Effects of Raspberry Juice on Microbial Flora. Max O Malley Grade 9 Central Catholic High School

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1 Effects of Raspberry Juice on Microbial Flora Max O Malley Grade 9 Central Catholic High School

2 Antimicrobial effects of berries Types of berries Raspberry Cranberry Blueberry Previous Studies Cancer Cells antioxidant activity Staphylococcus aureus and Escherichia coli

3 Raspberry Health Benefits Cancer and disorders Cardiovascular Immune System Heart Disease Digestion

4 Escherichia coli Inhabit intestinal tract Many different types Many are not harmful Different types of infections Serves as a common prokaryotic cell model

5 E. coli and Raspberry Juice E. coli and other bacteria can cause many infections Raspberry Juice is claimed to prevent these Does Raspberry juice affect E. coli populations?

6 Purpose To determine if raspberry juice has an affect on E. coli survivorship in vitro

7 Hypothesis Null Hypothesis: Raspberry juice does not significantly affect E. coli survivorship in vitro. Alternative Hypothesis: Raspberry juice does affect E. coli survivorship in vitro.

8 Materials Labeling tape Ethanol E. coli DH5 Alpha Micropipettes Micro rack Micro tubes Raspberry Juice Sterile Dilution Fluid (100mM KH₂PO₄, 100mM K₂HPO₄, 10mM MgSO₄, 1mM NaCl) Luria Broth agar plates Spread bar Incubator Matches Klett Spectrophotometer

9 Procedure 1. E. coli was grown overnight in sterile LB 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 (37 C) until a density of 50 Klett spectrophotometer units was reached. This represents a cell density of approximately 10⁸ cells/ ml. 4. The culture was diluted in sterile dilution fluid to a concentration of approximately 10⁵ cells/ml. 5. Sterilized Raspberry juice was mixed with the appropriate amount of SDF to create raspberry juice concentrations of 1%, 0.1%, and 0.01%.

10 Procedure μl of cell culture was then added to the raspberry juice solutions, yielding a final volume of 10 ml and a cell density of approximately 10 3 cells/ml. 7. The solutions were vortexed and allowed to sit at room temperature for 15 minutes. 8. After vortexing to evenly suspend the cells, 100 μl aliquots were removed from the tubes and spread on a set of regular LB plates. 9. The plates were incubated at 37 C for 24 hours. 10. The resulting colonies were counted visually. Each colony was assumed to have arisen from one cell.

11 Chart of Liquid Concentration 0 R 0.01 R 0.1 R 1 R E. coli 0.1 ml 0.1 ml 0.1 ml 0.1 ml Sterile Dilution Fluid 9.9 ml 9.89 ml 9.8 ml 8.9 ml Raspberry Juice 0 ml 0.01 ml 0.1 ml 1 ml Total Volume 10 ml 10 ml 10 ml 10 ml

12 Colony Count Raspberry juice effects on E. coli survivorship Average E. coli Colony Survivorship Concentrations of Raspberry Juice P-value: R 0.01 R 0.1 R 1 R

13 Dunnett s Test Analysis T-Critical (Significant) = 3.29 Variable Concentration T Value Interpretation 0.01% Raspberry Juice Significant 0.1% Raspberry Juice 0 Not Significant 1% Raspberry Juice 7.5 Significant

14 Conclusions The null hypothesis that raspberry juice does not significantly affect E. coli survivorship in vitro was rejected for the concentrations of 1% and 0.01%, but were not rejected for 0.1%.These results indicate that 1% raspberry juice positively influenced E. coli survivorship, and 0.01% negatively influenced E. coli survivorship.

15 Limitations The E. coli planting was not exactly synchronized, possibly contributing to varying colony counts. Does not account for dilution in the body

16 Extensions and Future Studies Larger sample sizes will be employed in the next round of testing. More variables will be tested. Higher and lower concentrations of raspberry juice will be tested. More species of bacteria will be tested.

17 References "Theodor Escherich and the Discovery of E. Coli." Bright Hub. N.p., n.d. Web. 03 Jan < "E-coli." TheFreeDictionary.com. N.p., n.d. Web. 03 Jan < Cooper, Geoffrey M. Cells As Experimental Models. U.S. National Library of Medicine, n.d. Web. 03 Jan < "E. Coli the Biotech Bacterium Biotech Learning Hub." Biotechnology Learning Hub RSS. N.p., n.d. Web. 03 Jan < terium>. "Food & Feed Research." Different Antimicrobial Effects of Raspberry Depending on the Method of Active Components Isolation. N.p., n.d. Web. 03 Jan < =31&cntnt01sortby=magazine_id&cntnt01sortorder=asc&cntnt01summarytemplate =current&cntnt01detailtemplate=detaljno&cntnt01cd_origpage=180&cntnt01maga zineid=153&cntnt01returnid=188>. "Raspberries." Raspberries. N.p., n.d. Web. 03 Jan <