EFFECT OF AcHA ON STRATUM CORNEUM 175 was measured by Cutometer. The skin-softening effect was evaluated by the ratio of skin stretch length before and after the application. EVALUATION OF THE TREATMENT EFFECT OF A LOTION CONTAINING 0.2% AcHA FOR DRY SKIN CONDITION Four healthy male volunteers (26 - 46 yr) were selected for this study. All measurements were done in a room at constant temperature and constant humidity (25øC and 50% RH) Thirty minutes before the measurement, the skin surface of the ventral forearm was washed with soap and water to avoid the influence of residual oil and skin debris. The skin surface conductance and transepidermal water loss (TEWL) (baseline values) were measured by using a SKICON-200 hygrometer and Evaporimeter ER! (Servomeal, Stockholm, Sweden), respectively. Then, artificially dry skin was induced on the ventral forearm with 5 ml of 5 w/w% sodium dodecyl sulfate (SDS) by an Emanuel cup (inner diameter: 24 mm) for 30 min. After the treated region was washed with water and dried with a towel, photographs of silicon negative replica (adhesive silicon polymer) of the skin were taken. The photographs show the skin surface structure induced by SDS before the treatment. One ventral forearm was treated with a lotion containing 0.2 w/w% AcHA once a day and the other ventral forearm was treated with a lotion containing 0.2 w/w% HA (Table I). The treatment was continued for one week. The barrier function and the skin-moisturizing effect were evaluated by the ratios of values of TEWL and the skin conductance before and after the application, respectively. To observe the effect of AcHA on dry skin condition, replicas of the treated region were also taken from each volunteer. From the observation of the replicas obtained from the skin before and after the treatment, the effect of improvement on the volunteer skin was evaluated. RESULTS EFFECT OF AcHA ON DYNAMIC ELASTIC MODULUS OF STRATUM CORNEUM The time course of dynamic elastic modulus of the stratum corneum after the application of water or AcHA is shown in Figure 2. Evidently, the stratum corneum was plasticized rapidly by a 2-pl water application. Then, the dynamic elastic modulus gradually returned to the initial value as water evaporated and slightly exceeded the initial level. The minimum elastic modulus ratio was 0.45 at 20 min, and the recovery time was 60 min. On the other hand, there was a remarkable improvement in the skin-softening effect when the sample was treated with 0.2 w/w% AcHA of which the degree of substitution of acetyl groups (DS) was 2.6 - 3.8. This figure shows that the dynamic elastic modulus ratio of the sample, which was treated with AcHA of which the DS was 3.0 - 3.5, was around 0.3 for 100 min. The results indicate an excellent skin-softening effect for the stratum corneum. Upon the use of AcHA with a DS of 3.8, the modulus ratio returned to 1.0 at 100 min. Figure 3 shows the relationship between the modulus ratio at 100 min and the DS value of AcHA. There was an optimal DS value around 3.3. Figure 4 shows the synergic effect on skin softening of the combination of various humectants. When 0.2 w/w% HA was applied, the recovered value of softness was slightly lower than that of water. Since HA returned to approximately 0.9 at 60 min, HA had a low skin-softening effect. On the other hand, the stratum corneum treated

176 JOURNAL OF COSMETIC SCIENCE 1.2 1.0 0.8 0.6 0.4 0.2 0.0 ' 0 50 100 Time / min ! Figure 2. Time course of the changes in the dynamic elastic modulus of guinea pig stratum corneum treated with water or sodium acetylhyaluronate (AcHA). The y-axis shows the ratio of the dynamic elastic modulus, E't/E't_o, where E't is the dynamic elastic modulus of stratum corneum treated with a test solution at a specific time after application, and E't_ o is that of non-treated stratum corneum. The stratum corneum was treated with 2 pl of water or AcHA solution. The x marks represent the modulus of the stratum corneum treated with water. The closed upside-down triangle (¾) shows treatment with AcHA, of which the degree of substitution of acetyl group (DS) is 2.6. For the open circle (O), DS is 3.0 for the closed circle (0), DS is 3.5 for the open square ([•), DS is 3.7 for the closed triangle ('), DS is 3.8. with 5 w/w% glycerol maintained the modulus value at about 0.7, showing its relatively high skin-softening effect. If HA was used with glycerol, a synergic effect on skin- softening appeared, which has been reported by Ozawa eta/, (13). If AcHA was used with glycerol, it also showed the synergic effect. EFFECT OF AcHA ON THE WATER-HOLDING CAPACITY OF HUMAN SKIN Figure 5 shows the time course of the ratio of human skin surface conductance before and after the application of the sample. Treatment with HA slightly lowered the initial value of the skin conductance ratio over the measuring period. On the other hand, treatment with AcHA showed an immediate sharp increase of the ratio 10 min after the applica- tion. Then, the value of the ratio gradually decreased to approximately 1.4 at 60 min and remained the same up to 120 min after the application. Analysis of the in vivo water- holding capacity demonstrated that AcHA significantly increased the water-holding capacity of human skin and that the capacity was greater than that of HA for 60 min after the application.