Influence of temperature on Saccharomyces cerevisiae fermentation speed

Influence of temperature on Saccharomyces cerevisiae fermentation speed

We are finally reunited, the three of us to develop our diabolical project… Our aim? Becoming rich by making ... BREAD ! But we want to process really quickly to sell our bread because this money will allow us to go to … IBIZA this summer !

So now the question is: How can we make our bread dough rise as fast as possible?

It is known that BREAD dough rises thanks to the process of yeast fermentation. Fermentation is a metabolic process which may occur without oxygen. In dough, yeasts convert a sugar (a carbohydrate for the more scientist) into alcohol and CO2. There is an equation for this process, for those who aren’t just interested in food :

C6H12O6 → 2 C2H5OH + 2 CO2`

We can see that one molecule of sugar is turned into two alcohol molecules and two carbon dioxide molecules. There is the explanation of the entire process :

http://test.classconnection.s3.amazonaws.com/529/flashcards/475529/jpg/lactic-acid-fermentation.jpg

But how to speed up this reaction? We decided to test the influence of the temperature on fermentation, to figure out what is the optimal one.

Experiment :

The only product of the reaction of fermentation that we can observe was the CO2 (gas). So we decided to measure the speed of the release of CO2 in order to estimate the reaction’s speed. We wanted to compare this speed at different temperatures.

Protocol :

• Add 15g of glucose in 300 mL of water in a plastic bottle
• Drill each bottle cap with a screw to introduce the tube (containing colorant) in the bottle
• Incubate all 4 bottles at 25, 27, 35, 37, 40°C for 10 minutes to bring the solution at correct temperature.
• Add 15g of bakery yeast in each bottle
• The capillary, going out of the incubator, is fixed horizontally using tape
• Make 3 marks at 10 cm of distance of each other.
• Measure the time the colorant takes to travel the 10 cm marked

→ Repeat this experiment as many times as possible to have a better approximation, to calculate the average speed of the colorant that is relevant to the reaction speed.

Set up for 30°C

Set up for 25°C

Set up for 35 and 37°C

Bottle in an incubator

Results :

The ink moved extremely slowly through the tube. Over a total time of 1h40 minutes, it traveled for 19mm, over 1h, then almost stopped.

Average speed (really low) =0,000556cm/s ~19,8mm/h.

 
The first value (at time zero) shows that the reaction doesn’t occur instantly but takes some time.

The ink moves slower over the first 20cm and faster over the second 20cm.

Average speed for the 2nd 20cm =1,7cm/s.

For the incubators at 30, 35 and 37°C, we observed that the ink starts by moving slowly, then speeds up really fast at one point, then slows down a few seconds later, and continues to push the ink until the end of the tube.

Our hypothesis to explain this phenomenon is that CO2 accumulates in front of the ink, therefore the pressure builds up, to a point where CO2 diffuses through the ink, which speeds up. After, the pressure goes down, so does the speed of the ink.

The measurements for 35 and 37°C were done on portions of the tube where the ink was going fast, so it doesn’t show accurately the speed of the ink.

For the fast-moving ink :

Average speed =2,28cm/s at 35°C

Average speed =2,48cm/s at 37°C

Conclusion :

We could conclude that the optimal temperature between 25, 30, 35, 37°C for bakery yeast fermentation is 37°C.

25°C is not enough for the fermentation to occur. Enzymes helping in yeast metabolism might not be functional at this temperature.

We put the bottle from the 25°C incubator into another one at 35°C, the reaction occurred and the colorant moved fast, so we’re sure that our set up had no problem.

Now we could ask ourselves if the pressure influence yeasts metabolism. We could repeat this experiment but changing the parameter studied : at a constant temperature, but at different pressure.

We could also compare different types of yeast, (with beer yeast) in order to show whether different type of yeast have different optimal conditions.

