**2. Function of human sebaceous fatty acid "C16:1∆6"**

Human skin lipids, including sphingosine and C16:1Δ6, provide a defense against external factors and function to maintain homeostasis. It is well established that human skin lipids contribute to the defense mechanism termed "self-sterilization" (Wille & Kydonieus, 2003) and the antimicrobial activities of each lipid component have been evaluated (Arikawa et al., 2002; Harder et al., 2010). A significant difference was found in the fatty acid composition of sebum lipids between AD patients and healthy controls (Takigawa et al., 2005). We describe the results of our investigation in the section below.

Improvement of Atopic Dermatitis by Human Sebaceous Fatty Acids and Related Lipids 311

and the total level of C16:0 ranged from 0.88 to 7.64 μg/cm2 with a mean of 3.43 μg/cm2 (14.9 to 28.2% with a mean of 22.1% in percent of total). In contrast, the level of C16:1Δ6 (0.08 to 0.53 μg/cm2 with a mean of 0.26 μg/cm2) and the percent of total (1.1 to 8.7% with a mean of 3.0% in percent of total) in the skin of AD patients were lower than in the healthy controls, although the amounts of C16:0 (0.53 to 2.84 μg/cm2 with a mean of 1.44 μg/cm2) and the percent of total (6.8 to 30.7% with a mean of 16.5% in percent of total) were similar to those of the healthy controls. A significant decrease in free C16:1Δ6 content in nonlesional

Colonization of *S. aureus* has been reported in AD patients and is regarded as an exacerbation factor (Akiyama et al., 1996). Antimicrobial agents, including C16:1Δ6, inhibited the growth of *S. aureus* at a final concentration of 100 mg/l in the following order; Benzalkonium chloride = Triclosan > C16:1Δ6 > Trichlorocarbanilide, while they inhibited the growth of *S. epidermidis* in the following order; Benzalkonium chloride = Triclosan> C16:1Δ6 = Trichlorocarbanilide (Fig. 3). Whereas C16:1Δ6 could inhibit the growth of *S. aureus* over a concentration range from 100 to 1000 ppm, it did not greatly

In addition, the antibacterial mechanism of C16:1Δ6 against *S. aureus* and *S. epidermidis* was investigated using protoplasts treated with lysostaphin. The results of this experiment indicated that there was no difference in the effectiveness of C16:1Δ6 against *S. aureus* and *S. epidermidis*, and suggested that the antibacterial action of C16:1Δ6 against Staphylococcal bacteria was caused by disruption of the cell membrane, not disruption of the cell wall, and

**7**

**B**

**Log number of survivors(CFU/ml)**

Fig. 3. Comparison of the antimicrobial effects of C16:1Δ6 against *Staphylococcus aureus* ATCC 12600T (left panel) and *Staphylococcus epidermidis* JCM 2414T (right panel).

An inverse correlation between *S. aureus* colonization and the level of C16:1Δ6 in AD patients was found (Fig. 4). In addition, the colonization of *S. aureus* was reduced by topical application of C16:1Δ6 to the skin of six to eight AD patients in a small-scale clinical study. Based on these investigations, it was considered that the decreased level of

**C16:1Δ6**

*Staphylococcus epidermidis* **JCM 2414T**

**Time(hr) 0 5 10 15 20 25**

**50mM phosphate buffer(pH4.5) Test concentration : 100 ppm**

**Triclosan**

**Benzalkonium chloride**

**Trichlorocarbanilide**

that this disruption resulted in leakage of bacterial components (Data not shown).

**C16:1Δ6**

*Staphylococcus aureus* **ATCC 12600T**

**Triclosan**

**50mM phosphate buffer(pH4.5) Test concentration : 100 ppm**

**0 5 10 15 20 25 Time(hr)**

**2.4 Topical application of C16:1∆6 to AD patients** 

**Benzalkonium chloride**

**Trichlorocarbanilide**

skin from AD patients compared with healthy controls was found.

inhibit *S. epidermidis*.

**7**

**A**

**6 5**

**4 3**

**Log number of survivors(CFU/ml)**

**2**

**2.3 Antibacterial activity of C16:1∆6: Selective antibacterial activity** 
