j. Soc. Cosmet. Chem., 46, 181-190 (July/August 1995) Photochemical alterations in human hair. Part I1' Analysis of melanin EDO HOTING, Hans Schwarzkopf GmbH, Hamburg, Germany,' MONIKA ZIMMERMANN and HARTWIG HOCKER, Deutsches Wollforschungs Institut an der TH Aachen e.V., Aachen, Germany. Received February 28, 1995. Synopsis A brief investigation was carried out of changes in the melanin pigment that occur during the irradiation of hair with simulated UV-B, UV-A, visible light, infrared light, and total sunlight. After six weeks of irradiation, the pigments were isolated from hair samples and the changes in their physicocharacteristics were evaluated. The melanin appears to be affected most by visible light, although the pigment of black and dark brown hair (predominantly eumelanin) is less changed than that of light-brown hair (a mixture of eu- and pheomelanins). The weight losses of melanin correlate with photochemically induced alterations of the melanin polymer. IR spectroscopic investigations characterize the eumelanin by five peaks and the pheomelanin by six peaks in the range of 1800 to 500 cm-•. The intensity of some of these peaks decreases, depending on the irradiation wavelength and on the origin of melanin pigments. INTRODUCTION Melanin constitutes the primary pigment present in the skin and hair of vertebrates. The particular color of the tissue is determined by the quantity and location of the pigment granules as well as by the chemical nature of the pigment (eu- or pheomelanin) (1-3). The black eumelanin is insoluble in acids and alkali and contains no sulfur (2). The reddish-brown pheomelanin is alkali-soluble and contains 8-12 % sulfur (2). The highly complex structures of these forms of melanin remain unresolved enzymatic conversion of tyrosine or 5,6-dihydroxyindole (DHI) led to formation of pigments of random and unpredictable structure. According to Crippa (3) and Boudier (4), the eumelanin is a product of copolymerization of DHI and DHI-caboxylic acid. Cysteine is involved in the formation of pheomelanin by reaction with dopaquinone to form benzothiazine deriv- atives the process intermediates polymerize to form the reddish-brown pigment (3). While it is generally accepted that melanin has a photoprotective effect, the mech- anism of its action has not been elucidated. It is likely that in this respect several modes 181

182 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS of action are operative, and it was shown that melanin has the ability to scavenge molecular oxygen as well as photochemically formed reactive oxygen species such as 02- and H202 (5-8). The photooxidation of melanin is accompanied by bleaching (6,9,10), a process well known and practiced in the hair care field. Eumelanin and pheomelanin respond in a different manner to irradiation. Chedekel et al. (11) found that under alkaline conditions the photolysis of pheomelanin leads to a hundredfold higher production on biologically active OH' radicals and O2-anions than eumelanin. The highly damaging potential of these compounds led Menon et al. (12) to suggest that pheomelanin might be a poor protectant against sun-induced skin cancer. A somewhat different perspective on the reactivity of these pigments was provided by Wolfram and Albrecht (13), who examined the photo and chemical oxidation of pheo- and eumelanins in hair. They found pheomelanin to be more resistant to such oxidative attacks than eumelanin. The purpose of this investigation was to obtain a better understanding of the effect of sunlight on the melanin and hair. Our goal was to provide some explanations on the molecular level. We were fully aware of the difficulties associated with the lack of precise knowledge regarding the complex structure of melanin as well as with the uncertainties of the effect that different segments of the sunlight spectrum may have on the photo- reactions of the pigment. We have thus limited our experiments to black and light- brown hair and carried out the irradiation study with UV-B, UV-A, visible light, and IR segments of the sunlight spectrum. The work was performed in a specially designed irradiation unit, which was described in detail in the first part of this study (14). An enzymatic technique was used for the isolation of the melanin pigments from hair (15). The isolated pigment was further purified by removal of proteins and lipids. The melanin granules were weighed and characterized by means of IR spectroscopy. The results are discussed in relation to the nature of the pigments and irradiation parameters. MATERIALS AND METHODS HAIR SAMPLES Untreated black and light-brown hair was obtained from Herzig Co. as 25-cm-long tresses of European origin. PURIFICATION OF HAIR The hair was extracted for 5 min with dichloromethane and for 30 min with diethylether to remove the sebum and traces of naphthalene, a conservation agent applied by the hair trader. Finally, the hair was washed with a non-ionic detergent, rinsed, and stored at ambient humidity. IRRADIATION OF THE HAIR The hair tresses were irradiated for a period of 6 weeks (1008 h) in individual com- partments with UV-B, UV-A, visible light, IR, or global radiation at RH 70 %