374 JOURNAL OF COSMETIC SCIENCE Table III Comparison of Water Permeation Parameters for Pairs of Thumbnails, Index Fingernails, and Long Fingernails From the Same Donor Nail Thickness (mm) J (mg cm-2h -•) J* (mg cm-•h -1) Total water uptake (mean + SEM) (mean _+ SEM) (mean _+ SEM) (mg cm-3) a Thumbnails (n = 2) 0.66 + 0.02 6.72 + 1.947 0.44 + 0.1277 242.01 + 8.90 Index fingernails (n = 2) 0.56 _+ 0.10 14.52 + 1.08 0.80 + 0.08 283.31 + 32.52 Long fingernails (n = 2) 0.52 _+ 0.06 15.48 * 2.35 0.79 -+ 0.03 361.88 _+ 71.06 a Calculated by dividing total water present in nails (from extractions) by the product of cell area and nail thickness (assuming lateral diffusion is negligible). t Indicates statistically significantly different fromJ of index and long fingernails (ANOVA, tx = 0.05). '•t Indicates statistically significantly different from J* of index and long fingernails (ANOVA, tx = 0.1). level, a difference at the 90% confidence level between the J* values of the thumbnails and other fingernails, using such a small sample size (n = 2), suggests that structural differences exist between the thumbnails and other fingernails. However, the total water uptake values were not statistically significantly different from each other for the three sets of fingernails by ANOVA at the 0.05 level of significance. Thus, our results are suggestive, but not conclusive, that thumbnails have a unique structure and possibly a tighter cross-linked keratin network, as compared to other fingernails, which results in their lowJ and J* values. Moreover, the structure of the nail keratin network appears to be similar for the index and long fingernails from the same donor, and nail thickness appears to be a primary cause for variability in their permeation parameters. Our results indicate that there is a large interindividual variability in nail permeation. Normalizing the parameters for nail thickness helps to remove one source of variability between nails. Further, nail location is important, and thumbnails appear to possess a tighter cross-linked keratin structure than do toenails and possibly other fingernails. EFFECT OF pH ON IN VITRO NAIL PERMEATION OF WATER The normalized water flux values obtained under conditions of acidic, neutral, and alkaline pH are shown in Table IV. A statistically significant increase in the permeation of water under alkaline conditions was observed in experiments using the standard receptor (relative to the neutral gel) at the 0.05 level of significance. However, the flux increase was less than twofold. For the experiments wherein the donor and receptor contained the same buffer, the J* values at acidic and alkaline pH were not statistically Table IV Effect of pH on the Permeation of Water Through Toenails j* (mg cm-•h -•) (mean + SEM, n = 3-6) pH Standard receptor Receptor = Donor 1.96 1.52 + 0.08 1.95 + 0.17 7.01 1.48 _+ 0.10 1.61 + 0.16 11.27 2.09 _+ 0.13• -- 11.69 -- 1.39 _+ 0.10 Indicates statistically significantly different from neutral conditions (t-test, tx = 0.05).

NAIL PERMEATION 375 significantly different from neutral conditions (t-test, ot -- 0.05). Hence, wide variations in pH did not appear to affect the overall intrinsic permeability properties of the human nail. High concentrations of alkalis are known to catalyze [3-elimination of disulfide linkages in hair keratin, thereby producing reactive intermediates, which can then cross-link with other residues in the polypeptide chain. The reaction is reversible in the presence of excess water. The alkaline degradation of cystine linkages in hair has been shown to occur with alkali treatment at pH values • i 1.0 and at elevated temperature (40øC) (9). Thus, the small effect observed in our experiments at alkaline pH using the standard receptor may be attributed to some degradation of the cystine linkages in nail keratin. The effect of pH on nail permeability had been investigated earlier by other researchers using ionizable drugs, and conflicting results had been reported. It was found that pH had little effect on the permeation of the antifungal drug miconazole (6). However, in the case of model compounds like benzoic acid and pyridine, it was found that pH indeed affected permeation and that the undissociated drug preferentially permeated the nail plate (5,7). Thus, it was unclear whether pH had an effect on the nature of the nail plate itself or on the ionization of the drugs used in these studies. In our experiments using water as a probe, we have demonstrated that pH does not affect intrinsic nail permeability properties. EFFECT OF TEMPERATURE ON IN VITRO NAIL PERMEATION OF WATER Water permeation through human nails was significantly enhanced with increasing temperature. The relationship between normalized permeability coefficient for water and temperature is defined by the Arrhenius equation as shown below: Ea log P* = log Po* 2.303RT (3) where Po* is a factor independent of temperature and is related to the number of molecules entering the diffusion process and the probability that these molecules have sufficient energy to engage in diffusion, E a = energy of activation in calories/mole or kJ/mole, R = gas constant, and T = absolute temperature (10). Figure 3 shows the Arrhenius plot of normalized permeability coefficient for water. As predicted by equation 3, a linear relationship between log P* and reciprocal temperature was observed. The energy of activation (Ea), computed from the slope of the plot, was found to be 28.4 kJ/mole (approximately 7.2 kcal/mole). Skin permeability has been documented to be temperature-sensitive, with higher temperatures resulting in in- creased permeation (11). This increased permeation is thought to result from a loss of crystallinity on heating. The E• value for the permeation of water through the nail is closer to the value for delipidized human stratum corneum (E• = 6.3 kcal/mole) than for intact stratum corneum (E• = 14.3 kcal/mole) (12). TEST FOR BARRIER INTEGRITY OF NAILS AFTER pH AND TEMPERATURE STUDIES The normalized flux values before and after pH and temperature studies, shown in Table V, are essentially the same. These results suggest that pH/temperature treatments do not