J. Soc. Cosmetic Chemists, 16, 747-761 (1965) An the Improved Method for Testing Safety of Hair Dye Preparations STEVEN CARSON, Ph.D., MYRON S. WEINBERG, Ph.D., and RICHARD GOLDHAMER, B.S.* Presented November •, 196•, New York City Synopsis--The Draize procedure for subacute dennal studies in rabbits was compared with a modified procedure in which certain of the critical parameters were modified for the evalua- tion of oxidation hair dyes. These included limitation of the duration of contact to one hour, clipping the hair to •/• to •/• in. instead of complete depilation, and thorough washing of the application sites after the one-hour contact. The results revealed that the modified method was equally sensitive with respect to the toxicological parameters examined. A further modi- fication was employed in which the sites of application were excised and analyzed for hemo- globin content as an index of erythema. Addition of this procedure made it possible to detect effects within one hour of application. Significant differences were demonstrated between the irritation potential of 3 and 12% hydrogen peroxide. INTRODUCTION Irritation is manifested by a tissue system in response to stimuli of either exogenous or endogenous origins. The characteristic reactions include rubescence (erythema), edema, inflammation, and possible im- pairment of the integrity of the associated vasculature, with necrosis and tissue degradation if the stimulus is sufficiently intense or prolonged. In this report, two dermal test procedures employing rabbits have been compared for their utility in the evaluation of systemic safety and irritation potential of oxidation hair dyes. One of these methods, origi- nally described by Draize (1), has been widely applied to the testing of cosmetics, topical pharmaceuticals, and industrial or agricultural chemi- * Food and Drug Research Laboratories, Inc., Maurice Avenue at 58th Street, Maspeth, New York. 747

748 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS cals. A modified test procedure was used which more closely approxi- mates use conditions. Oxidation hair dye formulations characteris- tically contain ammoniated bases which for use are mixed with an ap- propriate quantity and concentration of hydrogen peroxide. These dyes are used under specified conditions which include limited contact with tlm hair, followed by thorough sk_ampooing and immediate rinsing. These conditions are employed both in the home and in the beauty salon. In the Draize test prolonged (six-hour) contact is maintained with the test material by means of either a rubber dam. or plastic sleeve wrapped around the trunk of the rabbits. The condition of the skin is TABLE I Evaluation of Skin Reactions Erythema and Eschar Formation No erythema 0 Very slight erythema (barely perceptible) 1 Well defined erythema 2 Moderate to severe erythema 3 Severe erythema (beet redness) to slight eschar formation (inJuries in depth) 4 _ Total possible erythema score 4 Edema Formation No edema 0 Very slight edema (barely perceptible) 1 Slight edema (edges of area well defined by definite raising) 2 Moderate edema (raised approximately 1 mm.) 3 Severe edema (raised more than 1 min. and extending beyond area of exposure) 4 -- Total possible edema score 4 scored daily in accordance with Draize's grading system. (Table I) for evaluating skin reaction, whereby emphasis is placed on the degree of edema, erythema, and esehar formation. Under these test conditions, in which daily (5 days per week) applica- tions to the abraded skin are made for 6 hours for 21 days, or to the in- tact skin for 90 days, even the most innocuous materials dieit adverse changes in the skin of rabbits. As a result of this treatment regimen the skin becomes dry and sealy, cracks, and thickens, with subsequent sloughing. Relatively large body areas become denuded, but regrowth of hair is generally observed in these areas. Since the test procedure precludes washing of the application area, it is intrinsically responsible for many of the physical changes observed. The effects due to any active components in the formulation are superimposed on these back- ground reactions.