HAIR BLEACHING AND WAVING 165 oeH 3 NH NH oe0 HE NH oe0 I oeH• [H 2 S S [H 2 EO I EH I NH I EO EH 3 Figure 3. N,N'-bisacetyl-L-cystine-bismethylamide (ACM). hair research allows the separation of unfractionated protein extracts. Radiolabeling of the investigated keratins increases the detection sensitivity so that very small samples are sufficient, allowing for high resolution of the protein components during electro- phoresis. The first step of the procedure is the reductive solubilization of the keratins followed by the alkylation of the cysteine residues thus obtained with iodo(2-•4C)-acetic acid. The separation parameter for the first dimension is the charge of the proteins, whereas in the second dimension it is the apparent molecular weight in the presence of sodium dodecyl sulfate (SDS). Proteins are located by fluorography (Figure 6). For full experimental details, refer to Marshall et al. (18). After investigation of more than one hundred different single hair samples under iden- tical conditions, it was found that a basic pattern was always repeated (18-20). How- ever, different exogenous or endogenous influences cause variations. Fluorographs showing the changes in patterns obtained with untreated (Figure 6) and bleached human hair (Figure 7) illustrate some of the possibilities of this technique for detecting the chemical changes that occur during the cosmetic process. The changes in the fluorographs of the bleached samples compared with those of un- treated hair are: Area 1.1 is weaker, areas 1 and 2 are unchanged with regard to the molecular weight of the proteins but are obviously modified with respect to the charge,

166 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS lOO- 5'0 80- 60- 20- ,, su[fonic ucid ø •' • "'•"'••• • '•,, .'•'•.• disu monoxide I I I 1 2 3 lime / hours Figure 4. Alkaline peroxide oxidation (pH 9.5) of the model disulfide N,N'-bisacetyl-L-cystine-bismeth- ylamide (ACM). Formation of oxidation products versus treatment time (oxidation products are plotted as percentage of the original disulfide concentration) (Schumacher-Hamedat (14)). Treatment solution: 10-2mol/l ACM, 10 ml/1 H202 (35%), 0.5 ml/1 ammonia (25%), 2g/1 tetrasodium pyrophosphate pH 9.5 . 30øC. Analytical procedure: HPTLC (chloroform: methanol:acetic acid/95: 3: 5/v:v:v) detection with tert.-butylhypochlorite, o-toluidine, KI in acetic acid quantification by densitometry. areas 4 and 5 are absent, and areas 6 and 7 are weak and also modified in terms of the charge. These results can be explained as being the expected changes in electrophoretic behavior resulting from the conversion of the native proteins to cysteic acid-containing proteins. The latter cannot form the fiuorographically detectable S-carboxymethyl derivative. Thus an absence of spots (e.g. areas 4 and 5) or a weaker appearance (e.g. areas 1.1, 6, 7) results, depending on the amount of oxidized sulfur. Shifting of proteins, as in areas 1, 2, 6, and 7, is a consequence of the changes in the ionic character of the sulfonic acid-containing proteins. For further details on the changed electrophoretic behavior of oxidized keratins, refer to Wittig et al. (19). PERMANENT WAVING OF HAIR The permanent waving process usually comprises two different steps: During the first the cystine of hair is partially reduced and subsequently, disulfide bridges are oxida- tively rebuilt during the second step. The most common reagents used are alkaline ammonium thioglycollate during reduction, and acidic solutions of hydrogen peroxide