j. Soc. Cosmet. Chem., 44, 181-195 (July/August 1993) Investigations into biomechanisms of the moisturizing function of lanolin E. W. CLARK and I. STEEL, Westbrook Lanolin Company, Laisterdyke, Bradford, West Yorkshire BD4 8AU, England Received January I2, I993. Synopsis A study of the lanolin/human stratum corneum system has been made, both in vitro and in vivo, to investigate where lanolin absorbed by skin lies within the tissues, and to determine possible mechanisms of the moisturizing action. Skin treated with lanolin rendered electron-opaque by lipophilic doping with either lead lanolate or lead oleate was examined by freeze-fracture and transmission electron microscopy of tissue specimens in order to identify the location and state of lanolin within the stratum corneum. Evidence was found that most of the electron-opaque material, presumed to be doped lanolin, occupied intercellular spaces down as far as the stratum granulosum, replacing natural lipid structures lost during sample preparation. In some cases applied material had penetrated into corneocytes and also appeared to have been taken up into the lipid strata of some trilaminar cell envelopes, some of which remained intact. Lanolin appeared to have a special affinity for regions of cell junction. Although this could possibly have been lead marker which had migrated, there appears to be no clear reason why the marker should have greater affinity for natural skin lipids than for lanolin. There was further evidence that lanolin within intercellular spaces had spontaneously emulsified epidermal water as a w/o emulsion with a droplet size of approximately 40 nm, thus providing a possible moisture reservoir. INTRODUCTION The emollient or moisturizing action of lanolin on human skin is well established (1). Nothing is known of the biomechanisms involved, but considerable indirect evidence exists: 1. Microprofilometry and intracorneal cohesography of skin treated with lanolin or some lanolin derivatives demonstrated pronounced smoothing (2). 2. Reduction in TEWL of 30% to 32% after lanolin application has been reported (3,4). 3. It has been shown (2) that films of lanolin statically immersed in water become hydrated by spontaneous absorption of water to form a fine w/o emulsion and, conversely, that hydrous lanolin emulsions exposed to air lose water by migration through the substrate and evaporation from the surface lanolin thus permits a two-way transport of water. The droplet size in spontaneously formed emulsions was measured at 50 nm to 6 }xm, this dimension being significant in relation to obser- vations described herein. 181
182 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS 4. The penetration of topically applied lanolin down as far as the stratum lucidum has been demonstrated by tape stripping and chemical analysis (5). 5. Lanolin has some components in common with the natural intercellular lipids, such as free and esterified cholesterol and free and esterified fatty acids some lanolic acids are hydroxy acids, and others are esterified with diols to form diesters with two long acyl chains, as in the case of ceramides. This earlier work led to the hypothesis that the moisturizing effect of lanolin was a result of absorption into the stratum corneum, where it reduced TEWL by partial occlusion of the internal water pathways or other means, and possibly also of lanolin within the stratum corneum absorbing natural epidermal moisture and thus acting as a reservoir, releasing water if the moisture gradient across the stratum corneum should favor such loss. The present work explores this hypothesis in three separate stages. EXPERIMENTAL STAGE 1 This was an initial in vitro exploratory stage involving the application of lanolin to excised skin, which, after suitable fixing and sectioning, was examined by transmission electron microscopy. The lanolin was insufficiently electron-opaque, however, to be differentiated from other structures within the stratum corneum specimens. Therefore, this approach was discontinued and details are not reported here. STAGE 2: OUTLINE This stage of the study was subdivided into two sections involving different method- ologies. Section 1 consisted of treating the surface of human skin in vivo with anhydrous lanolin, and after a time removing the surface lanolin and examining it by freeze-fracture and scanning electron microscopy in order to study the microstructural characteristics. Section 2 involved the doping of lanolin with lead to increase its electron opacity before application to human skin in vitro at a specific loading. Prepared specimens of treated skin sections were then examined by transmission electron microscopy. The principle of lead doping to increase electron opacity had been previously used by Ghadially and co-workers (6) in studies of skin penetration by petrolatum. In those studies, doping was carried out by adding lead nitrate in the form of a physical sus- pension. Lead nitrate is soluble in water but insoluble in petrolatum and other lipids, and thus there is the possibility of migration of some of the lead salt out of the petrolatum into hydrous areas. In this part •)f our work, such a potential problem was avoided by doping lanolin with lead lanolate, which is the lead salt of the naturally occurring fatty acids of lanolin, at the level of 1.3% of lead as Pb. Lead lanolate has good solubility in lanolin but negligible solubility in water, and the possibility of migration into hydrous areas may be assumed to be negligible. There is at least the possibility of selective migration of lead salt into other lipids, but there seems to be no reason why these should have a greater affinity for lead salt than in the case of lanolin. Moreover, in a later section of
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