**1. Introduction**

Plant-based proteins are innovative ingredients with fast-growing applications in the food industry. They are sources of important bioactive compounds and ingredients that are used in the production of functional foods [1]. Sustainability, ethical implications, population growth, variety, and the formulation of healthier products are among their main advantages over animal proteins. However, on the other hand, most plant-based protein preparations obtained by aqueous extraction methods are characterized by poor aqueous solubility, weak functionality (i.e., gelling, emulsification, and foaming), and a high degree of complexity, as well as susceptibility to pH, ionic strength, and temperature, which limits their applications in the industrial sector [2]. Moreover, they are less digestible and have less ability to transport other important nutrients, such as calcium and iron. Therefore, efficient modification processes are needed to improve the value of plant-based proteins.

One of the methods of protein extraction from plant-based materials that retain their original functional properties is the dry tribo-electrostatic separation method. This technology relies on the efficiency of milling to mechanically dissociate the proteins, which are subsequently separated by an air stream based on particle size and density [3,4]. As a result, the dry fractionation method has many advantages, including having no chemical residues, a minimal impact on the techno-functional properties, and the loss of insoluble protein, as

**Citation:** Miedzianka, J.; Walkowiak, K.; Zieli ´nska-Dawidziak, M.; Zambrowicz, A.; Wolny, S.; Kita, A. The Functional and Physicochemical Properties of Rice Protein Concentrate Subjected to Acetylation. *Molecules* **2023**, *28*, 770. https:// doi.org/10.3390/molecules28020770

Academic Editor: Susy Piovesana

Received: 13 November 2022 Revised: 27 December 2022 Accepted: 9 January 2023 Published: 12 January 2023

**Copyright:** © 2023 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

well as having low energy consumption. The low protein extraction yield is the main disadvantage [3]. Based on the studies of Tabtabaei et al. [5], dry tribo-electrostatic separation can produce protein-rich plant fractions from navy bean flour which were characterized by superior solubility, emulsifying, and foaming when compared to wet process isolates. Vitelli et al. [6] investigated the performance of electrostatic separation to produce proteinand carbohydrate-rich fractions from hammer-milled navy bean flour. The results revealed that the samples collected from the middle and top part of the plate had a significantly higher protein content and, therefore, a lower starch content than the samples collected from the bottom of the plate.

Some of the functional properties can also be changed, inter alia, by chemical modifications [7]. Chemically modified proteins are produced by the addition of new functional moieties or the elimination of the components from the protein's structure. In proteins, the most reactive nucleophilic groups are the *E*-amino groups of lysyl residues and the SH group of cysteine. Therefore, these amino acids undergo the most modification. Examples of chemical modification include acetylation and succinylation based on the use of acetic or succinic anhydride, respectively, as the acylating agent. Acylation is a type of modification that has been widely used due to its efficiency, low cost, and ease of operation. Hydrophilic functional groups, such as hydroxyls, ε-amino groups, and phenols, are susceptible to acetylation [8]. Acetylation is advantageous from a technological point of view because the protein concentrates and isolates obtained can have improved solubility, emulsifying ability, and foaming and water-binding capacity and also stabilize the emulsions and determine their color. Additionally, it has been scientifically proven that modification through acetylation may cause changes in the chemical composition of the obtained preparations without adversely affecting their nutritional value [8–10]. However, the chemical modification methods (including acetylation with acetic anhydride) are not favorable for food applications as they use chemicals and produce chemical byproducts. Additionally, the modified proteins can be less digestible and are not utilized in animal feeding tests.

One of the plant-based alternatives to meat or soy protein is rice (*Oryza sativa* L.). It is a staple cereal and is widely consumed around the world. It has high nutritional value, being a source of starch, mineral salts, vitamins, and dietary fiber. The protein content of rice is relatively small (about 3%). However, rice proteins have been recognized as highly nutritious, hypoallergenic, and particularly healthful for human consumption. Additionally, the antioxidant and nutraceutical properties of rice are associated with a reduction in the risk of oxidative stress, which contributes to the prevention of hypercholesterolaemia [11,12].

Commercial rice protein concentrate is first subjected to alkaline extraction, and then the proteins are precipitated by adjusting the pH to their isoelectric point, where the nonprotein components are isolated by enzymatic processes [12,13]. Depending on factors such as the rice cultivar and the degree of milling of the rice, the protein content of the products of these treatments ranges from 65% to 90%. Rice protein concentrate (RPC) produced on an industrial scale is popular in the food and pharmaceutical industries, mainly in protein supplements and also in drink shakes, bars, or gels. Additionally, studies indicate that RPC can be used as a value-added ingredient in the production of bread [14] or biscuits [15]. It is considered as a substitute for soy protein as it has no beany flavor [16]. However, its powder presents fine particles and poor dissolution properties, limiting its use.

So far, rice protein has been subjected to chemical modification by phosphorylation with sodium trimetaphosphate (STMP) [17,18] and alkaline deamidation [19]. As for acetylation using acetic acid anhydride, authors have analyzed the physicochemical and functional properties of rice bran protein concentrate [17] and rice protein successively subjected to trypsin hydrolysis [20]. The novelty of our research is the analysis of commercial RPC, and it is known that among the factors which have a great influence on the properties of modified plant protein preparations are the protein's origin and method of isolation, as well as the conditions of the modification process [21]. The poor functional properties of RPC limit its use, but due to the high biological value of these proteins, it

is justified to search for modification methods that might lead to the improvement of its functional properties. Therefore, the aim of this work was to try to improve the functional properties of commercial RPC by acetylation using acetic anhydride and to analyze the influence of the modification on the physicochemical properties (chemical composition, structure, protein profile, and digestibility).
