378 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS safety of matehals following their application to the skin of laboratory animals. These are methods which research laboratories have employed for a long time, and which in the hands of experienced pharmacologists, have a high degree of reliability. They, therefore, can be used effectively to screen new chemicals and chemical mixtures. Our aim, in the laboratory, is to eliminate those preparations which may be irritating to the skin or, through absorption, show systemic toxicity. Using the animal as a substitute for the human, we use the same signposts as the dermatologist to judge effects. We look for reactions which may take the form, in order of severity, of primary irritation, sensitization or systemic toxicity. We have found that the response to a substance applied to the skin of the rabbit is reproducible, and often similar, to the response in the human. Potential hazards are evaluated by the use of laboratory animals which are raised under controlled conditions, and which react to the same irritants in a similar manner. Derreal changes in these animals, following application of an irritant, vary from slight erythema and edema to overt necrosis. These changes may be seen by gross observation or they may be magnified by histological section. Observations are made following single and repeated contact. A single application may cause no reaction while repeated applica- tions could result in skin responses which may or may not return to normal before each succeeding application. Some products are found which show reactions foilowing several applications. This response may disappear or become very mild after continuous treatment. As a result of many years of experience we have found that the rabbit, by virtue of its large skin area, ease of handling, maintenance and avail- ability, is the animal of choice for the screening of compounds. Although histologically the skin of the rabbit differs markedly from that of man in the relative thickness of the epidermal layer, and has a vigorous hair growth, we will attempt to show that results obtained from its use can be correlated with the responses which would occur in the human. The pig has been suggested as a better experimental animal, because of its lack of hair and thick epidermal skin layer, but its size makes it a difficult experimental subject. Sensitivity and general toxicity do not go hand in hand and are not interchangeable. Sensitivity is a random reaction and does not respond to a concentration gradient. An animal or human responds to a sensitizing agent if its threshold, which may be of a very low concentration, is reached. Sensitization in animals depends upon as many variables as sensitization in man. Factors of age, diet, previous or present illness, environmental conditions, sex and method of application all play a part. Animal experience indicates that strong sensitizers may be picked up in the guinea pig and

RABBIT SKIN STUDIES IN EVALUATING COSMETIC SAFETY 379 kept from man, but the weak reactors may not be eliminated but must' be studied employing the human patch test techniques. The laboratory techniques for studying sensitization in the guinea pig follow the exact timing procedure which the dermatologist employs in the human. Instead of a patch technique, the material is injected intradermally: (as an 0.1% solution or suspension) every other day for ten applications. Following a two weeks' rest a challenging dose is injected, and the resulting response compared to the preceding reactions. Sensitization is present if the challenging dose causes a greater skin response than that evoked by any earlier injection. Primary irritation, which denotes a definite skin response to a single application of a cosmetic, can be readily studied in the rabbit. The patch test technique is employed, and a known amount of material is applied to the backs of animals the fur of which has been previously removed with the aid of an electric clipper. Several abrasions are made on one side of each animal. These abrasions penetrate to the dermis but do not go through it, eliminating bleeding. The patch used is a 2"x 2" gauze and a standard dose of 0.5 g or 0.5 ml of the same is applied under each patch. If a powder is being studied, a thick paste is made with water, applied to the skin and, then covered with the patch. Each animal receives one patch on its intact side and one patch over the abraded area. After the patches are applied they are kept in place by means of a rubber dam. This also reduces evaporation. The animals are immobilized in a holder for 24 hours, after' which time the patches are removed and the degree of skin reaction noted. Readings are also made 72 hours after the patches are removed. Scoring is based upon two responses--erythema and edema. Each response is further divided according to degree, and a numerical value given for each stage. Erythema may manifest itself by slight pink to beet red, while edema may be barely perceptible or may produce a raised area more than 1 mm in height at the site of treatment. In order to determine the extent of correlation when this procedure is followed in the rabbit, with the response which the human would show to the same preparations the following experiment was performed. Four prepara- tions were screened for their activity on both the rabbit and the human.. The first was a depilatory in a cream form, the second a revised lornrelation of a nail polish remover, the third a shampoo containing a new anti-dandruff ingredient, and the fourth a cuticle softener. When these four materials were each patch tested on the backs of groups of rabbits the animals res- ponded by showing a marked edema and erythema. After these results were obtained 24-hour patch tests were made with each of these preparations under the direction of a dermatologist. 25 humans received each of these materials as a single patch, and the area was examined 24 and 72 hours after exposure.