Eumelanin and pheomelanin in tissue samples can be specifically measured as the markers pyrrole‐2,3,5‐tricarboxylic acid (PTCA) and 4‐amino‐3‐hydroxyphenylalanine after acidic permanganate oxidation and hydroiodic acid hydrolysis, respectively. Those degradation methods, although widely applied, are not easily performed in most laboratories. To overcome this difficulty, we developed alkaline H₂O₂ oxidation in 1 M K₂CO₃ that produces, in addition to the eumelanin marker PTCA, thiazole‐2,4,5‐tricarboxylic acid (TTCA) and thiazole‐4,5‐dicarboxylic acid (TDCA) as markers for pheomelanin and pyrrole‐2,3‐dicarboxylic acid (PDCA) as a marker for 5,6‐dihydroxyindole‐derived eumelanin. Those four degradation products can be easily separated by HPLC and analyzed with ultraviolet detection. The alkaline H₂O₂ oxidation method is simple, reproducible and applicable to all pigmented tissues. Its application to characterize eumelanin and pheomelanin in human hair shows that PTCA and TTCA serve as specific markers for eumelanin and pheomelanin, respectively, although some caution is needed regarding the artificial production of TTCA from eumelanic tissue proteins.