the effect of muscle supplement on yeast cell growth michael gemperle central catholic high school...
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The Effect of Muscle Supplement
on Yeast Cell Growth
Michael GemperleCentral Catholic High SchoolGrade 9
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Problem
• Companies selling muscle supplements claim the supplements will help increase muscle growth.
• In 2014 approximately 32 billion dollars in supplements were sold globally.
• Are cells affected by muscle supplements?
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Genuine Muscle Milk®• Popular brand of muscle supplement• Consumed in liquid form• The major active ingredient is whey protein
Nutrition Facts
Major Ingredients: Calcium Sodium Casseinate, Milk Protein Isolate, Whey Protein Concentrate
Grams % Daily Value
Total Fat 7 11
Cholesterol 10 3
Sodium 260 11
Potassium 620 18
Total Carbs 6 2
Protein 20 40
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Whey Protein• Mixture of globular proteins isolated from whey, a
byproduct of cheese production• Contains all the essential amino acids• Fast digesting protein, 80-90% absorbed by body• Marketed as a dietary and muscular supplement
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Muscle• Muscle anatomy and physiology are partly
maintained by the available amino acid pool.
• Proteins, made of amino acid chains, help build muscle tissue.
• Foods containing high amounts of protein include eggs, meat, fish, and milk.
• Many muscle supplements are enriched with protein (whey) to help repair and grow muscle tissue used during exercise/workouts.
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Yeast
• Unicellular eukaryotic Fungi
• Common model organism
• Saccharomyces cerevisiae - species of budding yeast used in research and food industry
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Purpose
To determine if Genuine Muscle Milk® has a significant effect on
the growth of Saccharomyces cerevisiae.
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Experiment Hypotheses
Hypothesis:Genuine Muscle Milk® will significantly
increase yeast cell growth.
Null Hypothesis:Genuine Muscle Milk ® will have no
significant effect on yeast cell growth.
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Materials
• Klett-Summerson Photoelectric Colorimeter• Saccharomyces cerevisiae• Genuine Muscle Milk® Protein Nutrition Shake• Sterile YE broth (1% Yeast Extract)• Sterile YEP broth (1% Yeast Extract, 2%
Peptone)• Sterile test tubes• Sterile side-arm flasks• Sterile pipettes• Sterile water
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Colorimeter• Klett-Summerson Photoelectric Colorimeter
• Scale readings (Absorbance) are directly proportional to the concentration, according to Beer’s Law
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Procedure
1. 8.5 ml of YE broth was added to 16 sterile side arm flasks.
2. 8.5 ml of YEP broth was added to a new set of 16 sterile side arm flasks.
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Procedure3. Sterile water and Muscle
Milk® were added to 5 flasks of both YE and YEP broth to make a 0.1% dilution.
4. Sterile water and Muscle Milk® were added to 5 flasks of both YE and YEP broth to make a 1% dilution.
5. 0.5 ml of yeast cells were added to 15 of both YE and YEP flasks.
+ 8.5 ml broth+ 0.9 ml sterile water+ 0.1 ml Muscle Milk ®
+ 0.5 ml yeast cells 1%
0.1%
+ 8.5 ml broth+ 0.99 ml sterile water+ 0.01 ml Muscle Milk®
+ 0.5 ml yeast cells
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Procedure
6. Klett readings were taken every 30 minutes for 15 hours.
7. The final reading taken at stationary growth phase.
8. Data was recorded and analyzed.
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Data Analysis
-20
-10
0
10
20
30
40
50
0% YEP0.1% YEP1% YEP0% YE0.1% YE1% YE
Mean
Ch
an
ge in
Ab
sorb
an
ce (
ku
)
Hours
Mean Change in Absorbance verses Time
рYE3 = 0.5334рYEP3= 0.2889
рYE15 = 6.893E-4рYEP15= 4.244E-5
рYE12 = 5.581E-4рYEP12= 7.935E-4
рYE9 = 3.7E-3рYEP9= 0.5798
рYE6 = 0.5341рYEP6= 0.6126
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Data Analysis
0.00% 0.10% 1.00%0
5
10
15
20
25
30
35
40
45
50
30.8
23.5
7
Mean Total Absorbance Increase; YE
Aver-age In-crease, YE
Percent Muscle Milk®
Overa
ll C
hange in A
bso
rbance
(ku
)
0.00% 0.10% 1.00%0
5
10
15
20
25
30
35
40
45
50
34
44
25.8
Mean Total Absorbance Increase; YEP
Aver-age In-crease, YEP
Overa
ll C
hange in A
bso
rbance
(ku
)
Percent Muscle Milk®
р= 6.893E-4 р= 4.244E-5
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Analysis of Variance (ANOVA)
H0: μ0%= μ0.1%= μ1%
H1: not all means are equalα = 0.05
Media
F critical F calculated p-value Interpretation
YE 4.1028 16.4434 6.893E-4 Reject H0
YEP 3.8853 26.1258 4.244E-5 Reject H0
Reject Ho; There is a significant difference in the means.
