic dental plaque that is actively forming acid. The third requirement is that the inhibitory effect be retained for one hour Inhibitor Glucose Buffer AIk(31i Acid REACTIONS AFFECTING EQUILIBRIUM pH REACHED WITHIN SALIVA SEDIMENT SEDIMENT GLASS ELECTRO • Glucose j/•• CO 2 • H20 •' ..•- Figure 2.

ENZYME INHIBITORS FOR DENTIFRICES 269 or more after the test chemical has been replaced by a control solution. Re- tention is important since sodium bicarbonate is effective in raising the pH of plaques temporarily but the effect is transient as one might expect, bi- carbonate used as dentifrice does not seem to have an influence on dental caries (9). A sketch of the theory of our measurements is shown in Fig. 2. A meas- ured quantity of salivary sediment is placed against a glass electrode and held in place with a thimble of nylon mesh. The solution in contact with the sediment contains 0.2 molar glucose and 0.01 molar bicarbonate buffer. The glucose diffusing into the sediment is converted to acid by bacterial enzymes. The acid gradually accumulates, lowering the pH of the sediment in contact with the glass electrode. After a few minutes a steady state is reached, at which time the acid being formed is balanced by loss through diffusion of acid to the surrounding buffer and by titration of acid by buffer diffusing inward. At this time the pH differential between the salivary sediment and the surrounding solution is an index of acid formation rate. When a test solution is added to the bicarbonate-glucose buffer, the acid production rate may fail, allowing the sediment pH to rise and level off at a higher pH. The effect of 0.1 per cent sodium fluoride is shown in Fig. 3 with a diagram of the accessory equipment used for measuring pH. Next the test solution is removed and a control solution is placed in contact with RECORDER p H AUTOMATIC SWITCH ........... .... ß ,,,:•, i• _.•• .... - :-.q.• .... • T• 4 o•. 4 S 6 ? 8 pH Figure 3.