The document discusses an experiment on the effect of different salt concentrations on yeast cell growth. Saccharomyces cerevisiae yeast cells were exposed to solutions with salt concentrations ranging from 0.3g to 1.5g. Optical density measurements found that after 24 hours, yeast density increased in all samples, contradicting the hypothesis that high salt would inhibit growth. This suggests yeast have a higher salt tolerance than expected and can tolerate salt quantities thought to be harmful. Repeating the experiment with even higher salt levels could help determine the threshold where growth is reduced.

1. Yeast Cells vs Different Salt
Concentrations
By: Rawan Gebreil, Jasmine Arreola

2. Background research on Saccharomyces cerevisiae
● Saccharomyces cerevisiae is a species of yeast (single-celled fungus microorganisms). It belongs to Kingdom
Fungi and Domain Eukaryote. Production of fermented beverages and breads.Spoilage of foods and
beverages. Processing food waste. Production of food ingredients as a probiotic. Production of industrial
ethanol. The inability to use nitrate and ability to ferment various carbohydrates are typical characteristics of
Saccharomyces.
● S. cerevisiae is used in baking; the carbon dioxide generated by the fermentation is used as a leavening agent
in bread and other baked goods. Saccharomyces when translated means “sugar fungus”. That is what this yeast
uses for food. They are found in the wild growing on the skins of grapes and other fruits. Saccharomyces
cerevisiae has a rich metabolism that enables it to survive or grow in a wide range of environments eclipsing
those found in either fruits or bark, with varying nutrient availabilities—with both low and high carbon and
nitrogen concentrations. Serving as a nutrient source for bacterial, faunal and protistan predators, soil yeasts
contribute to essential ecological processes such as the mineralization of organic material and dissipation of
carbon and energy through the soil ecosystem.

3. Project objective:
What impact does salt have on the yeast growth
process?

4. Hypothesis and Predictions
➔ We hypothesized, that too much salt will have a negative effect on the yeast growth
and it will kill most of them.
● We predict that:
- 0.25g of salt concentration would not have a big effect on the yeast cells by not
reducing their cell density
- Starting from 0.5g of salt, it would start to affect the yeast growth by reducing their
density and kill them off
- 1.5g of salt, will have the most effect by killing most of the yeast cells and reducing
their density extremely

5. Materiels
➢ Salt
➢ Yeast
➢ YPD
➢ Distilled Water
➢ (10)- Test tubes
➢ (6) 200mL beakers
➢ (11) - 1 mL Serological Pipets
➢ (2) - 10 mL Disposable Plastic
Pipettes
➢ (12) - 3 mL Disposable Plastic
Pipettes
➢ (17) - Cuvettes
➢ Electronic scale
➢ Spectrophotometer
➢ Cotton rounds

6. Methods
● The methods employed in this experiment are strikingly similar to that of Lab E:
Working with Cell Cultures (Activities 1 and 2).
● To begin we started by preparing our 5 different salt solutions.
Salt Solutions (% weight)
0.3g salt in 49.7ml DI water = 0.7%
0.5g salt in 49.5ml DI water = 1%
0.7g salt in 49.3ml DI water = 1.5%
1.0g salt in 49ml DI water = 2%
1.5g salt in 48.5ml DI water = 3%

7. Methods #1
➔ Preparing our Yeast Culture
● 0.25g of yeast to 100 ml of DI water
➔ Preparing our Samples
● Obtained 5 test tubes and labeled them 1 thru 5
● Obtain 45 ml of YPD
● Use a 10 mL pipette to add 9 mL of YPD broth to each tube
● Use a 1.0 mL pipette to add 0.8 mL of salt solution to tubes 1-5 according to the table below.
● Use a 1.0 mL pipette to add 0.2 mL of the Yeast Culture to each tube.
Tube Broth Salt Solutions Yeast
1 9.0 0.8mL of 0.3g solution 0.2 mL
2 9.0 0.8mL of 0.5g solution 0.2 mL
3 9.0 0.8mL of 0.7g solution 0.2 mL
4 9.0 0.8mL of 1.0g solution 0.2 mL
5 9.0 0.8mL of 1.5g solution 0.2 mL

