370 JOURNAL OF COSMETIC SCIENCE Various clinical methods for evaluating the efficacy of antibacterial soap products have been reported in the literature (1,5-7). These methods typically measure the ability of a product to reduce the resident flora through a continuous action or evaluate the ability of a product to reduce the level of marker organisms on artificially contaminated hands. These methods, however, do not measure the ability of an antibacterial agent to remain on the skin for a prolonged period and continue suppressing growth of acquired skin pathogens in situ. This antibacterial substantivity or residual effectiveness is an impor- tant characteristic to measure in evaluating the overall performance of an antibacterial product. Aly and Maibach (8) reported the development of a method to evaluate the residual effectiveness of a single application of topical antibacterial products against various skin pathogens. This method incorporated the occlusion of treated skin sites inoculated with bacteria and the enumeration of the bacteria remaining after five hours of occlusion via the cup scrub technique. Residual antibacterial effectiveness was determined by com- paring the differences in growth and survival of the test organism on sites treated with either the antimicrobial agent or a vehicle. Finkey et al. (9) and Scala et al. (10) used this method to evaluate antimicrobial soaps following repeated application. These studies did not, however, establish how quickly the antibacterial effect occurs or how long it lasts. Our objective was to determine the rate and duration of the antibacterial action of antibacterial soaps under simulated use conditions. We modified the previously reported residual effectiveness test procedure (9,10) by inoculating the sites either immediately or 24 hours after the final wash and harvesting the surviving organisms after various intervals of occlusion. MATERIALS AND METHODS Two studies illustrate the method, one study examining the immediate antibacterial efficacy of the bar soap containing 1.5% TCC relative to the vehicle bar, while the second study examines the prolonged antibacterial efficacy of the same two bars. STUDY POPULATION Twenty (20) subjects, male and female, ages 18 to 65 years, were enrolled in each study. These subjects were screened for normal skin on the forearms. In order to clear the skin of any residual antimicrobials previously used, subjects were instructed to use only the non-medicated personal cleansing bar soap provided in place of their regular personal cleansing product during a 7-14-day washout period. They were also instructed to refrain from using any antibacterial soaps, medicated lotions and creams, and dandruff shampoos until the study was completed. All subjects provided written, informed consent. TEST MATERIALS Two bar soaps were examined in each study. The antibacterial soap contained 1.5% 3, 4, 4'-trichlorocarbanilide (TCC). The other soap was the vehicle soap bar containing no antimicrobial agents.

ANTIBACTERIAL SUBSTANTIVITY 371 APPLICATION OF TEST PRODUCTS In each study, assignment of the test and placebo soaps to the subjects was made according to a computer-generated randomization. Following the 7-14-day washout period, subjects performed a supervised wash of their forearms three (3) times per day at the laboratory with their assigned bar of soap. Washes were at least one hour apart. To assess immediate residual effectiveness after washing, subjects performed seven washes: three washes on days 1 and 2 and one wash on day 3 of the test period. For evaluation of prolonged residual effectiveness, subjects performed nine washes: three washes on days 1, 2, and 3. Subjects were permitted to shower and bathe during the test period, using a non-antibacterial bar soap. However, they were instructed not to wash their arms. Showering and bathing was restricted after the last wash. Subjects were permitted to use a non-medicated lotion on their forearms whenever the forearms became dry, except after the final wash. At each wash visit, the subjects were closely supervised as they washed their own arms. Each forearm was washed separately with the appropriate soap. The volar surface of the forearm and the bar soap were wetted under running tap water maintained at 95 ø- 100øF. The bar was then rubbed on the forearm using an up-and-down motion from the wrist to the elbow for 15 seconds. The lather was then rubbed on the forearm using the hands in the same motion for another 45 seconds, followed by a 15-second rinse. Each forearm was patted dry with a paper towel. INOCULATION OF TEST SITES On the last day of the test period (immediately after the seventh wash or 24 hours after the ninth wash), three (3) circular test sites, 3.0 cm in diameter, were marked on the volar surface of each forearm. These circular sites were evenly spaced along the midsec- tion of each forearm, avoiding the area on the wrist and the elbow crease. Each test site was inoculated with S. a•reus, strain 502A (ATCC # 27217) that had been grown at 35 ø + 2øC for 20 5 2 hours in trypticase soy broth (TSB). For inoculation, a ten-fold dilution of the S. aureus broth culture was made, and 10 lal of the diluted bacterial culture was applied to each test site. The inoculum concentration ranged between 3.4 x 105 and 7.8 x 105 colony-forming units (CFUs) for the inoculation immediately fol- lowing the final wash and between 1.2 x 10 6 and 1.5 x 10 6 CFUs for the inoculation 24 hours following the final wash. Using a sterile inoculating loop, the inoculum was spread within the center of the test site while remaining 4 to 5 mm from the marked edge. Each inoculated test site was immediately occluded with a 25-mm Hill Top Chamber © without the non-woven absorbent pad. The chambers were secured to the skin with an adhesive dressing (Durapore ©, 3M). Sites remained occluded for intervals of 30 minutes, two hours, or five hours. HARVESTING OF SURVIVING ORGANISMS At the completion of each occlusion interval, the chambers were removed and the sites were sampled for surviving organisms using the cup scrub technique (1,2). The sampling