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The Effects of Gatorade on Microbial Survivorship
Christopher Samek Grade 9 Pittsburgh Central Catholic High School 1st Year in PJAS
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Problem Gatorade is constantly consumed to rehydrate the body.
At what concentrations does Gatorade affect the survivorship of human microflora?
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Sports Drinks First sports drink created by a chemist, William Owen, in the U.K. in 1927. In 1968, sports drinks began to revolutionize sports. Used for rapid replacement of electrolytes and fluid lost during exercise. Restores depleted muscle carbohydrate storages that cause muscle failure and fatigue.
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Gatorade Ingredients Water Glucose Sucrose Dextrose
Total Carbohydrates 34g Citric acid Natural flavor Sodium citrate Monopotassium phosphate Gum arabic Red 40 Glycerol ester of rosin Protein 0g Sodium 270 mg Potassium 75mg Total Fat 0g
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Microbial Flora Microorganisms that reside in or on the surface of the body Respiratory, urinary, digestive tracts, and skin Consists of mostly bacteria, fungus, protists, and archea Most are harmless, but some can cause disease Most studies based on ingested materials study human cells, but human are a collection of cells and symbiotants Estimated 10% body mass is symbiotants
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Staphylococcus epidermidis
Human skin flora Gram (+) bacterium Surrounded by a simple cell wall Most forms are non-pathogenic Forms biofilms on plastic devices
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Escherichia coli Large and diverse group of gram (-) bacteria
Surrounded by an extra cell wall composed of lipopolysaccharides Free living, symbiotes, or pathogens Most strains are non-pathogenic Serves as a common prokaryotic cell model
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Purpose To determine what effect Gatorade has on Staph e. and E. coli survivorship
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Hypotheses Null : Gatorade will not have a significant effect on the survivorship of E. coli or Staph. e. Alternate : Gatorade will have a significant effect on the survivorship of the E. coli and/or Staph. e.
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Materials 64 LB Agar Plates (0.5% yeast extract, 1% tryptone, 1% sodium chloride) Escherichia Coli (DH5-alpha) Staphylococcus Epidermidis Sterile Dilution Fluid [SDF] (100mM KH2PO4, 100mM K2HPO4, 10mM MgSO4, 1mM NaCl) Sterile test tubes Racks Sterile spreader bars Matches Incubator Ethanol Bunsen burner Vortex Gatorade Micropipettes Turn table Labelling tape 0.22 Micron sterile filter
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Procedure E. coli and Staph. e. were grown overnight in sterile LB media. Samples of the overnight culture were added to fresh media in a sterile sidearm flask. The cultures were incubated until a density of 50 Klett spectrophotometer units was reached. This represents a cell density of approximately 10^8 - 10^9 cells/mL. The cultures were diluted in sterile dilution fluid to a concentration of approximately 10^5 cells/mL. The Gatorade was diluted with sterile dilution fluid to concentrations of 0%, 5%, 10%, and 25% to total 9.9 mL.
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Table of Concentrations
0% Concentration 5% 10% Concentration 25% Concentration Sterile Water 9.9 mL 9.4 mL 8.9 mL 7.4 mL Gatorade 0 mL 0.5 mL 1 mL 2.5 mL Staph. e. / E. coli 0.1 mL Total Volume 10 mL
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Procedure 0.1 mL of cell culture was then added to the test tubes, yielding a final volume of 10 mL and a cell density of approximately 10^3 cells/mL. The tubes were allowed to incubate at room temperature for 10 minutes. After vortexing to evenly suspend cells, 0.1 mL. aliquots were removed from the tubes and spread on LB agar plates. The plates incubated at 37°C overnight. The resulting colonies were counted. Each colony is assumed to have arisen from one cell.
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Results P-Value = 0.49 Not Significant Surviving Colonies
0% Concentrations of Gatorade
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Results Survivorship of Staph. e. when Exposed to
P-Value = 1.10E-34 Significant Survivorship of Staph. e. when Exposed to Various Concentrations of Gatorade 0% 5% 10% 25%
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Dunnett’s Test (Staph. e.)
T-Critical – 2.88 Alpha – .05 Concentration of Gatorade T-Value Significant? 5% 8.93 Significant 10% 5.40 25% 11.12
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Conclusion Staph. e. The null hypothesis can be rejected.
The alternate hypothesis can be accepted. Gatorade promoted bacterial growth in small quantities. Gatorade killed the bacteria in larger quantities. E. coli The null hypothesis fails to be rejected. The alternate hypothesis can be rejected. Gatorade had no effect on the E. Coli in any amount.
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Limitations / Extensions
Plating not perfectly synchronized Slightly shorter or longer exposure time Only two species tested Extensions More trials to create a better basis for evaluating the results Vary exposure time in a two-factor experiment Use agar infused plates
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References
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Liquid Exposure Raw Data (E. coli)
0% 5% 10% 25% 85 80 71 89 81 96 88 82 91 84 79 74 77 90 76 86 87
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Liquid Exposure Raw Data (Staph. e.)
0% 5% 10% 25% 392 569 302 203 406 582 317 198 421 579 298 187 394 566 319 196 412 304 199 399 578 301 184 414 589 297 197 411 572 294 201
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E. coli Anova Anova: Single Factor SUMMARY Groups Count Sum Average
Variance Column 1 8 678 84.75 Column 2 681 85.125 Column 3 653 81.625 Column 4 659 82.375 ANOVA Source of Variation SS df MS F P-value F crit Between Groups 3 Within Groups 28 Total 31
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Staph. e. Anova Anova: Single Factor SUMMARY Groups Count Sum Average
Variance Column 1 8 3249 Column 2 4601 68.125 Column 3 2432 304 Column 4 1565 ANOVA Source of Variation SS df MS F P-value F crit Between Groups 3 E-34 Within Groups 28 Total 31
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