EVALUATING IRRITATION POTENTIAL 295 1 ß 0 1 r=O.$1 p=O.07 0 1 2 3 4 5 6 7 In Vitro Ranking Figure 3. Correlation between the ranking of antibacterial liquid soaps using the pH rise test and the ranking of antibacterial liquid soaps using the modified soap chamber test. Zein test. Zein is a protein that is insoluble in aqueous solution unless denatured by irritating detergent products. In this test (3), fifty milligrams of zein from Sigma (St. Louis, MO) are incubated with a solution of the finished product at 1% of the dry extract. The solutions are mixed at room temperature at pH 7.0 for one hour. After incubation, the solubilized zein is isolated and assayed for protein using the BCA protein assay kit (Pierce Chemical). The more zein that is solubilized, the more irritating the product. The results are expressed as follows: Zein = milligrams of solubilized zein per milliliter of solution IN VIVO METHODS A modified soap chamber test was performed according to the method of Simion et al. (11) in early April by an independent testing laboratory. In this test, 0.3 ml of the products was applied at 5% dry extract to the panelist's forearms using occlusive Hill Top Chambers. Twenty-four hours later, the patches were removed, and the sites were

296 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS 6• 2• r = -0.26 p = 0.57 0 1 2 3 4 5 6 7 In Vitro Ranking Figure 4. Correlation between the ranking of antibacterial liquid soaps using the zein test and the ranking of antibacterial liquid soaps using the modified soap chamber test. rinsed with running tap water and patted dry. The panelists waited three hours before skin irritation assessments of redness (clinical and instrumental, using the Minolta Chroma Meter) and barrier damage (Servo Med Evaporimeter). The products were reapplied for a further 21 hours and assessed as above, as well as 24 hours after patch removal. The products were ranked on their overall irritation score based on their propensity to produce erythema and skin barrier damage. RESULTS IN VITRO RESULTS Table II summarizes the results of the three in vitro studies. In order to compare the irritation prediction scores for the three methods, the results have been expressed as a percentage of the score obtained with a solution of SLS at 1% in each of the tests. There is some variation as to how the three tests rank the products. The collagen swelling test ranks Product B as the mildest and Product F as the most irritating. The pH rise test