544 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS Table X Antimicrobial effect of p-hydroxybenzoates mixed with formaldehyde (18) No. of tubes with growth Dilution of No. of tubes challenged % p-hydroxy- Formaldehyde challenge benzoates [ppm culture Yeasts and Bacteria moulds _ 0 8 None 13/13 7/7 10-3 13/13 5/7 lO-6 13/13 s/7 0 53 None 10/13 7/7 10-3 4/13 4/7 10-6 0/13 3/7 0.165 8 None 7/13 3/7 10-3 5/13a 0/7 10-6 3/lab 0/7 0.165 54 None 0/13 1/7 10-3 0/13 0/7 10-6 0/13 0/7 to both bacteria and fungi. They also showed that p-chlorophenoxetol at 0.2% provided very favourable results. The authors chose Phenoxetol for their investigations because of its known activity against Pseudomonas pyocyanea-- a serious potential pathogen which may grow in vaccines and other medicinals. In their conclusions, these authors stated that Phenoxetol supplements the antibacterial activity of antibiotics, (a point already referred to above in considering penicillin-Phenoxetol systems) provides adequate antifungal activity and has excellent stability. Furthermore, this product is not inactivated by materials rich in protein, in contrast to bithio- nol and quaternary antibacterial agents, e.g. cetyl pyridinium chloride, which are inactivated by fatty acids present in protein (19). No benefit is obtained when using a combination of a material such as benzalkonium chloride with hexachlorophane, where a loss of antibacterial activity is observed (20). When considering a particular component for a potentially synergistic combination for use in preservation, the components must be compatible, firsfly with each other and secondly, with the material into which they are incorporated and which they are intended to preserve. Another recent publication describing the synergism of combinations of antibiotics and other well-known antimicrobial agents (21) describes the combined use of an antibiotic, e.g. neomycin and a product of the Phen- oxetol type, in providing an effective system for the treatment of ear

SYNERGISM IN VITRO OF CERTAIN ANTIMICROBIAL AGENTS 545 infections. This preparation was both bactericidal and bacteriostatic against gram-negative and gram-positive organisms. In the cosmetic and semi-cosmetic/pharmaceutical fields much has to be recommended on the basis of the bacteriological results given above for using such effective systems in products such as body deodorants, anti- dandruff preparations, shampoos, antimycotic foot preparations, after- shave preparations and cosmetic-serum products. Janistyn (22), for ex- ample, has especially recommended the use of a combination of Phenoxetol with other antiseptic agents in shaving lotions and other preparations required for use as effective antiseptic agents in the treatment of infectious skin diseases, e.g. dermatitis. Considerable work by Clausen and Raugstad (23) has included com- binations of many different antimicrobial agents including benzalkonium chloride, aminacrine hydrochloride, esters of p-hydroxybenzoic acid, benzyl alcohol, Phenoxetol and its derivatives, dibromo-propamidine isethionate, etc. Some of the results obtained by Clausen et al (23) are shown in Tables XI-XIII. The synergistic combination of propylene Phenoxetol at 1.0% and aminacrine hydrochloride at 0.05% in aqueous solution in the absence, or presence, of normal serum has an excellent bactericidal effect. The compatibility of this system with serum (i.e. a protein rich medium) is another feature worth mentioning, as this is in contrast to products such as quaternary antimicrobial agents which are inactivated by such media. This combination has an excellent effect on Pseudomonas pyocyanea which, as we have seen, appears to be highly resistant especially in the presence of organic matter. This synergistic combination was also effective against E. coli, although a weaker effect was displayed against gram-positive micro- organisms. The results in Tables XI-XIII were obtained from tests conducted in 20% horse serum. Since the antimicrobial effect of many substances is considerably reduced by blood, the results obtained give an idea of the antiseptic effect of the products involved, i.e. a determination of their usefulness in preventing infection in topical therapy. We have already seen that the glycol ethers, e.g. Phenoxetol and its derivatives, are readily compatible with organic material of this type and combined with their non- irritant effect on the skin they may be considered well suited to external therapy. Quaternary compounds on the other hand, e.g. benzalkonium chloride or cetyl pyridinium chloride, as well as organic mercury corn-