MOISTURIZING FUNCTION OF LANOLIN 187 ß ! ! Figure 4. Lanolin/marker incorporated in membranes attached to, and detached from, the skin surface. control (Figure 6), where the intercellular spaces appear white. There also appears to be a heavy concentration of electron-opaque material at a junction between the corneocytes (arrowed) in Figure 5 (perhaps a desmosome remnant), and more evidence of this is reported in Stage 3. STAGE 3 Outline and methodology. Although this was basically a repeat of Stage 2, Section 2, freeze substitution was used throughout in the preparation of specimens, and there was a change in the lead-doping technique. Instead of using lead lanolate, doping was carried out with lead oleate, which has better lipid solubility and is easier to wash free from residual inorganic lead salts. Examination of thin films of the lead-oleate-doped lanolin by electron microscopy showed it to be quite homogeneous, with no dense particles. Tissue specimens were not stained, so that electron-opaque areas indicate the presence of lead oleate and presumably, therefore, of lanolin, unless selective migration of lead salt into other lipids has occurred, against expectation. Results. Figure 7 confirms the penetration of lanolin deep into the stratum corneum. It also confirms findings reported in Stage 1, Section 2, of the existence of a structure with high lead and presumably lanolin content forming a layer on, or near, the surface of some corneocytes bounding intercellular spaces, more clearly seen in an enlargement of the area marked 'L' in Figure 8. Similar layers on or close to, and parallel with, corneocyte surfaces are very clear in Figure 9 and 10, where signs of a lameliar structure

188 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS Figure 5. Concentration of lanolin/marker (arrowed) at cell junction, and lanolin visible in intercellular spaces down to the stratum granulosum. can be seen. Also visible in Figure 10 is apparent evidence of water droplets emulsified in the lanolin. The corneocytes themselves appear to be permeable to lanolin. Figures 11 and 12 show what seem to be concentrated accretions of lanolin (arrowed) within some corneocytes. The penetration of corneocytes by cholesterol was postulated by Garson et al. (14). Figure 12 also shows (arrows) concentrations of electron-opaque material at cell sur- faces. One of the objects of our work was to investigate the possibility of spontaneous emul- sification of water in lanolin absorbed in stratum corneum, as is known to occur in vitro. Such an effect appears to be manifested not only in Figure 10 but also in Figure 13 (arrows), showing a mass of lanolin within an intercellular space, which is honeycombed with holes up to about 40 nm in diameter. These could be where emulsified water droplets had existed before the fixation of the specimen. Lead oleate without lanolin would not be expected to show this effect. The size of droplet observed is a close approximation to that of many of those shown in Figure 1, where epidermal water had migrated into a lanolin film on the skin surface, and strengthens the evidence pointing to spontaneous emulsion formation within the stratum corneum. If this does in fact occur, then lanolin would seem to provide a second mechanism by which essential moisture can be stored within the stratum corneum, in addition to the water held in the lipid bilayer structures.