2.1.3. Spectrophotometric Analysis

After the extraction, purified pigments were dissolved in 500 μL of NaOH 1 M and its UV-Visible spectrum was measured in a UV spectrophotometer (UV 1600 PC Spectrophotometer, VWR International) by using M.Wave professional 2.0. A standard graph for estimation was used and was made using synthetic melanin. NaOH 1 M was used as a blank and the instrument was set in a range of 200–800 nm for the analysis. The correlation between absorbance and wavelength was defined. To determine the concentration of extracted melanin, synthetic DHN (1,8-DiHydroxyNaphthalene) melanin (Thermo-Fisher Scientific, Waltham, MA, USA) was prepared in 1 M NaOH at concentrations of 500 mg/mL and as reported in [27], a standard curve at 650 nm was obtained.
