STRESS TOLERANCE YEAST STRAIN FROM PAPAYA WASTES FOR BIOETHANOL PRODUCTION

Papaya wastes is naturally fermented and potentially overgrown by microorganisms such yeasts. The research aims to determine the presence of ethanol-fermenting yeasts on papaya wastes which has the ability to tolerate high glucose and ethanol contents. Yeasts isolated from papaya wastes with using Potato Dextrose Agar/PDA which modified with 3% yeast extract/YE and 10 ppm Amoxicillin, then incubated for 48h at room temperature. The isolates were identified macroscopic and microscopically then the yeast-like isolates cultured on Nutrient Broth/NB with the addition of 3% YE, 10 ppm Amoxicillin and 30% glucose or alcohol to be tested for the tolerance ability towards high glucose and alcohol. Yeasts presence on high glucose and alcohol media was determine by UV-Vis spectrophotometer by measuring optical density (OD) for UV absorbance at lambda=600nm. The isolate with highest OD at glucose and alcohol media grown at papaya wastes for 72h and the ethanol contents measured by chromium dichromate oxidation methods every 24h. Species identification performed using sequence analysis of the rRNA gene internal transcribed spacer (ITS) region with using primers ITS1 (5'-TCCGTAGGTGAACCTGCGG-3') and ITS4 (5'-TCCTCCGCTTATTGATATGC-3'), the sequences compared with the GenBank database using Basic Local Alignment Search Tools/BLAST algorithm. The results showed that three yeast-like isolates found from papaya wastes, with Y1 isolate identified as best isolate with the OD of 0.4699 at 30% glucose media and OD of 0.0960 at 30% alcohol media, with the highest ethanol fermented at 48h was 4.34%. The rate of isolate identification by sequence analysis resulted 96.20% (531/552) identical with Pichia sp. strain AQGWD 7.