]. Soc. Cosmet. Chem., 27, 509-531 (November 1976) Characterizing Cosmetic Effects and Skin Morphology by Scanning Electron Microscopy CHARLES A. GABBER, Ph.D. and C. T. NIGHTINGALE, B.S.* Presented May 30, 1975, SCC Seminar, St. Louis, MO Synopsis: The SCANNING ELECTRON MICROSCOPE has developed into an im- r•ortant tool for characterizing the effects of COSMETIC PRODUCTS on HUMAN SKIN. Methods are described for "replicating" areas of stratum corncure as in vivo impressions, or "negative replicas." The "negatives" are replicated again to produce "positives," which are then studied in the scanning electron microscope. Through the use of selected control subjects, it has been possible to define the differences between dry and normal skin, the former generally exhibiting larger uplifting desquamating layers (flakes) of stratum corneum. Experimental findings note that beneficial clinical effects are generally accom- panied by a reduction in the amount and/or size of the desquam.ating material. Examples will be given for (1) moisturizing produ,cts, (2) protein additives, (3) abradent cleansing products, and (4) moistnrizing soaps. The effects, in some instances, are so subtle that unless the microscopy is done as Before and After pairs on the same identical areas, the effects will not be recognized. Once good micrographs are obtained, it is sometimes possible to quantirate the resuks through the use of Quant/met Image Analyzing Com- puter, t a tool originally developed by metallurgists, but which has great potential for quantifying cosmetic effects. INTRODUCTION The characterization and definition of the effects of cosmetic products on human hair and skin has always been one of the most important responsibili- ties of the cosmetic chemist. Although the actual formulation of new prod- ucts is generally based on fundamental principles of physical and surface chemistry, the measurement of the efficacy of such materials has generally been performed by subjective clinical evaluation. Only in recent years, in part •Structure Probe, Inc., P.O. Box 342, West Chester, PA 19380. timanco, Inc., Monsey, N.Y. 509

510 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS due to the rising demands for the scientific documentation of advertising claims, has interest been focused on the development of reliable quantitative measurements for product efficacy on human hair and skin. We have found electron microscopy, and in particular, scanning electron microscopy (SEM) to be a powerful tool for scientifically documenting the effects of materials applied to human hair and skin. The transmission elec- tron microscope (TEM) also has its place, but since SEM is basically a sur- face (topographical) tool, and since studies of products on hair and skin in- volve surfaces (protein deposits on the outside of a hair shaft skin roughness is a surface phenomonen), the SEM is generally the instrument of choice. The SEM and TEM, rather than "competing" with each other, are actually complementary, and most important studies ultimately involve the use of both. The present work is not the first application of SEM for cosmetic studies. In fact, as early as the 1940s (1, 2) various methods of replicating the outer cuticle structure of hair were devised through the use of platinum and carbon techniques. The SEM, which really started to develop as a scientific tool in the late 19fi0s, was quickly utilized to examine human hair, and Wolfram and Lindemann (3) published a study of the structure and morphology of hair. Further studies have demonstrated yet additional information on hair (4, 5) and, more recently, some very novel studies have been published on the elucidation of the actual structure of the cuticle (fi). One of the most in- ventire approaches was for the study of porosity within human hair (7), which describes a method for the polymerization of methacrylate toohomer in situ. within the holes or pores that exist within the hair structure. The hair is then degraded away, leaving the now "plastic pores" (or "ghosts") avail- able for collection and characterization by SEM. A method for demonstrating the repair of damaged hair and a classification scheme for identifying and quantitating different types of damage has now been reported by DiBianca (8). This latter method was used to substantiate the claim that a hair care product can repair damaged hair. Although the amount that is now known and published about human hair is substantial, by comparison very little SEM and/or TEM work has been published about human skin. Virtually no SEM or TEM studies have been published that demonstrate the effects or efficacy of specific products on the stratum eorneum. This absence of data is not for lack of interest, but because of the enormous experimental difficulties encountered with skin studies. These difficulties have, therefore, severely limited the application of SEM and/or TEM techniques to the skin area. For TEM studies, a biopsy sample must be (1) "fixed" (hardened to keep it from degrading) (2) embedded (so it can be sectioned) (3) sectioned (4) "stained" to bring out contrast and (5) photographed within the high vacuum of the electron microscope. Put quite simply, the changes brought about by the fixing, embedding, and staining are enormous as compared to