Method development of flow cytometry analysis of yeast and lactic acid bacteria viability for alcohol fermentation
(2023) KMBM01 20231Applied Microbiology
Biotechnology
- Abstract
- The usage of yeast in conventional brewing techniques involving alcoholic fermentation is an age-old process. Analyzing the viability and obtaining physiological information regarding the microbial diversity of the fermentation samples plays a pivotal role in obtaining real-time information as well as developing a more efficient process design. While traditionally, methods like methylene blue cell staining and/or colony counting techniques provide viability information, their arduous nature and general inconvenience has resulted in the adoption of flow cytometry to not only analyze individual cells but also in-depth cell physiological information in a matter of minutes. The primary focus of this project was to develop a method which... (More)
- The usage of yeast in conventional brewing techniques involving alcoholic fermentation is an age-old process. Analyzing the viability and obtaining physiological information regarding the microbial diversity of the fermentation samples plays a pivotal role in obtaining real-time information as well as developing a more efficient process design. While traditionally, methods like methylene blue cell staining and/or colony counting techniques provide viability information, their arduous nature and general inconvenience has resulted in the adoption of flow cytometry to not only analyze individual cells but also in-depth cell physiological information in a matter of minutes. The primary focus of this project was to develop a method which utilized flow cytometry to monitor the microbial diversity of the complicated fermentation matrix, in particular the cell concentration and viability of the yeast Saccharomyces cerevisiae and Lacticaseibacillus casei. The developed method offers the quantitative viability analysis of rehydrated freeze-dried yeast and potential lactic acid bacteria present in the fermentation matrix through flow cytometry and better understand the microbial interactions within the fermentation tanks. (Less)
- Popular Abstract
- Striving for absolut perfection
Have you ever consumed alcohol and wondered how an ordinary organism such as yeast can create something as extraordinary as vodka? This is what happens almost everyday at The Absolut Company, the third largest spirit producer in the world! With yeast playing such a crucial role in ensuring a final product of highest quality, understanding its viability over the course of the fermentation process provides information that can be used to optimize the process flow to the highest possible efficiency. My master’s project aims to analyze the viability of yeast and lactic acid bacteria by developing a method using the analytical technique of flow cytometry.
Yeast has been used in the process of both food... (More) - Striving for absolut perfection
Have you ever consumed alcohol and wondered how an ordinary organism such as yeast can create something as extraordinary as vodka? This is what happens almost everyday at The Absolut Company, the third largest spirit producer in the world! With yeast playing such a crucial role in ensuring a final product of highest quality, understanding its viability over the course of the fermentation process provides information that can be used to optimize the process flow to the highest possible efficiency. My master’s project aims to analyze the viability of yeast and lactic acid bacteria by developing a method using the analytical technique of flow cytometry.
Yeast has been used in the process of both food and alcoholic fermentation for centuries. While understanding the exact mechanism of fermentation took a long time, advances in the science of fermentation microbiology have undoubtedly got us there. In The Absolut Company, yeast is added to a “mash” made up of water, wheat flour, and a combination of enzymes and allowed to ferment in the fermentation tanks.
Lactic acid bacteria are also fairly common in alcoholic fermentations. They are predominantly unwelcome though, as they can potentially compete with yeast and reduce the final yield. Therefore, monitoring the viability and growth of lactic acid bacteria is also a necessity as it can be an indicator of upcoming hygiene-related problems in the fermentation tanks. While a number of existing analytical techniques are capable of analyzing the viability of yeasts and lactic acid bacteria during fermentation,
they all pose significant disadvantages. For instance, methods such as colony forming unit analysis and optical microscopy analysis provide the viability information we are looking for but at the expense of time, energy, and resources. Despite taking a day or two before obtaining usable results, the colony forming unit method still only provides information regarding the live cells and nothing for the dead and total cell populations.
This is where flow cytometry makes an entry as it not only is super fast and efficient, it also only requires a small amount of resources and provides simultaneous quantitative information regarding both the yeast and bacterial cells in a matter of minutes. While flow cytometry has been used in the analysis of yeast cells since the 70s, the cells in question have always been from a pure culture. The samples analyzed during the course of this project are diluted from the complicated mash matrix which poses a significant disadvantage due to the bacterial cells being too small and getting lost within the mash particles. Once diluted, the samples are stained with different stains specific to live, dead, and damaged cells and finally analyzed using the flow cytometer.
In this report, you will see the various settings and trial and error involved in setting up a flow cytometry analysis method, specific to the flow cytometer at hand, for the combined quantitative analysis of yeast and lactic acid bacteria in a fermentation mash sample. (Less)
Please use this url to cite or link to this publication:
http://lup.lub.lu.se/student-papers/record/9125957
- author
- Komahan, Megha LU
- supervisor
- organization
- course
- KMBM01 20231
- year
- 2023
- type
- H2 - Master's Degree (Two Years)
- subject
- keywords
- Flow cytometry, yeast, lactic acid bacteria, fermentation, cell viability, applied microbiology
- language
- English
- id
- 9125957
- date added to LUP
- 2023-06-19 12:20:23
- date last changed
- 2023-06-19 12:20:23
@misc{9125957,
abstract = {{The usage of yeast in conventional brewing techniques involving alcoholic fermentation is an age-old process. Analyzing the viability and obtaining physiological information regarding the microbial diversity of the fermentation samples plays a pivotal role in obtaining real-time information as well as developing a more efficient process design. While traditionally, methods like methylene blue cell staining and/or colony counting techniques provide viability information, their arduous nature and general inconvenience has resulted in the adoption of flow cytometry to not only analyze individual cells but also in-depth cell physiological information in a matter of minutes. The primary focus of this project was to develop a method which utilized flow cytometry to monitor the microbial diversity of the complicated fermentation matrix, in particular the cell concentration and viability of the yeast Saccharomyces cerevisiae and Lacticaseibacillus casei. The developed method offers the quantitative viability analysis of rehydrated freeze-dried yeast and potential lactic acid bacteria present in the fermentation matrix through flow cytometry and better understand the microbial interactions within the fermentation tanks.}},
author = {{Komahan, Megha}},
language = {{eng}},
note = {{Student Paper}},
title = {{Method development of flow cytometry analysis of yeast and lactic acid bacteria viability for alcohol fermentation}},
year = {{2023}},
}