PADIMATE-O 121 Table I Effects of Application Vehicle and Skin Viability on Absorption of Padimate-O Through Hairless Guinea Pig Skin Radiolabel recovered, percent of applied dose Ethanol Lotion Viable Viable Nonviable Receptor fluid 18.2 --- 1.7 4.3 --- 0.4 6.8 --- 1.5 Stratum corneum• 7.4 +-- 1.3 1.3 --- 0.2 1.7 --- 0.3 Viable skin layer 2 16.9 --- 1.9 6.0 --- 1.0 4.3 --- 1.2 Total absorbed 42.5 --- 2.1 11.6 --- 1.1 12.7 +-- 2.5 24-h wash 49.1 +-- 1.8 73.8 --- 3.9 74.1 +-- 3.9 Total recovered 91.6 --- 1.5 85.4 --- 3.9 86.8 --- 5.0 Each value is the mean + S.E. of four or five determinations in each of three animals (ethanol-viable, 1otion-nonviable) or six animals (lotion-viable). • Surface layer of skin removed by cellophane-tape stripping. 2 Skin remaining after tape stripping. tion of Padimate-O does not appear to depend on maintaining skin viability in the diffusion cells. More than half of the absorbed compound remained in the skin at the end of the 24-h experiment (Table I). The percentage of the applied dose absorbed was significantly higher in the receptor fluid and skin fractions when the ethanol vehicle was used. Presumably this was due, at least in part, to the increased levels of Padimate-O that partitioned into the skin as the ethanol evaporated. Substantial amounts (between 10 and 20%) of the absorbed Padimate-O were found in the stratum corneum. This is expected for a very lipophilic compound and helps to account for the effectiveness of this sunscreen agent. Padimate-O was also found in the epidermal and papillary dermal tissue that remained after removal of the stratum cor- neum by tape stripping. This viable tissue is 15 to 20 times thicker than the stratum corneum, which accounts for the larger amounts of Padimate-O found in this layer. Overall absorption of Padimate-O in the ethanol vehicle was four times greater than that in the lotion vehicle. Recoveries of approximately 80-93% of the applied dose were typically found. Padimate-O and NMPABAO are lipophilic chemicals, as evidenced by a log octanol/ water partition coefficient value of 3.86 for Padimate-O (data not shown). Bovine serum albumin was added to the receptor fluid to increase the solubility of these compounds and thereby facilitate partitioning from skin into the receptor fluid. However, total percutaneous absorption must include not only the compound absorbed into the receptor fluid but also that material absorbed and found remaining in the skin at the end of the experiment. Limited solubility of test compounds in the receptor fluid can result in an underestimation of skin absorption unless skin levels of the test compound and metab- olites are included as being percutaneously absorbed. A portion of the absorbed Padimate-O was hydrolyzed to DMABA by esterase in skin during percutaneous absorption (Table II). However, no difference was observed for metabolism in viable and nonviable skin. The viability of skin was maintained to

122 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS Table II Effects of Application Vehicle and Skin Viability on Esterase Hydrolysis of Padimate-O in Hairless Guinea Pig Skin Percent of absorbed dose hydrolyzed to DMABA Ethanol Lotion Viable Viable Nonviable Receptor fluid fraction 0-6 h 29.9 + 7.1 8.3 + 1.3 10.9 + 1.6 6-12 h 29.8 + 8.3 10.4 + 2.2 9.9 + 1.1 12-18 h 27.6 + 7.4 9.7 --- 2.1 8.9 + 2.9 18-24 h 26.0 --- 7.8 9.6 + 2.6 10.6 + 4.3 Skin 5.4 -+ 2.3 2.3 -+ 0.7 2.7 -+ 0.9 Each value is the mean + S.E. of four or five determinations in each of three animals (ethanol-viable, lotion-nonviable) or six animals (lotion-viable). observe the acetylation of PABA and benzocaine in vitro in diffusion cells (13). However, esterase activity in skin is stable and does not require cofactors. Esterase activity has previously been observed histochemically in nonliving stratum corneum (14). No sig- nificant differences in the percentages metabolized were found for the receptor fluid fractions within each individual group. Therefore, just as the rate of absorption of Padimate-O remained relatively constant throughout the 24-h experiment, so did the percentage metabolized. Only a small amount of DMABA was found in the skin of each group. The metabolism of Padimate-O was 2-3 times greater with the ethanol vehicle, which was previously shown to markedly enhance percutaneous absorption. Whether increased absorption of Padimate-O results directly in increased metabolism of the compound is not clear. Possibly, pathways of absorption (and exposure to metabolic enzymes) differ during metabolism when a compound is applied in different vehicles. The percutaneous absorption of NMPABAO from the lotion vehicle was less than half of the absorption of Padimate-O (Table III). As with Padimate-O, absorption of NM- PABAO was similar through viable and nonviable skin. Approximately 1% of NMPA- BAO penetrated into each receptor fluid fraction of both groups throughout the 24-h experiments. The levels of the nitrosamine in skin at 24 h were substantially lower than the levels of Padimate-O. NMPABAO was metabolized to NMPABA by esterase activity in skin during percu- taneous absorption (Table IV). The differences in metabolism of NMPABAO within the receptor fluid fractions of either viable or nonviable skin were not significant. However, significant differences were found for metabolism in viable and nonviable skin sections approximately four times as much NMPABA was recovered in the viable receptor fluid fractions. The reason for the effect of skin viability on the metabolism of NMPABAO and the absence of an effect on the metabolism of Padimate-O is not clear. Possibly, different isozymes of esterase are involved in the hydrolysis of the two compounds. Also, a reduction in the pH of nonviable skin may have an effect on enzyme activity. Little or no NMPABA remained in the skin of each group at the end of the experiments. These