COSMETIC FILMS ON THE SKIN (16, 17), the principle in sebum which makes it porous was found to be the presence of branched-chain aliphatic compounds. The development of synthetic branched-chain compounds similar to those found in sebum was undertaken. Formulations of cosmetic bases incorporating these compounds were applied to skin, and the porosity of these films on the skin was checked. EXPERIMENTAL Collection and Analysis of Natural JVMF The NMF of the stratum corneum was extracted from the skin of 35 healthy human subjects, ranging in age from 14 to 65 years of age. Areas on the forearms and the backs of these subjects were used. Open-end cylinders with diameters from 3-5 cm were pressed on the skin with gentle pressure. The cylinders were filled with ether and left in contact with the skin with occasional shaking for 5 minutes. The ether was removed, and after the skin was dry, ethanol was added to the cylinders and left in contact with the skin for 5 minutes, with occasional shaking. After the alcohol was removed and the skin was dry, distilled water was added to the cylinders and left in contact with the skin for 15 minutes with occasional shaking. The water extracts were removed from the skin, pooled, and immediately evaporated under high vacuum at room temperature. The residue was dissolved in water and chroma- tographed on a Dowex 50 column. A total of 200 fractions was col leeted. Each fraction consisted of 5.7 ml. For the first 50 fractions 0.3/V HC1, for the next 50 fractions 0.5/V HC1, for the next 50 fractions 1.0 _N HC1, and for the last 50 fractions 2.0 _N HC1 were used. The con- tent of each fraction was quantitatively determined and its chemical composition determined with paper chromatograms. Figure 1 shows the results of this analysis. This analysis served as the basis for the preparation of a synthetic NMF. In the following experiments this synthetic NMF was used.* E•cacy of Synthetic rs. JVatural JVMF The efficacy of synthetic NMF (trade name, Aqualizer E-J) has been determined in 171 single tests on living human skin. Extraction of Skin--The sebum and natural moisturizing factor were extracted from the inner forearm or back of healthy human subjects. * The use of this product as a moisturizing material in cosmetic preparations is covered by U.S. Patent 3,231,472, English Patent 1,004,774, and French Patent 1,329,616.

152 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS Adjacent areas on the same subject were extracted. Glass cylinders, open at both ends (4 cm diameter), were attached to the skin with rubber bands. Five cc of ether was poured into each cylinder and kept in contact with the skin for 2 minutes. The ether was then removed, and this extraction was repeated five times. After the ether extraction, the same area was extracted with distilled water. Five cc of water was placed in each cylinder and left in contact with the skin for 2 minutes. 5•gar amino acid compounds Lactic Tyc, I Lgs. Arg. $•, .GI.y. aCidca+. K + •,a AW • i•,. Gtu. rh•on. Va,. •o. Leu. iLeu 600' • I I I I I I •0 •20 o.sct Figure 1. Analysis of NMF on Dowex 50. Column 7 X 700 mm p = 1200 H20 sample weight = 320 mg fraction volume = 5.7 mi. The water was then removed and the skin dried with a warm air current for 2 minutes. This extraction with water and alternate drying was re- peated ten times. The skin was then allowed to remain untouched for 30 minutes. Reimpregnation--Next, a given amount of a 5% test solution (6 mg/ cm 2, on dry basis) was applied to one of the extracted areas and massaged in with a glass rod. The skin was then dried for 5 minutes with a warm air current (at 39 øC) and for five additional minutes at room tempera- ture. The application of test solution and massaging was repeated three times. After the third application of test solution, the skin area was blotted with filter paper to remove all excess test solution. This blotting was done before drying the skin for the third time. Removal of Skin Cells--Ninety minutes after the reimpregnation, the stratum corneum of the skin was scraped off with a sharp scalpel or razor blade. To eliminate material that only clings to the surface, the outermost layer of cells scraped off was discarded. Skin cells from the adjacent extracted but not reimpregnated area and skin cells from un- touched adjacent skin •'ere also scraped off to furnish material for comparison. The three types of skin (untreated extracted only and