j. Soc. Cosmet. Chem., 46, 39-52 (January/February 1995) A simple and sensitive method using protein loss measurements to evaluate damage to human hair during combing SUKHVINDER S. SANDHU, RAM RAMACHANDRAN, and CLARENCE R. ROBBINS, Technology Center, Colgate Palmolive Company, Piscataway, NJ 08854-1340. Received June 1 O, 1994. Presented at the Annual Meeting of the Society of Cosmetic Chemists, New York, December 8-9, 1994. Synopsis A simple method to quantify hair damage during combing or brushing has been developed. The method involves collecting hair fragments that are chipped from hair during combing and quantitatively measuring the amount of protein using a colorimetric procedure capable of detecting as little as 5 Ixg of protein per mi. Using this procedure on hair tresses in the laboratory and on live heads (half-head tests), we were able to demonstrate significant differences in protein loss during post-shampoo combing of undamaged and chemically damaged hair previously treated with various shampoos/conditioners. The method is applicable to different types of hair tested, namely, Caucasian, Asian, Oriental and Negroid. INTRODUCTION The cuticle forms a protective barrier for each hair fiber. The cuticle governs the frictional properties of hair fibers and is also largely responsible for maintaining the structural integrity of hair. Many investigators have shown that the cuticle is gradually chipped, fragmented, and worn away by the abrasive action of combing, brushing, shampooing, and teasing of human hair (1-4). It is also known that gradual loss of cuticle layers can eventually lead to complete fibrillation of the fiber (4,5). Various investigators have used a number of techniques such as scanning electron microscopy [SEM (1,3,4)], combing measurements using an Instron Tensile Tester (4,6,7), fric- tional measurements (8), tensile measurements (9, 10) and measurement of the sound or raspiness caused when combing (11) to evaluate hair damage due to combing and/or the conditioning effects of various products. Some of these techniques require expensive instrumentation (e.g., SEM), some are qualitative (11), and others lack sensitivity [tensile measurements (10)], simplicity (SEM), and/or the precision desired in an ana- lytical technical for routine use to study surface damage. Previously we published a simple technique to assess surface damage to hair caused by oxidative/reductive treatments such as bleaching and permanent waving (12). Since hair 39

40 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS is primarily a proteinaceous material, we hypothesized that measurements of the amount of protein abraded from hair could be used as an index to assess damage caused by abrasive actions. The method involves shaking hair in water (wet abrasion) and quan- titatively measuring, via a colorimetric procedure, the amount of protein abraded/eroded from hair during shaking. During the course of these studies, it was observed that, unless the hair is very badly damaged and totally stripped of cuticle layers, the cuticle is the primary source of protein loss during abrasion (12). We have extended this work to show that protein loss measurements can also be used to quantify hair damage due to combing. We have demonstrated that this technique can also be used to measure hair damage on live heads (half-head tests) from combing. Furthermore, this technique can differentiate the conditioning/protective efficacy of current conditioning shampoos. MATERIALS AND METHODS Caucasian and Oriental Hair fibers used in these studies were commercial samples purchased from DeMeo Brothers, New York. Asian (Indian) and Negroid hair were taken from private individuals. All hair samples used in these studies, with the excep- tion of Negroid hair, were approximately 9-11 inches long the Negroid hair samples were --5.5 to 7 inches in length. Using these hair samples, hair tresses, each weighing 3.0 g (1.0 g for Negroid hair), were prepared for various experiments. Reagents for protein estimation were obtained from Sigma Chemical Company, St. Louis, MO. All other reagents used were of analytical grade. EVALUATION OF POST-SHAMPOO PROTEIN LOSS DURING COMBING Laboratory studies using hair tresses. Laboratory studies to determine protein loss from hair during combing were conducted using a minimum of three tresses for each shampoo treatment as follows: The tresses were wet individually under running tap water (-- 105øF), pre-washed with SLES (20% @ 1 ml per tress, 0.5 ml for Negroid hair) and rinsed extensively for one minute under running tap water to remove any surfactant residue. Each tress was then shampooed individually, with the test product (1 ml per tress, 0.5 ml for Negroid hair) applied uniformly with a syringe, the hair gently rubbed between the fingers and occasionally on the palm of the hand to generate foam for a total of one minute and rinsed extensively for at least one minute to remove any shampoo residue. Finally, each tress was combed gently under running tap water to detangle the hair, and the excess water was squeezed out. To induce protein loss during combing, each tress was combed wet, by hand, using a fine-toothed nylon comb (20-22 teeth per linear inch) @ 50-100 strokes (always the same number of strokes in each test). During combing, the tress was held firmly in one hand and combed with the other hand, using a firm, smooth, vigorous combing action for the entire length of the tress and occa- sionally changing the side of the hair tress being combed. After every five strokes, the comb was dipped into a beaker containing 50 ml of distilled water, and after ten strokes, the tress was also dipped into the same water to recover loose/chipped protein fragments from hair. The water suspension from each tress was then tested for turbidity in a spectrophotometer at 600 nm and for protein concentration as described below. The data was statistically analyzed using an in-house computer program for one-factor analysis of variance (1-way ANOVA) to establish whether the observed differences between treat- ments are significant at 95% confidence level (p = 0.05).