Dark- or Light-Colored Melanins: Generating Pigments Using Fe2+ and H2O2

We have studied the formation of melanin-like pigments from catechol or pyrogallol and a wide range of other phenolic compounds using Fe²⁺ and H₂O₂. Combining UV_Vis spectroscopic measurements and size-exclusion chromatography analyses we evaluated the impact of the intensity of the oxidation reaction by varying the concentration of H₂O₂ present in the reaction mixtures. All compounds tested, except tyrosine, reacted readily leading to mixtures that were black, brown or yellow-orange in color. For many compounds tested, the use of increasing concentrations of H₂O₂ resulted in either precipitation of the pigment or the formation of a soluble, lighter-colored pigment. With catechol or pyrogallol as model compounds, and using different concentrations of H₂O₂, several materials were synthesized, purified and dried. The physic-chemical properties of these materials were compared to the properties of melanin-like pigments synthesized from the same precursors using air-oxidation in an alkaline environment. For both precursors, a distinct chemical change, as judged from FT-IR spectroscopy, was introduced in the melanin structures when using H₂O₂ as the oxidizing agent and the relative intensity of this distinct signal strengthened with increasing concentration of H₂O₂ used in the reaction. In general, our results suggest that depending on the precursor molecule and the intensity of the oxidizing reaction conditions involved, light- or dark-colored melanin-like pigments can be generated. This may be an important factor when evaluating the visible outlook of histological or archeological specimens: the presence of a lighter color or the absence of a dark color may not necessarily mean the absence of melanin-like biomolecules.