2.3.6. Anthocyanin Content

The anthocyanin content (AC) was determined by using the procedure reported in literature, with some modifications [21]. 1.0 g of LOMW was suspended in 10.0 mL of methanolic solution of HCl (1.0% *v*/*v*). The solution was incubated at 60 ◦C under stirring for 1 h and then filtered in a volumetric flask (10.0 mL). The absorbance was spectrophotometrically measured at 657 and 530 nm. The net absorbance was calculated according to the following equation:

$$\text{Net absorbedance} = \text{Absorbance at 530 nm} - 0.25 \text{ (Absorbance at 657 nm)} \tag{1}$$

AC was calculated on the basis of cyanidin-3-glycoside (molecular weight equal to 449.1 g mol−<sup>1</sup> and extinction coefficient of 29,600) [22]. AC was expressed in milligrams of anthocyanins per gram of LOMW (mg AC/g LOMW), according to the following equation:

$$\text{AC (mg/g)} = \text{((Net absorber)/29,600)} \times \text{MW} \times \text{FT} \times \text{(V/(Sample weight))} \tag{2}$$

where MW is the molecular weight of cyanidin-3-glycoside, FT is the dilution factor, V is the total volume (mL). Each measurement was performed in triplicate and data expressed as means (±SD).