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Dunnett’s Testtd =
Mi – Mc
√2MSEnh
YE α = 0.05
Percent Muscle Milk® t critical t d
Interpretatio
n
0.1 2.57 -1.738 Not Significant
1.0 2.57 -5.665 Significant YEP α = 0.05
Percent Muscle Milk® t critical t d
Interpretatio
n
0.1 2.50 3.965 Significant
1.0 2.50 -3.251 Significant
Control sample is 0% Muscle Milk®
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Conclusion
YE: Experiment Hypothesis: Not Supported
The yeast cells exposed to Muscle Milk did not show a significant increase in growth.
YEP: Experiment Hypothesis: Supported
The yeast cells exposed to Muscle Milk showed a significant increase in growth.
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Interpretation• At the 1% Muscle Milk concentration the solution
was opaque making it difficult to use with colorimeter.
o Cell growth in both YE and YEP broth for the 1% Muscle Milk was significantly less than for 0%.
• A Student-t test was conducted on the mean growth in YE and YEP broth at 0% Muscle Milk.
o There was no significant difference in the growth between the YE and the YEP broth at the 0% exposure.
• Increase in cell growth in YEP broth at 0.1% Muscle Milk is likely the result of the additional nutrition.
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Limitations/Extensions• Limitations:
Time required to reach stationary phase growth
Temperature conditions were not ideal 30 minute reading interval limited replicates
• Extensions Increase number of replicates Test additional Muscle Milk ® concentrations Test growth using whey isolate Quantify cell growth using hemocytometer or
plating
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Works Cited1. Bio Factsheet, Number 114, “Practical Techniques in MicrobiologyII: Measuring the
Growth of Microorganisms”, September 2002, <www.curriculumpress.co.uk>.2. Current Protocols in Molecular Biology 13.0.1-13.0.4 (John Wiley & Sons Inc 2008)3. David Lariviere, Forbes "Nutritional Supplements Flexing Muscles As Growth Industry“
<http://www.forbes.com/sites/davidlariviere/2013/04/18/nutritional-supplements-flexing-their-muscles-as-growth-industry>.
4. Exploreyeast "Yeast, a Cellular Model“ <http://www.exploreyeast.com/article/yeast-cellular-model>.
5. Heidi Skolnik, Andrea Chernus "How our bodies use protein“ <http://www.humankinetics.com/excerpts/excerpts/how-our-bodies-use-protein>.
6. Hemi Weingarten "What's In Muscle Milk?" <http://blog.fooducate.com/2011/12/05/whats-in-muscle-milk>.
7. “How Fast Do You Digest Whey Protein?” <http://www.livestrong.com/article/520142-how-fast-do-you-digest-whey-protein>.
8. Jeff S. Volek, Ph.D., R.D, “Whey vs. casein protein” <http://www.nutritionexpress.com/showarticle.aspx?articleid=787>.
9. Kansas State University, “Yeast Experiments: Baker’s Yeast and It’s Life Cycle”, <http://www.phys.ksu.edu/gene/a1.html>.
10. Megan Nantel "Why Is Yeast Used in Research?" <http://futurescienceleaders.org/researchers2012/2013/03/why-is-yeast-used-in-research>.
11. Operating Instructions Klett Photoelectric Colorimeter, Scienceware,005B-0997-5H-1-IH, REV 5/10.
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Thank you
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95% Confidence Interval for Total Mean Absorbance Increase
0% YE 0.1% YE 1% YE0
5
10
15
20
25
30
35
40
45
50
YE
Abso
rbance
(ku
)
0% YEP 0.1% YEP 1% YEP0
5
10
15
20
25
30
35
40
45
50
YEP
Abso
rbance
(ku
)
р= 6.893E-4 р= 4.244E-5