358 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS with the densitometer. If growth occurred it was manifested by an increase in turbidity in the broth. The results are tabulated in Table II. DISCUSSION AND RESULTS To prepare the salicylanilides, the standard procedure of reacting the substituted salicylic acid with a substituted aniline in the presence of phosphorus trichloride was employed. According to the method of LeMaire (1) the preparation of the 4'-bromothiosalicylanilide gave TABLE II Antibacterial Activity of Salicylanilides and Cyclic Salicylanilides Minimum Inhibitory Concentration Range X 10 2 Compound Against S. aureus Against A. fecalis Salicylanilide 4'-Bromosalicylanilide 3,5- Dibromo-3'-trifluoromethyl- salicylanilide 4'-Bromothiosalicylanilide 4',5-Dibromosalicyl anilide 3 '-Trifluoromethylsalicylanilide 2'-Chloro-5'-trifiuoromethyl- salicylanilide 3-Phenyl-l,3-benzoxazine-2,4-dionc 6,8- Dibromo-3- (3-trifluoromethyl- phenyl)-l,3-benzoxazinc-2,4-dione 3-(4-Bromopbenyl)-l,3- bcnzothiazinc-2,4-dione 6- Bromo-3- (4-bromophenyl)- 1,3- benzoxazine-2,4-dione 3- (3-Trifiuoromethylphenyl)-l,3- benzoxazine-2,4-dione 3-(3-Trifiuoromethyl-2-chlorophenyl)- 1,3-benzoxazine-2,4-dione 1:10-1:100 1:10-1:100 1:100-1:1000 1:1-1:10 1:1000-1:10,000 1:10-1:100 1:100-1:1000 1:1-1:10 1:1000-1:10,000 1:1-1:10 1:1000-1:10,000 1:1-1:10 1:1000-1:10,000 1:1-1:10 1:1-1:10 1:1-1:10 1:1000-1:10,000 1:1-1:10 1:10-1:100 1:1-1:10 1:1000-1:10,000 1:10--1:100 1:100-1:1000 1:1-1:10 1:1000-1:10,000 1:1-1:10 the disulfide. It was found that when the order of mixing and reflux time were altered the mercapto derivative could be obtained instead of the disulfide. Thus, when a slurry of o-mercaptobenzoic acid, phospho- rus trichloride, and p-bromo-aniline in chlorobenzene was refluxed two hours, the mercapto derivative was obtained. To prepare the benzoxazinediones (I), the procedure of Stenseth (2) employing the reaction of a salicylanilide in a mixture of pyridine and acetonitrile with ethyl chloroformate was used. All of the products were characterized by infrared spectra and carbon-hydrogen analyses. It

CYCLIC SALICYLANILIDES 359 was found that the benzoxazinediones could be prepared by allowing an acetone solution of the salicylanilide saturated with phosgene to stand for three days. In general, the yield by the phosgene method was not particularly good. o o R I II It was thought that if an atom of sulfur were introduced into the benzoxazine molecule a more active antimicrobial agent might be formed. Thus, a benzothiazine-2,4-dione (II) was prepared employing the ethyl chloroformate procedure. The microbiological screening did not in- dicate increased activity of the cyclic derivative. Attempts were made to prepare compounds containing a sulfur atom in the 2-position. It was thought that this might be done by replacing the ethyl chloroformate with thionyl chloride. Crystalline solids were obtained which were not starting material. They also contained nitrogen but no sulfur. No positive identification has been made of these products. A comparison of the antibacterial activities of the various halogen and trifluoromethyl substituted salicylanilides with the mercapto- benzanilide showed that sulfur-containing compounds had no greater activity than compounds without sulfur. It was also determined that the cyclization of the various salicyl- anilides did not produce a change in antibacterial activity. A decrease in activity was shown by the cyclization of the thiobenzanilide. It is interesting to note that cyclization, which changed a secondary amide to a tertiary amide, did not result in a change in antibacterial activity. This would suggest that the mode of action of these agents is more likely physical blockage of the pores of the cell membrane of the micro- organisms rather than an actual binding or complexation within the microorganism. SUMMARY A number of salicylanilides, benzoxazinediones, and a benzoxathia- zinedione were prepared for evaluation as antibacterial agents. Anti- bacterial screening by the minimum inhibitory concentration method showed that the trifluoromethylsalicylanilides and their cyclic deriva- tives are potent antibacterial agents.