Citation: Postigo, V.; O’Sullivan, T.; Elink Schuurman, T.; Arroyo, T. Non-Conventional Yeast: Behavior under Pure Culture, Sequential and Aeration Conditions in Beer Fermentation. Foods 2022, 11, 3717. https://doi.org/10.3390/ foods11223717 Academic Editor: Francesco Grieco Received: 10 October 2022 Accepted: 16 November 2022 Published: 18 November 2022 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affil- iations. 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 Non-Conventional Yeast: Behavior under Pure Culture, Sequential and Aeration Conditions in Beer Fermentation Vanesa Postigo 1,2 , Tadhg O’Sullivan 3 , Tom Elink Schuurman 3 and Teresa Arroyo 1, * 1 Department of Agri-Food, Madrid Institute for Rural, Agriculture and Food Research and Development (IMIDRA), El Encín, A-2, km 38.2, 28805 Alcalá de Henares, Spain 2 Brewery La Cibeles, Petróleo 34, 28918 Leganés, Spain 3 Heineken Supply Chain B.V., Burgemeester Smeetsweg 1, 2382 PH Zoeterwoude, The Netherlands * Correspondence: teresa.arroyo@madrid.org Abstract: The use of wild yeasts, isolated from different environments, is becoming the most interest- ing option for the production of new beers. The objective of this study is to evaluate the potential of seven non-conventional yeast strains from five different species (Saccharomyces cerevisiae, Hanseni- aspora guilliermondii, Metschnikowia pulcherrima, Torulaspora delbrueckii, and Zygosaccharomyces bailii) isolated from Madrid agriculture to produce type ale beer. Wild yeast strains were evaluated at labo- ratory and pilot plant scales under different fermentation conditions (pure, aerated, and sequential culture). Strain S. cerevisiae SafAle S-04 was used as a reference. Throughout the fermentation of beer, volatile compounds were determined by GC and residual sugars by HPLC, among other parameters. The yeast strains used for the fermentation in pure culture conditions were unable to ferment maltose and maltotriose (0.73–1.18% v/v of ethanol). The results of the study under aerated conditions showed varying levels of higher alcohol and ester concentrations. It should be noted that the strain CLI 1057 (S. cerevisiae) fermented maltose in the presence of oxygen (Kluyver effect). This strain also showed a high production of 4-vinyl guaiacol, making it suitable for producing beers with a phenolic profile. Finally, three strains (H. guilliermondii, Z. bailii, and T. delbrueckii) were evaluated in sequential culture together with commercial strain and found to improve the organoleptic characteristics of the brewed beer. These approaches offer the opportunity to add new product characteristics to the beers. Keywords: non-conventional yeasts; non-Saccharomyces; ale beer; aeration; aromas 1. Introduction Beer drinking habits have changed over the last few years. The volume of beer produced grew enormously until 2013, when it started to decline, with the production in 2021 amounting to about 1.86 billion hL [1]. In this context, the craft beer industry has taken an important role in the market. Among other proposals, a commitment to the production of non-alcoholic and low-alcoholic beers is emerging [2]. Therefore, there is general importance in developing new products that meet the needs of consumers both in terms of flavors and healthier products. In this sense, interest has focused on non- conventional yeasts and the search for species and strains with low ethanol production due to their low fermentative capacity and new aromatic profiles [2–4]. Low ethanol production is often determined by the inability of some yeasts to ferment maltose, the major sugar in beer wort. Steensels and Verstrepen conducted a study with wild Saccharomyces yeast strains, where only 12% could ferment 50% of the available sugars in the wort [5]. On the other hand, the work by Methner et al., with 110 non-Saccharomyces yeasts showed that only 30% of the yeasts studied were able to ferment 25% of the maltose and maltotriose content of the wort [6]. It should be noted that not only is species screening important, but so is strain screening, as the ability to ferment the sugars in wort is often strain-dependent. Foods 2022, 11, 3717. https://doi.org/10.3390/foods11223717 https://www.mdpi.com/journal/foods