8. Methods #2
● Use a plastic dropper to transfer approx. 3 mL of YPD broth into one of the cuvettes. This is the “blank” you will
use to calibrate the spectrophotometer. Place the cuvette in the first (top) space of the left-side cuvette rack that
comes with the spectrophotometer.
● Gently invert culture tubes to resuspend yeast in the solution.
● Using a fresh plastic dropper for each tube, transfer approx. 3 mL of yeast from each tube into a cuvette. Place the
cuvettes in the left-side cuvette rack in order from top to bottom, with sample 1 at the top and sample 5 at the
bottom.
● The spectrophotometer should already be powered on. If not, turn on using the switch on the back, then follow
instructions on the LCD screen to let the machine perform its initial diagnostics.
● From the Home Screen on the spectrophotometer (“spec”), use the down arrow key to select “OD 600” and press
the return key.
● On the next screen, check that the Mode is set to “Direct.” If it is not, use the left or right arrow keys to change
the setting until it is. Use the down arrow key to select “Go” and press the return key.

9. Methods #3
● The spec will instruct you to place your “blank” in the sample compartment. Open the lid to the sample
compartment and place your cuvette containing only broth in the sample holder, making sure the clear
sides of the cuvette are on the left and right.
● BE SURE TO WIPE THE SIDES OF THE CUVETTE WITH A LAB TISSUE TO REMOVE ANY
SMUDGES.
● Close the lid to the sample compartment and press the “0.00” button on the spec.
● Once the display on the spec reads 0.000, open the compartment and remove the “blank” cuvette. Place it
in the top position of the right-side rack.
● You are now ready to take readings of your yeast samples. Begin with sample 1 at the top of the left-side
rack. Wipe off the cuvette with a lab tissue and place in the sample compartment. Allow the reading to
stabilize and record the OD in the table below. Remove the sample and place into the right-side rack.
● Repeat the step above for the four remaining samples, recording the OD as you go.

10. Getting Our Results
● After we got our results from the spectrophotometer at time 0, we incubated our
samples in water bath set at 30 C for 24 hours.
● After 24 hours we repeated all the steps from the slide above (Methods #3) for
our samples.
● Our results after 24 hours went over the limit of our spectrophotometer of 2.5 OD
for each sample which meant we had to dilute each sample.
● We did a dilution of 1.0 mL sample to 9 mL of distilled water and we did that to
each sample.

11. Results - Measurements of Optical Density of Salt
Solutions
➔ 0.3g of salt
Time 0: Optical Density of 0.424
After 24 hours: Optical Density of 0.848
➔ 0.5g of salt
Time 0: Optical Density of 0.244
After 24 hours: Optical Density of 0.837
➔ 0.7g of salt
Time 0: Optical Density of 0.235
After 24 hours: Optical Density of 0.897
➔ 1.0g of salt
Time 0: Optical Density of 0.282
After 24 hours: Optical Density of 0.956
➔ 1.5g of salt
Time 0: Optical Density of 0.238
After 24 hours: Optical Density of 1.41

12. Discussion
● Based on our experiment our hypothesis did not support our results considering
that we said “too much salt will have a negative effect on the yeast growth”. Our
results in the experiment showed that adding more salt will have a positive effect
on yeast growth.
● It showed that yeast cells have a higher tolerance for high salt concentrations by
increasing their densities extremely after 24 hours and that they can tolerate
higher salt quantities.

13. How would we have done different?
● What we believed we could have done to expand our results was doing a second
trial of different samples with high quantities of salt to see where the density of
the yeast cells began to decrease and see how much salt is too much for them.

14. Thank you

15. Works Cited
https://vdoc.pub/documents/the-rise-of-yeast-how-the-sugar-fungus-shaped-civilisation-s2a26l6h6v00
https://effca.org/microbial-cultures/production-of-microbial-cultures/yeasts-in-food-production/
https://drfungus.org/knowledge-base/saccharomyces-species/
https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/saccharomyces-cerevisiae
https://wiki.yeastgenome.org/index.php/What_are_yeast%3F