1. Isolation and Screening of Ethanol Tolerant Yeast, for the Production of Indigenous Liquor P. Moopanar CSIR Supervisor: Sani Gumede DUT Supervisor: L. Reddy

2. Slide 2 © CSIR 2006 www.csir.co.za Introduction Production of traditional beer and its economical impact Figure 1: Cashew FruitFigure 2: Palm tree

3. Slide 3 © CSIR 2006 www.csir.co.za

4. Slide 4 © CSIR 2006 www.csir.co.za The focus of this research was to isolate and screen yeasts for the improvement of current production processes of indigenous liquor. Aim

5. Slide 5 © CSIR 2006 www.csir.co.za Objectives To isolate ethanol tolerant yeasts from traditional juices To test ethanol tolerance of the isolated yeast strains To prepare and validate a cell bank of the isolated yeasts to minimise process variation To determine the growth profile of the yeast isolates. Establishing the efficacy of combinations of selected isolates in marula juice fermentation, to produce the highest concentration of ethanol

6. Slide 6 © CSIR 2006 www.csir.co.za Isolation and selection of high ethanol producing yeasts from Marula, Cashew, & Palm Isolates were evaluated for ethanol tolerance at a range of concentrations Methodology Shake flask experiments were conducted in triplicate to determine growth profiles Purified isolates were cell banked for future evaluations, the cell bank was subsequently validated Combinations of the isolates were screened for ethanol productivity in marula juice

7. Slide 7 © CSIR 2006 www.csir.co.za Fig 1: KM01Fig 2: KM02 Fig 3: KC01 Results & Discussion

8. Slide 8 © CSIR 2006 www.csir.co.za Cell Growth during ethanol tolerance trial (9%)

9. Slide 9 © CSIR 2006 www.csir.co.za Cell Growth during ethanol tolerance trial (12%)

10. Slide 10 © CSIR 2006 www.csir.co.za Cryopreservation Yeast isolateAverage counts before Average counts in cryovials after cryopreservation (cfu.ml -1 ) % cell recovery cryopreservation KC012.47 * 10 8 1.55 *10 8 91.50% KM017.205 *10 7 6.23 *10 7 97.50% KM021.113*10 8 1.63*10 8 95.50%

11. Slide 11 © CSIR 2006 www.csir.co.za Growth curve determination

12. Slide 12 © CSIR 2006 www.csir.co.za Ethanol productivity rates

13. Slide 13 © CSIR 2006 www.csir.co.za Sugar consumption rates

14. Slide 14 © CSIR 2006 www.csir.co.za pH profile of yeast combinations

15. Slide 15 © CSIR 2006 www.csir.co.za Conclusion Successful isolation of 3 ethanol tolerant yeasts, KM01, KM02 and KC01 Combination of three isolates was most productive in terms of ethanol productivity rates, sugar utilization rates and the final ethanol concentration obtained

16. Slide 16 © CSIR 2006 www.csir.co.za Proposed Future Work Pathogenicity evaluation Ethanol productivity of individual isolates Lyophilisation Impact of starter culture on taste, texture and end product Final selection and identification of organism/s Productivity of starter culture in large scale application.

17. Slide 17 © CSIR 2006 www.csir.co.za CSIR Biosciences and the Bioprocess development Team, especially Sani Gumede, Zinhle Ngubane and Nodumo Zulu for their intellectual input and all the individuals who aided in the success of the project. Acknowledgements