Seleção de leveduras selvagens multirresistentes aos fatores de estresse do processo fermentativo industrial para produção de etanol em indústrias alcooleiras

Abstract

Contaminant yeasts in an industrial fermentation process may become predominant and lead to losses such as decreases in efficiency and fermentative yield. The selected strain of the plant itself prevails throughout the process, preventing undesirable yeasts from settling in the process. This study aimed to use the yeast diversity from the industrial fermentation process to obtain strains suitable for the selection of multiresistant isolates. The yeast colonies for selection were obtained from the fermented must and a dorn from the plant at the end of the 2014/2015 harvest. One sample was at the start of the fermentation (2 hours) and another at the end of the fermentation (after 8 hours). The macro morphological diversity was verified by surface plating in WLN medium. Mixed cultures were introduced in a fermentation cycle of selection for resistance to acidity, high Brix and high ethanol content. Crude cane molasses was used as a substrate. The fermentation started with 17ºBrix, each cycle lasted 24 hours, 4 cycles in the same Brix until reaching 22ºBrix. After each 24 hours a cell wash was performed with 2 mL dilute sulfuric acid (sterile) pH 2.5, left standing for 2 hours, after 2 hours a new fermentation began. With the conclusion of the selective fermentation totaling 24 days, the surface plating was performed in WLN medium to observe the morphological diversity after the selective pressure. Genetic variability of yeasts before and after 24 cycles. They were also evaluated by the MSP-PCR fingerprinting technique using the oligonucleotide (GTG)5. A kinetic evaluation was performed determining cell-specific growth measured by absorbance. Resistance tests were performed on ethanol, glucose and sucrose, pH and temperature. The SC-32 strain of the selection pressure, obtained greater combined resistance to all stress factors and was compared to CAT-1, in a fermentation (24 hours) with constant agitation, concentrations of 17 and 22ºBRIX, temperatures of 30 And 40ºC, analyzing cell viability sugar consumption (BRIX), mass produced (gravimetry) and pH, obtaining satisfactory results. The results obtained in this work demonstrated that the selection pressure did not vary the morphological diversity and varied the genetic diversity of the isolates and that the selection process did not provide selection pressure for yeasts with higher growth rates, being the isolates with better diversity performance. However, several other parameters, such as conversion to ethanol, fermentative efficiency, presence of acids, glycerol produced and maintenance costs of this and other strains should be evaluated for the proper choice of a yeast for the production of ethanol.