Drews’ Brews and the

Bioreactor Blues
Section AA
Stephanie Ford, Evan Epstein, Emily Thomas,
Kyle Elliott, Andrew Kullman

January 28, 2020

Summary
1. Introduction to the unit
2. Background and objectives
3. Experimental design
4. Data and analysis
5. Conclusions
6. Next steps and recommendations

Established recommendations for maximizing aerobic growth

Introduction to the Unit
● Bioreactor → production of ethanol, product of plant

● Experimental Goals:
○ Protocol for yeast production
○ Impact of media components on growth
○ Establish measurement protocols

Bioreactor success = Plant success

Bioreactor background
● Controlled and uncontrolled
parameters
● Aerobic and anaerobic phase
● More efficacious than small batch
● Increased growth → Increased
ethanol

Bioreactor allows control over growth and fermentation.

Yeast Background
● Growth factors:
○ Temperature
○ Sugars
○ pH
○ Dissolved oxygen
(DO)
○ Proteins and other
nutrient sources

Hypothesis: growth will be optimized with higher nutrient concentrations

Pfeiffer, T., & Morley, A. (2014). An evolutionary perspective on the Crabtree effect. Frontiers in Molecular Biosciences, 1. https://doi.org/10.3389/fmolb.2014.00017
Experimental Design
● Small-scale experiments in shakers
● Standard media1:
○ 2% peptone, 1% yeast extract, 2% glucose by
mass
● Constant temperature (35 ℃)
● Varied input values of:
○ Activation method
○ Glucose concentration
○ Initial pH (AC)
○ Yeast extract concentration (AB)
● Took samples at:
○ 0, 15 min, 30 min, 1 hr, 1.5 hr, 24 hr, 72 hr

1
Carothers, J.“Cell Growth Kinetics”
Glucose Variation

Optimal initial glucose concentration >5%

pH Variation - AC

Optimal pH <5
Yeast Extract Variation - AB

Optimal initial concentration between 2% and 8%

Data Conclusions
● Optimal sugar concentration: > 5%
● Optimal initial pH: < 5
● Optimal yeast extract concentration: < 8% and > 2%
● Notes:
○ May not be economically optimal
○ Not in triplicate
○ Measurement error

Higher nutrient content and a lower pH improve growth to a point

Next Steps
● Further optimize operational parameters
● Find effect of dry pitching at various % glucose
● Develop parameters and setpoints for bioreactor
● Strengthen conclusions and assumptions with HPLC data
○ OD as a valid measurement of growth

○ Proportionality of ethanol production to biomass growth

Issues and Recommendations
● Work with other lab sections to ensure time and materials for triplicate trials
● Corroborate OD results with HPLC
● Equipment limitations
○ Carefully plan use of shaker table and bioreactor

● Safety

Focus on bioreactor and cooperate with other groups

Experimental Recap
Expectation:
● Optimal growth at higher nutrient concentrations

Result:
● Determined recommendations for media components

Importance:
Questions?
Sources
Pfeiffer, T., & Morley, A. (2014). An evolutionary perspective on the Crabtree effect. Frontiers in Molecular
Biosciences, 1. https://doi.org/10.3389/fmolb.2014.00017

Westman, J. O., & Franzén, C. J. (2015). Current progress in high cell density yeast bioprocesses for bioethanol
production. Biotechnology Journal, 10(8), 1185–1195. https://doi.org/10.1002/biot.201400581

Carothers, J. (2020). Cell Growth Kinetics [PowerPoint slides]. Retrieved from

https://docs.google.com/presentation/d/1cUlJ0z0CKwz2BbArcaoWbzzOpReqILaueoR85UHo_Fg/edit#slide=id.g42d1
5d8386_0_79