Eumelanin separation was performed on a Waters e2695 system with 2478 UV detector at 269 nm using a C18 column (Atlantis T3, , 5-μm particle size). PTCA was determined in isocratic mode with a mobile phase of 18% methanol in 20-mM potassium phosphate buffer (pH 2.12) at a flow rate of under room temperature (injection volume: 80 μL). To determine pheomelanin, samples were analyzed with an Agilent 1100 series HPLC system consisting of, a C18 column (Atlantis T3, , 5-μm particle size), an electro-chemical detector (Agilent 35900E interface and LC-4C Amperometric Detector), a reference electrode: Ag/AgCl and a platinum working electrode. 4-AHP was eluted at a flow rate of in gradient mode: 0 to 30 min, from 100% A to 100% B, 30 to 40 min, 100% B, equilibrate column with 100% A for 15 min before the next run. The composition of the mobile phases were A: 0.1% acetic acid in 5% methanol with 1.5-mM octanesulfonic acid sodium salt and 0.1 mM EDTA (pH 4.0); mobile phase B: 0.1% acetic acid in 90% methanol with 1.5-mM octanesulfonic acid sodium salt and 0.1 mM EDTA (pH 4.0). The working voltage was set to .