Citation: Prokopová, A.; Gál, R.; Mokrejš, P.; Pavlaˇ cková, J. Preparation of Gelatin from Broiler Chicken Stomach Collagen. Foods 2023, 12, 127. https://doi.org/ 10.3390/foods12010127 Academic Editor: Jose Lorenzo Rodriguez Received: 23 November 2022 Revised: 17 December 2022 Accepted: 22 December 2022 Published: 27 December 2022 Copyright: © 2022 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/). foods Article Preparation of Gelatin from Broiler Chicken Stomach Collagen Aneta Prokopová 1 , Robert Gál 2 , Pavel Mokrejš 1, * and Jana Pavlaˇ cková 3 1 Department of Polymer Engineering, Faculty of Technology, Tomas Bata University in Zlín, Vavreˇ ckova 275, 760 01 Zlín, Czech Republic 2 Department of Food Technology, Faculty of Technology, Tomas Bata University in Zlín, Vavreˇ ckova 275, 760 01 Zlín, Czech Republic 3 Department of Lipids, Detergents and Cosmetics Technology, Faculty of Technology, Tomas Bata University in Zlín, Vavreˇ ckova 275, 760 01 Zlín, Czech Republic * Correspondence: mokrejs@utb.cz; Tel.: +420-576-031-230 Abstract: With the increasing consumption of poultry meat around the world, the use of chicken stomachs as a source of collagen is being offered. The objective of this study was to extract gelatin from the stomachs of broiler chickens and to estimate their gel strength, ash content, viscosity, gelling point, melting point, clarity and digestibility. An innovative biotechnological method based on the conditioning of collagen with a microbial endoproteinase (Protamex ® ) and hot-water extraction was used to control the chemical and thermal denaturation process of collagen to prepare gelatin. The experiments were planned using a Taguchi design, 2 factors at 3 levels; factor A for the amount of proteolytic enzyme (0.10, 0.15 and 0.20%) and factor B for the extraction temperature (55.0, 62.5 and 70.0 ◦ C). Data were statistically processed and analyzed at a significance level of 95%. The gelatin yield averaged 65 ± 8%; the gel strength ranged from 25 ± 1 to 439 ± 6 Bloom, the viscosity from 1.0 ± 0.4 to 3.40 ± 0.03 mPa·s, gelling point from 14.0 ± 2.0 to 22.0 ± 2.0 ◦ C, melting point from 28.0 ± 1.0 to 37.0 ± 1.0 ◦ C. The digestibility of gelatin was 100.0% in all samples; the ash content was very low (0.44 ± 0.02–0.81 ± 0.02%). The optimal conditions for the enzymatic treatment of collagen from chicken stomachs were achieved at a higher temperature (70.0 ◦ C) and a lower amount of enzyme (0.10–0.15%). Conditioning chicken collagen with a microbial endoproteinase is an economically and environmentally friendly processing method, an alternative to the usual acid- or alkaline-based treatment that is used industrially. The extracted products can be used for food and pharmaceutical applications. Keywords: biotechnology; chicken stomachs; collagen; enzyme conditioning; food; gelatin; meat by-products; pharmacy; proteins 1. Introduction Collagen is a protein biomolecule made of amino acids. Collagen is the most abundant structural protein in the extracellular matrix of various connective tissues in the body, responsible for strength and flexibility [1,2]. Hydrolysis is used for the biochemical break- down of collagen bonds. In an acidic environment, it is assumed that an electrophilic mechanism of hydrolysis occurs [3] in comparison to the nucleophilic mechanism that is common in an alkaline environment [4]. The extraction of gelatin based on the partial acid-controlled hydrolysis of the collagen structure is called type A gelatin, and the second one based on partial alkaline-controlled hydrolysis of the collagen structure is called type B gelatin. Both chemical methods are slow and costly and have an environmental footprint [5]. Compared to chemical agents, enzymes are more environmentally friendly, minimize pro- duction costs, and allow the desired functional properties of gelatins to be achieved [6]. The most used enzymes today include industrially produced microbial enzymes, animal enzymes trypsin and pepsin, as well as plant enzymes (e.g., papain) [7]. Enzymes are pure proteins, are fully biodegradable, and do not produce unwanted by-products [8]. Foods 2023, 12, 127. https://doi.org/10.3390/foods12010127 https://www.mdpi.com/journal/foods