*3.2. Acetylation of PPC*

Acetylated PPC samples were prepared using the method of Miedzianka et al. [24] with slight modifications. The pumpkin protein preparation was acetylated with acetic anhydride by adding different concentrations of modifying reagent (0.4, 1.0 and 2.0 mL of anhydride per 1 g of protein contained in the preparation) to the 1% aqueous suspension. The reaction took place over 30–90 min, and the pH was established at pH 7.5–8 by dropwise addition of 1 M NaOH. The solution was constantly monitored for changes in pH and maintained at the assumed pH with constant stirring. After this time, the precipitate was separated from the supernatant by centrifugation using a Biofuge 28 RS centrifuge (5260 rpm/min for 15 min, Heraeus Sepatech, Osterode, Germany). Excess modifying reagent was removed from the resulting precipitate, i.e., the precipitate was mixed with water until the liquid's conductance was close to that of distilled water (3–5 times). Then, it was bathed with distilled water until the conductance of liquid was similar to that of distilled water. Then, the protein was freeze-dried at a pressure of 63 Pa, with the heating shelves at 50 ◦C, for 24 h using a Christ Alpha 1-4 LSCplus (Osterode am Hatz, Germany), sieved through a sieve with a pore size of 420 µm and stored in a sealed plastic container at about −20 ◦C until further analysis. Untreated native PPC was used as the control. Acetylation was performed in two technological repetitions.
