Pancreatic α amylase

After the fully-cooked time had elapsed, pasta samples (100 mg) were processed as previously described by Rocchetti et al. [9 (link)].
Briefly, spaghetti samples (100 mg) were cooked for the fully-cooked time (see paragraph 2.6) and then minced in 4 mL of maleic buffer (pH 6) containing an enzyme mixture composed of amyloglucosidase (AMG; 4 mL; 300 U/mL; Megazyme, Wicklow, Ireland) and pancreatic α-amylase (40 mg; 3000 U/mg; Megazyme, Wicklow, Ireland). Samples were incubated in a shaking water bath at 37 °C. At 0, 30, 60, 120, and 180 min, the reactions were stopped with 8 mL of ethanol. Samples were then centrifuged at 2500 g for 10 min.
The amount of glucose was quantified by the glucose oxidase plus peroxidase (GOPOD)-based method (RESISTANT STARCH; Megazyme, Wicklow, Ireland).

Jelly fig (Ficus awkeotsang Makino) and, pea (Pisum sativum L.) for PE extraction, were purchased from a local market in Taipei, Taiwan. Soy protein was from Gemfont (Taipei). Methylene blue and sodium chloride were from Riedel-de Haën (Seelze, Germany). Acetone, methanol, hydrochloride solution and fluorescein isothiocyanate (FITC)-casein were from Merck (Darmstadt, Germany). Trypsin solution was from ThermoFisher Scientific (Waltham, MA, USA). Pancreatic α-amylase was from Megazyme (Bray, Ireland). Methyl red was from Ferak (Berlin, Germany). Polyvinylpolypyrrolidone (PVPP), ninhydrin, pectin, lipase, tannase, tannic acid, Folin–Ciocalteu reagent, sodium carbonate, dinitrosalicylic acid, bovine serum albumin (BSA) were from Sigma Aldrich (St. Louis, MO, USA). The DIAION^® HP-20 macroporous resin was from Mitsubishi Chemical (Tokyo, Japan). All chemicals were of reagent grade.

The cooked-to-optimum spaghetti samples (100 mg) were dispersed in 4 mL of maleic buffer (pH 6), containing an enzyme mixture composed of amyloglucosidase (AMG; 4 μL; 300 U/mL; Megazyme, Wicklow, Ireland) and pancreatic α-amylase (40 mg; 3000 U/mg; Megazyme, Wicklow, Ireland). Samples were incubated in a shaking water bath at 37 °C. At selected time intervals (i.e., 0, 30, 60, 120, and 180 min) the reaction was stopped by adding absolute ethanol. Samples were then centrifuged at 2500× g for 10 min. The amount of glucose was quantified as previously detailed, after the correction for glucose present in the AMG solution. Values were plotted on a graph vs. time, and the area under the hydrolysis curve (AUHC; 0–180 min) was measured by using the trapezoid rule. A starch hydrolysis index (HI) value was calculated as the AUHC with the product as a percentage of the corresponding area with white wheat bread [16 (link)].