By: Christian Wawrzonek The Metabolic Enhancer Effect on Microbes.
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Transcript of By: Christian Wawrzonek The Metabolic Enhancer Effect on Microbes.
By: Christian Wawrzonek
The Metabolic Enhancer Effect on Microbes
Supplements taken to increase exercise duration and reduce recovery time.
GNC is a common distributor of “metabolic enhancers.” Some for diet, vitamins, fitness, body care, and sports
nutrition. How many of them really work? One recommended drug is L-Glutamine Powder.
Metabolic Enhancers
Glutamine Claims Decreases muscle deterioration during a
workout.
Used as a fuel for cells.
May increase protein metabolism.
Can increase hormone production.
One of the 11 nonessential amino acids.
The body produces glutamine from glutamic acid and ammonia.
Found in many protein rich food sources.
Glutamine
Nitrogen carrier in the body. Precursor for immune cell division. Important for proper immune system functioning. When damaged, the body uses glutamine for
repair. Studies show an increase in recovery
speed and fewer infections in correlation to glutamine dosage.
Glutamine Uses
Saccharomyces cerevisiae Two strands of Saccharomyces cerevisiae
were used. Bakers Yeast: best known for alcohol
fermentation and bread baking. Laboratory strain: used in many
cell/biochemical investigations (Jones, CMU).
Saccharomyces cerevisiae as a Model The most investigated cell in the world.
Used in microbiology due to its ease of manipulation and rapid growth.
Eukaryotic, shares similar biochemistry, cell cycle, and genetics with more advanced organisms, including humans.
Purpose Do Metabolic Enhancers really work?
To determine if the metabolic enhancer L-Glutamine Powder has any effect on the population growth and cellular respiration of Saccharomyces cerevisiae.
Hypothesis and Null Hypothesis Null Hypothesis: There will be no
significant difference between the population growth and cellular respiration of Saccharomyces cerevisiae from the control.
Alternative Hypothesis: Glutamine supplementation will yield a faster rate of population growth and cellular respiration of Saccharomyces cerevisiae.
Materials
Sterile Side Arm Flasks L-Glutamine Powder Saccharomyces cerevisiae
(Lab Strain) Sterile Micro Pipettes Sterile Macro pipettes Klett Spectrophotometer YEPD media (0.5% Yeast
extract, 2% peptone, 2% glucose)
Sterile Deionized water Glutamine Solution (4% [ ])
Rapid Rise Saccharomyces cerevisiae (Cooking Strain), Red Star brand
Micro Pipettes Macro Pipettes Balloons (23 cm Unique
Industries) 125 ml Erlenmeyer Flasks Deionized Water Sucrose (1% final [ ]) 100mL Graduated Cylinders Plastic Tub Plastic Film Wrap L-Glutamine Powder (GNC
Pro Performance)
Experiment 1 Experiment 2
Procedure #11. Saccharomyces cerevisiae was grown overnight in sterile
YEPD media.2. A sample of the overnight culture was added to fresh
media in a sterile sidearm flask.3. The culture was placed in an incubator (30°C) until a
density of 150 Klett spectrophotometer units was reached. This represents the stock solution of yeast.
4. Glutamine powder was dissolved in sterile water at a concentration of (4%).
5. The glutamine solution, yeast stock, YEPD media, and sterile water were added to sterile side arm flasks as follows:
6. The flasks were swirled to evenly dissolve glutamine solution.
7. Klett spectrophotometer readings were recorded at times: (0,30,60,90,120,165,210,240,285, and 1200 minutes)
Table 1 Flask A(x5) Flask B (x5) Flask C (x5)
Yeast 0.1mL 0.1mL 0.1mL
GlutamineSolution
100µl 10µl 0µl
Sterile Water 4mL 4.9mL 5mL
YEPD Media 4.9mL 4.9mL 4.9mL
Total 10mL 10mL 10mL
Concentrationof Glutamine
1% 0.1% 0%
Procedure #2(1) Glutamine powder was dissolved in sterile water at a
concentration of (4%).(2) Sugar, water, and glutamine solution were added
to 12 flasks in the ratios as follows:
(3) 7 grams of Red Star rapid rise Cooking Yeast was added to each flask and balloons were immediately affixed to each flask.
