used), hydrogen peroxide, and a range of dye precursors (1). There are tw o chemical pro- cesses that take place during the permanent dying process, both of which contribute to observed fi nal color. First i s oxidation of melanin and previously deposited dyes that lightens the underlying hair color. Second is oxidation of dye precursors that have dif- fused inside hair to form colored chromophores. The rate of color formation is important: if too fast color will form outside hair or in the bottle before application is completed, and if too slow not enough color is formed during a typical development time of 30 min. Penetration of dyes inside hair is important for color longevity as the larger size and re- duced solubility of the dyes formed mean they are trapped inside hair, making them harder to be removed during washing. The basic mechanism of dye formation is an oxida- tive reaction between hydrogen peroxide and a primary intermediate to form a reactive intermediate called a diimine that then reacts with a coupler to form diphenylamine (leuco dye) which then undergoes a rapid oxidation to form a binuclear indo dye. Depend- ing on the coupler structure, there can be subsequent reactions with additional primary intermediates to form trinuclear and larger dyes (2,3). T he role of redox metals in accelerating this chemistry has been studied. A work by Kojma and coworkers showed via isotopically labeled dye precursors, and nano-secondary ion spectrometry that formed chromophores were localized in or close to melanin gran- ules and linked this to the presence of transition metals in melanin. A previously pub- lished work has shown that consumer hair contains redox metals such as copper and iron that are adsorbed during washing in tap water (4) and that these exogenous metals are present in the outer layers of the hair cuticle (5). The objective of this work was to determine whether these exogenous redox metals are infl uencing color formation rates and color uptake. M ATERIALS AND METHODS H AIR SOURCE C hemically virgin natural white Caucasian-source hair was purchased from International Hair Importers & Pr oducts Inc. (Gl endale, NY). Individual tresses (2 gm, 6in with hot wax tab at the top), formed by evenly blending hair from multiple ponytails, were used for all experiments. Colored hair was created by treating virgin hair tresses once with an oxidative permanent commercial colorant (Nice ʹN Easy Extra Light Blonde Shade, Clairol, Stamford, CT). The mixed colorant was thoroughly massaged onto hair at a dose of 4 g of product per g of hair and then incubated for 30 min in an oven at 30°C. The product was then completely rinsed from hair. Initial copper levels in hair were 20 μg/g. DETE RMINATION OF METAL CONTENT IN HAIR The metal content of hair samples was determined by inductively coupled plasma atomic spectroscopy (ICP-OES) with an Optima 5300 DV Optical Emission Spectrometer (Perkin Elmer Life and Analytical Sciences, Shelt on, CT). Samples of 100 mg of hair were di- gested overnight with 2 mL of high-purity concentrated nitric acid. The digestive mix- ture also contained 150 μL of 100 μg/g yttrium internal standard (Inorganic Ventures, REDOX METALS IN COLOR FORMATION IN A HAIR COLORANT 269

Christi anburg, VA). Following digestion, samples were heated to 70°–80°C for 1h, coole d to room temperature, and diluted to 15 mL with deionized water. Each hair sam- ple was analyzed in triplicate. DYE SOLU TIONS A soluti on of dye precursors was made in deionized water with 3% ammonium hydroxide (30% active) and a pH of 10.5. The molarity of the primary intermediate was 0.031 M (PPD, HDAP) and coupler was 0.032 M (AHT, Res). Solutions were made with 0, 0.2, 1.0, and 2.0 μg/g of copper nitrate. In a 2-mL glass vial, 1720 μL of dye solution was mixed with 180 μL of hydrogen peroxide (35% active) and either 100 μL of DI water or copper nitrate solution. Three vials were made for each dye, and copper level and images taken of the color change over time. The images were taken inside a constant illumina- tion light booth with a USB camera. Each image included a reference color chart. This chart was checked to be in-line with the initial values before each measurement. The images were captured and analyzed with National Instrument’s LabView (Austin, TX) applica- tion using Vision acquisition software. COLOR MEASUR EMENT Color measur ements were taken using a Konica Minolta CM-700d handheld spectropho- tometer (Ramsey, NJ). Settings used were as follows: D65 light, 10° observer, 3 mm aperture, and specular excluded. A total of eight color re adings were made on each tress: four on each s ide. HAIR TREATMEN TS (i) Infl uence of copper ions in hair on color uptake Hair tresses pretreated with one cycle of N ice ʹN Easy Extra Light Blonde colorant were washed with water containing copper ions (0.06 μg/g) to create four sets of 12 × 2 g, 6-cm tresses with copper levels of 73 (±7) μg/g, 52 (±7) μg/g, 35 (±7) μg/g, and 21 (±7) μg/g. Each set of 12 tresses was divided into sets of four tresses which were th en colored with a permanent colorant. The three colorant tints teste d were an intense me- dium blonde/red (77/44), a medium brown/violet (4/6), and an intensive red (66/46). Each tint was mixed with a 20-volume hydrogen peroxide developer (6%) in a 1:1 ratio and applied at a dose of 4 g of mix to each gram of hair. After 30 min, hair was rinsed for 2 min, dried, and color measured using a Minolta spectrophotometer. The hair was then washed with a shampoo containing 10.5% SLE1S, 1.5% SLS, and 1.0% cocami- dopropyl betaine surfactants for a total of 14 cycles with color fade measured using a Minolta spectrophotometer after 2, 6, 10, and 14 cycles. Each wash cycle consisted of applying 0.1 g/g shampoo to the hair switch and lathering for 30 s, followed by a 30-s rinse repeated for a tot al of two shampoo applications. Hair was then dried in a hotbox at 80°C. (ii) Infl uence of histidi ne chelant on color uptake JOURNAL OF COSMETIC SCIENCE 270