Improving biological control of industrial bioethanol production processes by using Saccharomyces cerevisiae's own weapons

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Effect of saccharomycin, a natural Saccharomyces cerevisiae biocide, on Hanseniaspora guilliermondii cells surface

Publication . Calvário, Joana; Silva, Nelly; Almeida, M. Gabriela; Albergaria, Helena; Eaton, Peter; Macedo, Anjos L.; Caldeira, Jorge

ABSTRACT: During spontaneous wine fermentations, most of the non-Saccharomyces yeasts present in grape musts show an early decline in their population. It was traditionally assumed that Saccharomyces cerevisiae (S.c.) prevalence was due to the higher resistance of this species to ethanol. However, wine fermentations performed with single cultures of non-Saccharomyces strains showed that those strains could withstand much higher ethanol levels [1]. It was then found that S.c. (strain CCMI 885) produced antimicrobial peptides (AMPs) that are responsible for the early death of the non Saccharomyces yeasts [2]. In previous work, we isolated, purified and sequenced those ntimicrobial peptides (AMPs) and found that they derive from the glyceraldehyde 3-phosphate dehydrogenase enzyme [3]. These GAPDH-derived AMPs compose the natural biocide secreted by S.c., which we named saccharomycin, and are effective against sensitive yeasts both in its natural/isolated and synthetic form.

2021-04Journal article

Open access

Evaluation of the biocontrol potential of a commercial yeast starter against fuel-ethanol fermentation contaminants

Publication . Branco, Patricia; Diniz, Mário; Albergaria, Helena

ABSTRACT: Lactic acid bacteria (LAB) and Brettanomyces bruxellensis are the main contaminants of bioethanol fermentations. Those contaminations affect Saccharomyces cerevisiae performance and reduce ethanol yields and productivity, leading to important economic losses. Currently, chemical treatments such as acid washing and/or antibiotics are used to control those contaminants. However, these control measures carry environmental risks, and more environmentally friendly methods are required. Several S. cerevisiae wine strains were found to secrete antimicrobial peptides (AMPs) during alcoholic fermentation that are active against LAB and B. bruxellensis strains. Thus, in the present study, we investigated if the fuel-ethanol commercial starter S. cerevisiae Ethanol Red (ER) also secretes those AMPs and evaluated its biocontrol potential by performing alcoholic fermentations with mixed-cultures of ER and B. bruxellensis strains and growth assays of LAB in ER pre-fermented supernatants. Results showed that all B. bruxellensis strains were significantly inhibited by the presence of ER, although LAB strains were less sensitive to ER fermentation metabolites. Peptides secreted by ER during alcoholic fermentation were purified by gel-filtration chromatography, and a bioactive fraction was analyzed by ELISA and mass spectrometry. Results confirmed that ER secretes the AMPs previously identified. That bioactive fraction was used to determine minimal inhibitory concentrations (MICs) against several LAB and B. bruxellensis strains. MICs of 1-2 mg/mL were found for B. bruxellensis strains and above 2 mg/mL for LAB. Our study demonstrates that the AMPs secreted by ER can be used as a natural preservative in fuel-ethanol fermentations.

2022-05Journal article

Open access

Wine Spoilage Control: Impact of Saccharomycin on Brettanomyces bruxellensis and Its Conjugated Effect with Sulfur Dioxide

Publication . Branco, Patricia; Coutinho, Rute; Malfeito-Ferreira, Manuel; Prista, Catarina; Albergaria, Helena

ABSTRACT: The yeast Brettanomyces bruxellensis is one of the most dangerous wine contaminants due to the production of phenolic off-flavors such as 4-ethylphenol. This microbial hazard is regularly tackled by addition of sulfur dioxide (SO2). Nevertheless, B. bruxellensis is frequently found at low levels (ca 10(3) cells/mL) in finished wines. Besides, consumers health concerns regarding the use of sulfur dioxide encouraged the search for alternative biocontrol measures. Recently, we found that Saccharomyces cerevisiae secretes a natural biocide (saccharomycin) that inhibits the growth of different B. bruxellensis strains during alcoholic fermentation. Here we investigated the ability of S. cerevisiae CCMI 885 to prevent B. bruxellensis ISA 2211 growth and 4-ethylphenol production in synthetic and true grape must fermentations. Results showed that B. bruxellensis growth and 4-ethylphenol production was significantly inhibited in both media, although the effect was more pronounced in synthetic grape must. The natural biocide was added to a simulated wine inoculated with 5 x 10(2) cells/mL of B. bruxellensis, which led to loss of culturability and viability (100% dead cells at day-12). The conjugated effect of saccharomycin with SO2 was evaluated in simulated wines at 10, 12, 13 and 14% (v/v) ethanol. Results showed that B. bruxellensis proliferation in wines at 13 and 14% (v/v) ethanol was completely prevented by addition of 1.0 mg/mL of saccharomycin with 25 mg/L of SO2, thus allowing to significantly reduce the SO2 levels commonly used in wines (150-200 mg/L).

2021-12Journal article

Open access

Evaluation of Biotechnological Active Peptides Secreted by Saccharomyces cerevisiae with Potential Skin Benefits

Publication . Mauricio, Elisabete; Branco, Patricia; Araújo, Ana Luiza Barros; Roma-Rodrigues, Catarina; Lima, Katelene; Duarte, Maria Paula; Fernandes, Alexandra; Albergaria, Helena

ABSTRACT:Biotechnological active peptides are gaining interest in the cosmetics industry due to their antimicrobial, anti-inflammatory, antioxidant, and anti-collagenase (ACE) effects, as well as wound healing properties, making them suitable for cosmetic formulations. The antimicrobial activity of peptides (2–10 kDa) secreted by Saccharomyces cerevisiae Ethanol-Red was evaluated against dermal pathogens using broth microdilution and challenge tests. ACE was assessed using a collagenase activity colorimetric assay, antioxidant activity via spectrophotometric monitoring of nitrotetrazolium blue chloride (NBT) reduction, and anti-inflammatory effects by quantifying TNF-α mRNA in lipopolysaccharides (LPS)-exposed dermal fibroblasts. Wound healing assays involved human fibroblasts, endothelial cells, and dermal keratinocytes. The peptides (2–10 kDa) exhibited antimicrobial activity against 10 dermal pathogens, with the Minimum Inhibitory Concentrations (MICs) ranging from 125 µg/mL for Staphylococcus aureus to 1000 µg/mL for Candida albicans and Streptococcus pyogenes. In the challenge test, peptides at their MICs reduced microbial counts significantly, fulfilling ISO 11930:2019 standards, except against Aspergillus brasiliensis. The peptides combined with MicrocareⓇ SB showed synergy, particularly against C. albicans and A. brasilensis. In vitro, the peptides inhibited collagenase activity by 41.8% and 94.5% at 250 and 1000 µg/mL, respectively, and demonstrated antioxidant capacity. Pre-incubation with peptides decreased TNF-α expression in fibroblasts, indicating anti-inflammatory effects. The peptides do not show to promote or inhibit the angiogenesis of endothelial cells, but are able to attenuate fibrosis, scar formation, and chronic inflammation during the final phases of the wound healing process. The peptides showed antimicrobial, antioxidant, ACE, and anti-inflammatory properties, highlighting their potential as multifunctional bioactive ingredients in skincare, warranting further optimization and exploration in cosmetic applications.

2024-09Journal article

Open access