64 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS ing on the backs of human subjects. After 6 hours of occlusion, treated sites were covered with a sterile bandage and the blister tops were allowed to fill overnight. Blister tops were cut off with scissors, remaining epidermal tissue was removed with a wool-tipped stick, and samples were stored in a desic- cator. Trypsinized SC was obtained from the abdominal region of cadavers and separated by the method of Kligman and Christophers (4). Epidermal tissue was gently removed with a wool-tipped stick and samples were stored in a desiccator until they were used. Sunburn exfoliated tissue was obtained in a single sheet from the back of an individual 10 days after 1.5 hours of ex- posure to summer sun and was stored in a desiccator. Hydration measurements were carried out .on 6 or 8 mm punches of dry SC mounted on nichrome wire hooks. Samples were weighed in one of two 15 x 15 x 20 cm chambers which had controlled environments. Within the chambers, up to 6 SC samples could be manipulated from storage hooks to a balance wire extending down from a Cahn RG Electrobalance • mounted on top of the chambers. In addition, samples could be transferred from one cham- ber to the other with a slidewire via a sliding door so that all measurements could be made without disturbing the environment in the chamber. Humidity was maintained by a salt solution within each chamber, which generated a constant RH (5). The following saturated solutions were used: magnesium nitrite (RH equals 30 per cent), potassium carbonate (RH equals 40 per cent), sodium nitrite (60 per cent), sodium chloride (75 per cent), potassium chloride (83 per cent), potassium nitrite (93 per cent), and potas- sium sulphate (97 per cent). The hydration chamber was mounted within a modified Hydro-Jac* incubator, which was coupled with a water bath to regu- late the temperature of the incubator (-0.2øC). The temperature was moni- tored with a 46-TU Telethermometer Thermistor and the humidity was mon- itored by means of Hygrodynamics • narrow-range sensors located within the chamber. Changes in temperature, humidity, and sample weight were re- corded simultaneously throughout the experiments on strip-chart recorders. Water content was measured as milligram H20 per milligram dry SC (mgl H20/mg SC). To study the effects of heat exposure, two 8 mm punches were cut from one sample harvested by each of the 3 methods. A dry weight was taken for each sample. The samples were transferred to the second chamber at 97 per cent RH for 48 hours, and hydrated weights were recorded. The samples were then wrapped in plastic and immersed in water at 60øC for 10 min. after which dry •Ventron Instruments Corp., Paramount, Calif. 90723. ]'Forma Scientific, Marietta, Ohio 45750. $Yellow Springs Instrument Co., Yellow Springs, Ohio 45387. •Hygrodynamics Inc., Silver Springs, Md. 20910.

WATER AND THE HORNY LAYER 65 and hydrated weights were again recorded. Effects of aqueous exposure were studied by immersing 8 mm punches of SC in distilled water for specified lengths of time. Each of 5 punches from a single piece of SC was immersed for 0, 1, 3, 10, or 24 hours. Following immer- sion, dry weights and water uptakes (97 per cent BH and 30øC for 48 hours) were recorded as described previously. Three sets of 5 were run, using the following 3 SC specimens: trypsin, cantharidin, and sunburn. Results The relationship between hydration of SC and BH or activity of water is shown in Fig. i for cantharidin blister and trypsin samples. Average dry weights and equilibrium hydrations for samples harvested by all three meth- ods are shown in Table I with statistical significance determined by analysis of variance and F-ratio test. Effects of exposure to water and to heat at 60øC are shown in Tables II and IH. The effect of temperature on the equilibrium water content at various re]a- five humidities has been discussed in detail by Spencer et al (3). The ability of the SC to retain water at equilibrium in vitro decreases with decreasing temperature at humidities be]ow 60 per cent. Above 60 per cent BH, this temperature dependence decreases with increasing BH until there is essen- tially no temperature effect on the equilibrium water content above 90 per cent. Kinetics of hydration of the SC from a dry weight to a steady state (SS) water content at 97 per cent BH were recorded for more than 45 samples. Analysis of hydration can be accomplished by treating the system as either diffusion of vapor into a membrane or pseudo-first order reaction of wate• with the membrane. Analysis of SC hydration as a diffusion problem can be defined as the sorp- tion of vapor by a plane sheet. Using an approximation for determining the average diffusion coefficient D from initial sorption data as described by described by Crank (6) and Scheuplein (7), one obtains the equation Wma. - 4 (*) where W equals mg H20/mg SC at time t after the dry sample has been placed in the hydration chamber Wmax equals the SS water weight (mg H20/mg SC) at 48 hours, • equals the average diffusion coefficient for the hydration t equals time (in minutes) and 1 is one-half the thickness of the SC. A plot of W/W,•ax versus t u will yield-• from the slope (m) squared as m 2 X -- 16.