If you want to know more : 

• Yeast (wikipedia) : https://en.wikipedia.org/wiki/Yeast
• Fermentation (wikipedia) : https://en.wikipedia.org/wiki/Fermentation
• Blog about baker yeast fermentation for the cooking of bread : http://www.snv.jussieu.fr/bmedia/blepain/3pain/pain2.htm
• Link to a video showing how the ink moves : https://drive.google.com/a/cri-paris.org/file/d/0Bydr3Agf3FV6bzhBWEViLWd5dEU/view?usp=sharing

Amandine Maire, Roxane Jannin and Mathieu Szotowski

Influence of temperature and pigmentation yeast cells concentration at stagnation phase

For the Thermal Wars final project, our group was interested by pigments, temperature and yeast. Yeast are unicellular microorganisms of the fungus kingdom. They are used in baking or in alcohol beverages. If the conditions allow it (temperature, pH…) and if they have nutrients, cells grow and divide over time. They can be different colors according to their pigments. Different colors absorb and reflect light in different ways, and this can be measured thanks to a spectrophotometer.

There are three main phases in cells’ growth (lag phase, exponential growth phase, stagnation phase), and we had already studied the exponential growth phase during the week. After stagnation, the cells start to die. Eventually, they multiply again, and the cycle starts over (if the conditions are favorable to their division).

Our teacher Tamara had different colored strains of yeast, and we knew that yeast are particularly resistant during stagnation. Therefore, we combined all these elements to come up with an experiment that interested us all : How does temperature and pigmentation of yeast influence the cell concentration at stagnation phase?

Indeed, we were  wondering if pigments or temperature have an influence on cell concentration at stagnation phase, and how these factors were interwoven. This research could help if you need to pick yeast strains (for a study at stagnation phase) in function of their color.

To answer this question, we used two methods : microscopy and spectrophotometry. We used two methods because it is better to base your study on several and different manipulations in case the tendencies vary between each method. (Also, Tamara said we were smart enough to do both).

So we got to work: we made our 11 different colored yeast cultures multiply at three different temperatures, 25°C, 30°C and 35°C. We waited overnight so they would be all grown up and then got to measuring. For both methods, we needed the cells to be a little less concentrated to actually be accurate so we made dilutions. We then counted the cells under a microscope and used the spectrophotometer to measure the optical density (which is supposed to be closely related to cell concentration).

Our results

Using those two methods, we analysed the data, and plotted those two graphs:

These were some of our results: the graph to the left represents the microscopy method, where we simply counted cells in a small sample of the culture. In that graph, we’re representing the cell concentration in function of temperature and we can see that as the temperature increases between 25 and 25°C, the concentration of the yeast increases as well.

To the right, we represented optical density (the measures taken with the spectrophotometer) in function of temperature and here we see a difference compared to the actual cell concentration: for 30°C, the optical density is much lower! We weren’t expecting those results since optical density is supposed to be a good indicator for cell concentration and thought that both the left and right graph would look almost identical.

Conclusion

From our results, we could conclude that proceeding through several methods is always safer. Our results did seem to indicate that for lower temperatures, the cell concentration at stagnation phase were lower than at higher temperatures, however there isn’t much else we could conclude. To make sure we had accurate results, we wanted to simply reproduce the experiment as much as we could. We think that optical density might also be influenced by the color of the strains, this would make its relation to cell concentration flawed!

With that in mind, we thought we would reproduce the same experiment but with a different goal: to evaluate the efficiency of spectrophotometry for different colored cells!

If you want to know more:

A nice article summing up the growth phases of yeast: http://capricorn.bc.edu/wp/pathways/biology-bootcamp/yeast-techniques/#growth

A brief course on cells and the dynamics of growth: http://www.columbia.edu/cu/biology/courses/c2005/lectures/lec1_11.html

Another course on bacterial growth & generation time: http://study.com/academy/lesson/what-is-bacterial-growth-generation-time-curves-phases-stages.html

A scientific article on the stationnary phase of the bacterial life cycle: https://www.ncbi.nlm.nih.gov/pubmed/8257118

A wikipedia article on Bacterial Growth : https://en.wikipedia.org/wiki/Bacterial_growth

Another scientific article on the dynamics of Saccharomyces cerevisiae’s growth: https://www.sciencedirect.com/science/article/pii/0168165692901508

To go further into the study of the factors influencing cell concentration at stationnary phase: http://schaechter.asmblog.org/schaechter/2013/02/bacterial-antidepressants-avoiding-stationary-phase-stress.html

Our presentation of the project: https://docs.google.com/presentation/d/1lwf07pkZp9DUC3q728xI5gc8gLztR-amWk84gNtI9nU/edit#slide=id.gc6f90357f_0_0

June VK, Selim BS & Jacques CK