The Effect of Oregano Oil on E. coli Dominic Veltri Grade 9 Pittsburgh Central Catholic High School

Purpose Problem Does Oregano Oil have antimicrobial activity? Would the consumption of Oregano Oil alter human microflora populations? To determine the effects of Oregano Oil on E. coli cell survivorship.

Oregano Oil Oregano Oil Made of natural components Used to cook Oil Used for therapy or in some doses cooking Made of natural components No major side effects Superior to antibiotics

Contents in Oregano Oil Leaves and flowers of oregano Naturally grown Carvacrol Thymol Vitamins A, C, E Iron Copper Calcium Other natural ingredients

Microbial Flora Present in the respiratory, urinary, and digestive tracts as well as on skin. Consists of mostly bacteria, few protists and fungi. Most are harmless, but some can cause diseases. Influences human anatomy, physiology, and overall health

Escherichia coli Large and diverse group of gram (-) bacteria Free living, symbionts, or pathogens Most strains are not pathogenic Common mammalian intestinal symbiont Serve as a common prokaryotic cell model Environmental prokaryotic model

Hypotheses Null: The Oregano Oil will have no significant effect on the survivorship of E. coli cells. Alternative: The Oregano Oil concentrations will have a significant negative effect on the survivorship of E. coli cells.

Materials Sterile test tubes Latex Gloves Laboratory Goggles Micropipettes Sidearm Flask Sterile Filters Incubator (37o C) Vortex Machine E. coli(DH5 Alpha) LB Agar Plates(LB Media) (1% Tryptone, 0.5% Yeast extract, 1% NaCl) Spread bars Ethanol Matches Sterile Dilution Fluid (SDF) (100mM KH2PO4, 100mM K2HPO4, 10mM MgSO4, 1mM NaCl) Oregano Oil Bunsen Burner

Procedure E. coli was grown until a density of 50 klett spectrophotometer density was reached. This was approximately 10^8 cells/mL. The culture was diluted in sterile dilution fluid to a concentration of approximately 10^5 cells/mL. Stocks of the variables were created. The following ingredients were added to the tubes to create the desired concentrations:

Concentration Table 0% (Control) 0.10% 1% 10% E. coli 0.1 ml   0% (Control) 0.10% 1% 10% E. coli 0.1 ml Sterile Water 9.9 ml 9.89 ml 9.8 ml 8.9 ml Oregano Oil 0 ml 0.01 ml 1 ml Total Volume 10 ml

The tubes were allowed to incubate at room temperature for 15 minutes The tubes were allowed to incubate at room temperature for 15 minutes. The tubes were mixed by inversion during this time period to promote oregano oil and cell contacts. 0.1mL of the aliquots were spread onto the agar plates. 7. The plates were then incubated for 24 hours and the resulting colonies were counted.

Oregano Oil’s Effect on E. coli Alpha: 0.05 p-value: 8.54E-17

Dunnett's Test 0.1% 65.44 1% 67.45 10% 68.9 T-Critical= 3.5 Alpha= 0.05 Concentration T- Value Significance 0.1% 65.44 Significant 1% 67.45 10% 68.9

Conclusions Oregano Oil has a very significant negative effect on E. coli Oregano Oil does show signs of having antimicrobial activity All doses of the variable show significant effect Oregano Oil seems to reduce survivorship of microflora populations Oregano Oil might be able to combat E. coli related illnesses

Limitations and Extensions Vary exposures Vary concentrations Other cultures of bacteria More replicates Test with other spices Narrow the range of concentrations Isolate and test ingredients One exposure time Exposure to small amount of light Wide range of variable concentrations Multiple ingredients

Sources Minton, Barbara L. "Oil of Oregano: A Powerhouse for the Alternative Medicine Cabinet." NaturalNews. N.p., n.d. Web. 07 Jan. 2017. "Oregano Oil Benefits Superior To Prescription Antibiotics." Dr. Axe. N.p., n.d. Web. 07 Jan. 2017. http://www2.centralcatholichs.com/extracurricular%20science/PJAS/

ANOVA Anova: Single Factor SUMMARY Groups Count Sum Average Variance Column 1 4 1035 258.75 94.91667 Column 2 56 14 10 Column 3 26 6.5 5.666667 Column 4 3 0.75 0.916667 ANOVA Source of Variation SS df MS F P-value F crit Between Groups 190361.5 63453.83 2276.371 8.54E-17 3.490295 Within Groups 334.5 12 27.875 Total 190696 15  

Data Analysis: Single Factor ANOVA Alpha: 0.05 P-value was 8.54E-17 Interpretation: Null is rejected, significant effect Type of Effect: The Oregano Oil appeared to have killed E. coli colonies

Data Analysis: Dunnett's Test .1% Concentration: Significant Effect 1% Concentration: Significant Effect 10% Concentration: Significant Effect