Download Antimicrobial Effects of Olive Oil

Antimicrobial Effects of Olive
Oil
Nathan Schollaert
9th Grade
Central Catholic High School
3rd Year participating in PJAS
Problem
• How does Olive Oil, a substance regarded as one of the
healthiest fats on earth, affect survivorship of the bacteria
Escherichia coli and Staphylococcus epidermidis?
Olive Oil
• Scientific Name: Olea Europaea
• Regarded as one of the world’s healthiest fats
• Used for cooking
• One of the world’s oldest cooking ingredients
Chemical Makeup of Olive Oil
• Olive Oil fat is a phospholipid
• Phospholipids have a special ability to become flexible and
semi-permeable
• Olive Oil fat is a monounsaturated fatty acid
Previous Studies
• Previous studies show that olive oil has positive effects on
certain bacteria in the human body
Microbial Flora
• Refers to microorganisms living in the gastrointestinal tract of the
human body
• Little is known about the association between humans and their
microbial flora
• The four effects are mutualistic, parasitic, pathogenic, and commensal
• Perform functions beneficial to the host, including the manufacturing
of essential vitamins, and the prevention of colonization by
undesirable microbes
• Human foods and supplements may have unintended effects on the
flora populations and their functions
Escherichia coli
• Largest group of known bacteria
• Resides in the lower human tract system
• Gram (-) Bacteria
• Common prokaryotic cell model used in scientific studies
Staphylococcus epidermidis
• Large group of Gram (+) Bacteria
• Resides on external layer of human skin
• Seen as a harmful bacteria
• Most strains are not harmful, as they reside on human skin
Purpose
To determine what effects Olive Oil might have on E. coli and
Staphylococcus epidermidis survivorship
Hypotheses
• Null Hypothesis- The Olive Oil will not have a
significant effect on E. coli or Staph survivorship
• Alternate Hypothesis- The Olive Oil will
significantly reduce survivorship on E. coli and
Staph
•
•
•
•
•
•
•
•
•
•
•
•
•
Materials
LB agar plates (0.5% yeast extract, 1% tryptone, 1% sodium chloride)
E. coli
Staphylococcus epidermidis
Sterile Dilution Fluid [SDF] (100mM KH2PO4, 100mM K2HPO4, 10mM MgSO4, 1mM NaCl)
Sterile Test Tubes
Sterile Spreader bars
Sterile Micropipets
Sterile Macropipets
Incubator
100% Virgin Olive Oil
Ethanol
Vortex
Klett Spectrophotometer
Procedure
1. E. coli and Staph were 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. The Olive Oil was sterile filtered through a 0.22 micron syringe filter.
Olive Oil was mixed with the appropriate amounts of SDF to create Olive
Oil concentrations of 10%, 1%, and 0.1%.
Procedure
6. 100 µL aliquots of cell culture was then added to the olive , yielding a final volume of
10 mL and a cell density of approximately 103 cells/mL.
7. The solutions were vortexed and allowed to sit at room temperature for a 10 minute
incubation period.
8. After vortexing to evenly suspend the cells, 100 µL aliquots were removed from the
tubes and spread on LB plates.
9. The plates were incubated at 37˚C for 18 hours.
10. The resulting colonies were counted visually. Each colony was assumed to have arisen
from one cell.
Concentration Table (ppm)
0% (Control)
0.1% Olive Oil
1% Olive Oil
10% Olive Oil
Microbe
0.1 ml
0.1 ml
0.1 ml
0.1 ml
SDF
9.9 ml
9.89 ml
9.8 ml
8.9 ml
Olive Oil
0 ml
0.01ml
0.1 ml
1 ml
Total Volume (ml)
10 ml
10 ml
10 ml
10 ml
Olive Oil Effects on E. coli
P-Value 9.73E-05
Dunnett’s Test Results (E. coli)
Concentration
Groups
T Value
T-Crit Value
Results
0.1%
2.72
3.10
Not Significant
1%
3.16
3.10
Significant
10%
7
3.10
Significant
Olive Oil Effects on Staph
P-Value 0.473591
Conclusion
• The null hypothesis was accepted for the 0.1% concentration of E.
coli. Null accepted for all Staph concentrations. Null hypothesis was
rejected for the concentrations of 1% and 10% of E. coli.
• The results and the statistical analyses indicate that 1% and 10% olive
oil concentrations significantly increased E. coli survivorship
Limitations
• Only one type of olive oil was tested
• Some cells could have had longer or shorter exposure times.
• The plating was not perfectly synchronized.
• Only Prokaryotic Cells were tested
• Only one type of exposure used
• Only one exposure time
Extensions
• One could use a Eukaryotic cell model, such as Yeast
• Prolonged exposure time to the E. coli and Staph
• Use different types of Olive Oil
• Synergistic effect of oil with other substances
Works Cited
• http://www.medicinenet.com/script/main/art.asp?articlekey=5549
http://web.uconn.edu/mcbstaff/graf/Student%20presentations/S%20epidermidis/s
epidermidis.html
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2807625/
http://www.mayoclinic.org/diseases-conditions/e-coli/basics/definition/con20032105
http://www.cdc.gov/ecoli/
http://www.mayoclinic.org/healthy-living/nutrition-and-healthy-eating/expertanswers/food-and-nutrition/faq-20058439
http://www.oliveoiltimes.com/olive-oil-health-benefits
http://www.cnn.com/2013/02/26/health/five-things-olive-oil/
https://www.pritikin.com/your-health/healthy-living/eating-right/1103-whats-wrongwith-olive-oil.html#.VKiIqUc8KrU
E. coli ANOVA
Anova: Single Factor
SUMMARY
Groups
Count
Sum
Average
Variance
Column 1
6
1952
325.3333
629.8667
Column 2
6
2290
381.6667
4712.667
Column 3
6
2342
390.3333
463.0667
Column 4
6
2980
496.6667
4333.867
ANOVA
Source of
Variation
SS
df
MS
Between
Groups
92040.67
3
30680.22
Within
Groups
50697.33
20
2534.867
142738
23
Total
F
12.10329
P-value
9.73E-05
F crit
3.098391
Staph ANOVA
Anova: Single Factor
SUMMARY
Groups
Count
Sum
Average
Variance
Column 1
6
2061
343.5
755.9
Column 2
6
1982
330.3333
997.4667
Column 3
6
1874
312.3333
4759.067
Column 4
6
1807
301.1667
3239.367
ANOVA
Source of
Variation
Between
Groups
Within
Groups
Total
SS
df
MS
6354.333
3
2118.111
48759
20
2437.95
55113.33
23
F
0.868808
P-value
0.473591
F crit
3.098391