MODEL SYSTEM FOR DANDRUFF INVESTIGATION 197 mutant resulted in an average increase of 88.1% in dandruff production, thus indicating the importance of this organism as an etiological agent. The appearance of appreciable levels of free fatty acids in the arti- ficial sebum was most probably the result of the hydrolytic activities of the microbial component of the system. Although fatty acids were pres- ent in the artificial sebum to which microorganisms were not added (Table IV), it appeared that an additional quantity of free fatty acids was required, particularly C•s monoenoic acid, to initiate the reaction. This would suggest that so•ne manner of fatty acid threshold must be exceeded to initiate sloughing. These data were further confirmed when cell-free fractions of extracellular medium constituents, containing demonstrable tributyrinase activity, produced an increase in per cent of C•s monoenoic acid and total free fatty acid content, and prolonged sloughing of at least moderate severity (Table V). It is thus apparent that extracellular lipases were capable of producing irritation in the absence of the organisms which initially synthesized them. The implications of these data for human dandruff are obvious. Hu- man sebum has been shown by Kellum (14) to be free of fatty acids as it is secreted by the sebaceous gland, yet appreciable levels are present in surface lipids. The probable source of these fatty acids is sebum tri- glycerides hydrolyzed by microbial enzymes since the three groups of organisms most commonly found on the scalp, P. ovale, diphtheroids, and staphylococci, are competent producers of lipase (15-17). Other data (18) have shown that fatty acids are irritants however, our attempts to apply specific concentrations of certain fatty acids to the human scalp have been unsuccessful due to the difficulty of maintaining these concen- trations on the scalp at all times. However, when attempts (Fig. 4) were made to produce guinea pig sloughing with the artificial sebum com- ponent which increases to the gxeatest degree on guinea pigs, oleic acid, a concentration-dependent sloughing reaction could readily be produced. In addition, dandruff is a disease appearing usually at puberty when sebaceous secretions on the scalp are increasing rapidly. It can be specu- lated that the concurrent onset of dandruff and extensive lipid secretion are more than coincidental. It should be remembered, however, that other physiological alterations occurring at this time could possibly in- duce dandruff-like conditions. (Received September 11, 1970)

198 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS REFERENCES (1) Emroohs, C. W., The isolation and pathogenicity of Pityrosporum ovale, Pub. Health Rep., 55, 1306 (1940). (2) Kile, R. L., and Engqnan, R. F., Further studies of the relation of Pityrosporon ovale to seborrheic eczema, Arch. Dermatol. Syphilol., 37, 289 (1938). (3) Reddish, G. F., The etiology of infectious dandruff, J. Soc. Cosmet. Chem., 3, 90 (1952). (4) MacLeod, J. M. H., and Dowling, G. B., An experimental study of the Pityrosporon of Malassez: Its morphology, cultivation, and pathogenicity, Brit. J. Dermatol., 40, 139 (1928). (5) Moore, M., and Kile, R. L., Pityrosporu•n ovalis as a causative agent of seborrheic der- matiris, Science, 81,277 (1935). (6) Rocha, G. L., Silva, C., Lima, A. O., and Goto, M., Experimental studies on Pityrosporum ovale: Its pathogenicity and antigenic capacity, J. Invest. Dermatol., 19, 289 (1952). (7) MartimScott, I., The Pityrosporum ovale, Brit. J. Dermatol., 64, 257 (1952). (8) Durfee, T., and Cousins, R. F., Practical tests of dandruff preparations, Stand. Rernedies, 22, 3 (1936). (9) Leone, R., Presence and significance of Pityrosporon ovalis in pityriases of the scalp, in figured seborrheic eczema and in various squamous dermatoses, Minerva Dermatol., 27, 93 (1952). (10) Spoor, H. J., Dandruff updated, Amer. Perfurn. Cosmet., 81, 81 (1966). (11) Baird-Parker, A. C., A classification of micrococci and staphylococci based on physiologi- cal and biochemical tests, J. Gen. Microbiol., 30, 409 (1963). (12) Metcalf, L. D., Schmitz, A. A., and Pelka, V. R., Rapid preparation of fatty acid esters from lipids for gas chromatographic analysis, Anal. Chem., 38, 514 (1966). (13) Gosse, R. M., and Vanderwyk, R. W., The relationship of nystatin-resistant strain of Pityrosporum ovale to dandruff, J. Soc. Cosmet. Chem., 20, 60• (1969). (14) Kellum, R. E., Human sebaceous gland 1ipids: Analysis by thin-layer chromatography, Arch. Dermatol., 95, 218 (1967). (15) Weary, P. E., Comedogenic potential of the lipid extract of Pityrosporum ovale, Ibid., 102, 84 (1970). (16) Kellum, R. E., Strangfeld, K., and Ray, L. F., Ache vulgaris. Studies in pathogenesis: Triglyceride hydrolysis by Corynebacterium acnes in vitro, Ibid., 101, 41 (1970). (17) Troller, J. A., and Bozeman, M. A., Isolati,on and characterization of a staphylococcal lipase, Appl. Microbiol., 20, 480 (1970). (18) Kellum, R. E., Ache vulgaris. Studies in pathogenesis: Relative irritancy of free fatty acids from C, to C•, Arch. Dermatol., 97, 722 (1968).