(4) The flasks were transferred to a warm (30 C) water bath.
(5) After 90 minutes of incubation, each balloon was removed from the flask (careful to prevent any leakage of gas; each balloon was pinched at the neck and twisted off).
Procedure#2 Continued(6) A plastic tub was filled with water.
(7)Each graduated cylinder was filled with water to the brim and sealed with plastic wrap.
(8)The cylinder was inverted and immersed into the water and the plastic wrap removed.
(9) The balloon was placed into the water with the mouth placed into the cylinder.
(10) The mouth was slowly released and the air was pumped into the graduated cylinder. The volume of gas was then recorded.
Table 2 Flask A (x3)
Flask B (x3)
Flask C (x3)
Flask D (x3)
Yeast 7 grams 7 grams 7 grams 7 grams
Sucrose(10%)
4mL 4mL 4mL 4mL
Distilled Water
31mL 35mL 35.9mL 36mL
Glutamine Solution (4%)
5mL 1mL .1mL 0mL
Total Volume
40mL 40mL 40mL 40mL
[Glutamine] 0.5% 0.1% .01% 0%
Glutamine Effect on Yeast Population
0
50
100
150
200
250
300
350
0 30 60 90 120 165 210 240 285 1200
Time (Minutes)
Kle
tt U
nits
0%0.10%
1%
Glutamine Effects on Yeast Respiration
0
20
40
60
80
100
120
140
0% 0.01% 0.10% 0.50%
[Glutamine]
CO
2 E
v(m
Ls)
Sig.Sig.
Sig.P>.05P>.05
P>.05
ANOVAsTest F value F critical P value Interp.
Respiration Experiment
14.9587 3.8625 0.000764 Significant
Growth Experiment
Growth at 120 Min.
19.227 4.256 0.000563 Significant
240 Min. 12.845 4.256 0.002307 Significant
285 Min. 4.515 4.256 0.043851 Significant
1200 Min. 0.858 4.256 0.455861 Insignificant
Time Variable Compared
t value Interpretation
120 Minutes 1% vs. c. 3.88 Significant
120 Minutes 0.1% vs. c. 6.1 Significant
240 Minutes 1% vs. c. 0.038 Insignificant
240 Minutes 0.1% vs. c. 4.51 Significant
285 Minutes 1% vs. c. 0.861 Insignificant
285 Minutes 0.1% vs. c. 2.12 Insignificant
Dunnett's Tests#1t critical = 3.22α = 0.05
Dunnett's Tests#2
Variable Compared
t value Interpretation
0% [stock] to 0.01% [stock]
4.41 Significant
0.1% vs C 5.08 Significant
0% [stock] to 1% [stock]
7 Significant
α = 0.05t critical = 3.73
Conclusions #1 Alternative Hypothesis Accepted:
Glutamine powder yielded a faster rate of population growth of Saccharomyces cerevisiae at 120,240, and 285 minutes.
Failed to reject the Null Hypothesis at the time of 1200 minutes.
Dunnett’s test was significant for both concentrations at 120 and 240 minutes.
Conclusions #2 Accepted Hypothesis: Glutamine
supplementation significantly increased cell respiration (as measured by CO2 evolution) at all concentrations (0.5%, 0.1%, 0.01%).
Null Hypothesis Rejected Dunnett’s test showed all concentrations
significant.
Limitations and Extensions Much larger sample sizes, Might require a team of
researchers. Slight variations in timing of readings for growth
experiment (Only one Klett Spec available) Possible slight variations in optical properties of
side arm flasks. Variation in the balloon elasticity (trap the gas in
more quantitative manner). Balloons placed awkwardly on flask limiting CO2
intake. Properly place balloons.
References http://www.online-medical-dictionary.org/L+Glutamine.asp?
q=L+Glutamine http://mental-health.emedtv.com/glutamine/does-
glutamine-work.html http://www.vitamins-supplements.org/amino-acids/
glutamine.php http://en.wikipedia.org/wiki/Glutamine http://genome.wellcome.ac.uk/doc_wtd020808.html http://biochemie.web.med.uni-muenchen.de/Yeast_Biol/
03%20Yeast%20Metabolism.pdf http://www.bodybuilding.com/store/glutpep.html http://mental-health.emedtv.com/glutamine/glutamine.html