Selection of yeast strains isolated during Alentejo wine productions to be used as starters

The knowledge of new species of yeasts potentialities with fermentative characteristics and/or with specific sensorial properties can be an important tool for the development of oenological processes with higher microbial complexity in a controlled system, conferring a higher wine quality and promoting innovative organoleptic characteristics to the final product. Identification and characterization of yeasts with certain oenological properties is very important to explore their fermentation potential and improve wine production. The aim of this study was the identification and characterization of yeast strains with oenological potential, isolated from different grape musts fermentations from Alentejo region. Initial screening allowed a selection of 16 different yeast strains from grape must from Touriga Nacional, Cabernet Sauvignon, Antão Vaz and Síria, at various stages of the fermentation process, to be used in immobilization systems, on the development of new oenological solutions. Batch cultures of these microorganisms were performed in order to characterize kinetic growth, carbohydrate consumption and ethanol production. Selected strains were characterized by their macroscopic and microscopic characteristics. DNA were extracted and amplified for strains identification and characterization [1]. The identification was done by the sequencing of PCR products obtained after amplification of the D1/D2 domain of the large subunit ribosomal RNA gene, and it was possible to identify Saccharomyces cerevisiae, Torulaspora delbrueckii, zygoascus meyerae, Pichia kudriavzevii, Rhodotorula mucilaginosa, Saccharomyces sp. and Hanseniaspora sp.. strains. Molecular analysis by M13-PCR allowed to characterize the genetic profiles for all the isolated yeast strains, showing differences on molecular profiles that allowed grouping according to phylogenetic proximity, both between different species and between strains of the same species. This approach allows better microbiologic knowledge about the grape must flora and to improve the selection of the most appropriate consortia to be used in the creation of new starters for immobilization solutions

New solutions of immobilized yeasts with oenological potential

The wine sector is expanding worldwide, with more and more wine producers entering this sector, which leads to an increase in wine production. Thus, it is important that companies have access to new technologies, optimizing their production. The development of new methodologies, such as immobilized yeasts has great oenological potential, since they have several oenological applications and allow to improve the quality / price relation of the final product. This type of methodology nowadays begins to be used by the producers, especially in the production of sparkling wines, during the second alcoholic fermentation. In addition, they can also be used in alcoholic and malolactic fermentations, in the production of late harvest wines or in the correction of some problems, such as excessive volatile acidity or late fermentation. The aim of this work was the production of new solutions of immobilized yeasts with oenological potential. A set of yeasts strains isolated during different stages of grape musts fermentation were immobilized in two different inorganic porous supports: volcanic tuff and expanded clay. The immobilization success was accessed by scanning electron microscopy and to validate the potential of the immobilization, the immobilized solutions were tested in microvinification assays during 7 days. After this period the stability of the matrixes and the presence of yeast cells on the immobilized system were confirmed by SEM. The results proved that both supports with immobilized yeasts allowed the conversion of must sugars into ethanol, and can be reused during five batches with high efficiency, displaying the potential use of these matrices in oenological context

Redução de compostos fenólicos de resíduos de lagares de azeite utilizando culturas de Coriolus versicolor

In this study, C. versicolor cultures were performed during 20 days with different RLA (olive mill wastes)subtracts, 50%, 75% and 100%. During this assay, samples were collected in regular time intervals, cell growth were evaluated and broth decolorization was observed. The organic matter and total phenolic compounds level in the culture broth showed a decrease in all RLA ratios. 50% and 75% RLA presented high total phenolic removal, 85% and 90% respectively. In the end of the assay, toxicological evaluation of the culture broth was performed against Artemia salina. These results showed a toxicological decrease of these residues after biological treatment with C. Versicolor

Toxicological and pharmacological properties of essential oils of Calamintha nepeta, Origanum virens and Thymus mastichina of Alentejo (Portugal)

Three autochthonous flavouring herbs from Alentejo (Portugal), Calamintha nepeta (syn. Clinopodium nepeta), Origanun virens and Thymus mastichina, were selected to evaluate toxicological, antioxidant, antiproliferative and antimicrobial potential of their essential oils (EOs). C. nepeta and T. mastichina EOs showed a high content of oxygenated monoterpenes (86–91%) while O. virens had similar content of oxygenated and hydrocarbon monoterpenes (45%). Toxicological assessment suggests high activity against A. salina (31.8 < CL50 < 128.4 mg/L) and very low toxicity in Swiss mice (DL50≥1500 mg/kg). EOs showed high antioxidant ability by DPPH radical scavenging assay (0.1–0.6 mg QE/mL EO), total reducing power method (0.2–1.7 mg QE/mL EO) and β-carotene/ linoleic acid system (11–501 mg QE/mL EO). An important antiproliferative effect against human breast tumour cell line was observed (88.9 < EC50 < 108.5 mg/L). Moreover, EOs presented a large antibacterial spectrum. Results point out the low toxicity and high antioxidant, antiproliferative and antimicrobial activities of EOs of these endemic aromatic plants, suggesting their potential use in biotechnological, food and/or pharmaceutical industries

Essential oils of Calamintha nepeta, Origanum vulgare and Thymus mastichina of Alentejo (Portugal): a pharmacological approach

Alentejo, in the south of Portugal, is rich in endemic aromatic plants, that are used as condiments and food additives by the local population. The aim of this study was to evaluate the antioxidant, antimicrobial and antiproliferative activities of EOs of autochthones Calamintha nepeta (L.) Savi (syn. Clinopodium nepeta (L.) Kuntze), Origanum vulgare L., and Thymus mastichina L. EOs were extracted from the aerial part of the plants by hydrodistillation and the chemical composition was analyzed by GC-FID and GC-MS. Antioxidant potential of the oils was evaluated by three different assays: DPPH radical, β-carotene/linoleic acid and reducing power methods. Antimicrobial activity of the oils was evaluated by a solid disk diffusion assay and minimal inhibitory concentrations (MIC) were determined by a microdilution broth method. Toxicity of the EOs was screened by the brine shrimp lethality test (LC50) and the oral lethal doses (DL50) were determined for mice. Cell viability was assessed by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay using MDAMB231 breast cancer cells

In silico, NMR and pharmacological evaluation of an hydroxyoxindole cholinesterase inhibitor

From a screening study of various potential inhibitors for cholinesterases (ChEs), compound (rac)-1 (4-((3-hydroxy-2-oxo-3-phenylindolin-1-yl) methyl) piperidin-1-ium chloride) showed an IC50 of 18 μM for butyrylcholinesterase (BuChE). Herein we present a toxicological and pharmacological evaluation of (rac)-1 to determine its potential for use as an alternative ChE inhibitor for the treatment of Alzheimer’s disease. The strategy adopted included in vivo and ex vivo studies with mouse models, Molecular Modelling and Saturation Transfer Difference (STD) NMR studies. Preliminary molecular docking studies were conducted with both (R) and (S)-1 with acetylcholinesterase (AChE) and BuChE, prior to advancing to the mouse model, and indeed favorable interactions were observed, with (R)-1 showing the best binding with AChE and (S)-1 with BuChE. STD-NMR studies were used to successfully validate these results. Toxicological studies were also conducted using the Artemia salina model, with donepezil as reference. It was found that in the in vivo mouse studies that (rac)-1 presented a slightly better inhibition of AChE (0.096 µmol.min−1.mg−1) than donepezil (0.112 µmol.min−1.mg−1) and the same level of inhibition for BuChE as donepezil (0.014 µmol.min−1.mg−1)