2.5. Amylose Content Determination (Iodine Assay)

Iodine colorimetric method was used to determine the amylose content. For the test, 0.010 g of the sample were dissolved in NaOH 0.1 M at 50 °C then neutralized with HCl 4 M and dilute up to 50 mL with ultrapure water. The iodine reagent solution was diluted in ultrapure water of a stock solution containing 7% w/w iodine and 5% w/w potassium iodide in a hydro-alcoholic mixture (90% v/v of ethanol). Then, 0.5 mL of the iodine reagent solution (0.2% w/w of iodine in water) were added to 10 mL of sample diluted solution. After mixing, the absorbance of solution was measured spectrophotometrically at 625 nm. The amylose content of the samples was determined by building up a calibration curve using pure amylose standard solutions (y = 0.1828 + 0.1320; R² = 0.994). Analyses were performed in triplicates.