THE EFFECTS OF OLIVE OIL AGAINST CIGARETTE SMOKE

THE EFFECTS OF OLIVE OIL AGAINST CIGARETTE SMOKE ON YEAST Peter Wagstaff

PROBLEM Cigarette smoking is the leading cause of preventable death in the United States. Quitting is ideal but not always easy or expedient.

CIGARETTE SMOKE Cigarette smoke contains thousands of chemicals, many of them toxic or cancer causing, such as arsenic and tar An estimated 480, 000 deaths are caused by cigarettes every year in the United States. Cigarettes are the leading cause of preventable death in the United States

OLIVE OIL Contains Oleic Acid Unsaturated fats are cited as a lung supporting nutrient by the National Lung Association Previous studies have reported that diets high in unsaturated fats can shield against some effects of smoking However, other studies have indicated that the burning of cooking oils can increase risk for lung cancer

WHY YEAST (SACCHAROMYCES CERERISAE) Yeast is used as a model because: Eukaryotic, just like human cells Most studied cell Many characteristics like nutrition and cell growth are similar to human cells Aerobic

PURPOSE Can unsaturated fats (olive oil) reduce the negative effects of cigarette smoke on yeast? Will oil alone affect yeast survivorship? Will smoke alone affect yeast survivorship?

HYPOTHESIS Null: The olive oil and cigarette smoke will NOT have a significant effect on yeast survivorship, individually or synergistically. Alternative 1: The presence of olive oil or cigarette smoke will have a significant individual effect on yeast survivorship. Alternative 2: Olive oil will significantly increase the survivorship of smoke stressed yeast.

MATERIALS Saccharomyces cerevisiae (Yeast) YEPD agar plates (1% yeast extract, 1. 5% agar, 2% glucose) Aldi brand virgin olive oil Sterile micropipettes and tips (200 ml) Sharpie Sterile dilution fluid (100 m. M KH 2 PO 4. 100 m. M K 2 HPO 4, 10 m. M Mg. SO 4, 1 m. M Na. Cl) Sterile spreader bars Ethyl alcohol Burner Marlboro cigarettes Stop watch Smoke Chamber Incubator Spectrophotometer

PROCEDURE 1. 2. 3. A culture Saccharomyces cerevisiae was grown. The culture was incubated at 30 degrees Celsius until a density of 50 Klett spectrophotometer units were reached. This represents a density of approximately 10⁷ cells/m. L. The culture was diluted with sterile dilution fluid to a concentration of 10^3 cells/m. L

PROCEDURE 4. 100μl of the cell suspension was pipetted into olive oil infused YEPD agar plates Agar infusion: After autoclaving YEPD agar, sterile olive oil was mixed with the molten agar to create the following infused plates; 0%, 0. 1%, and 10% 5. Six plates of each concentration were taped to the top of the smoke chamber and exposed to 0, 45, 90, and 130 seconds of smoke

PROCEDURE 7. A fresh cigarette was lit and fitted into the socket for each exposure, and the air pump was pumped every three seconds for consistent airflow. 8. The plates were covered and incubated at 30 degrees Celsius for 72 hours and the resulting colonies were quantified.

DOCUMENTATION OF SMOKE CHAMBER

DOCUMENTATION OF SMOKE CHAMBER

DOCUMENTATION OF SMOKE CHAMBER

ANOVA Single Factor � Stat test comparing the means of multiple groups � Alpha of 0. 05 is selected Two Factor � Can reveal synergistic affects of two variables

SMOKE EFFECTS ON YEAST SURVIVORSHIP Number of colonies Average Yeast a Survivorship 90 80 Overall P-value: 0. 00171 70 60 50 Yeast without oil 40 30 20 10 0 0 Smoke Exposure [seconds] 45 90 130

OIL EFFECT ON YEAST SURVIVORSHIP Number of colonies Average Yeast Survivorship 120 Overall P-value: 4. 24 E-06 100 80 60 Yeast with out smoke 40 20 0 0% Concentration[Oil] 0% 1% 10%

DUNNETT’S TEST The Dunnett's test is a follow up to the ANOVA that quantifies the difference between a variable and control group to find significant variation.

DUNNETT'S TEST Group T Value Significance 0. 1% Oil 2. 432 Not Significant 1% Oil 0. 024 Not Significant 10% Oil -4. 936 Significant Oil did not begin to have a significant effect on yeast until the concentration reached 10% T-Crit = 3. 098391

SMOKE AND OIL EFFECTS ON STRESSED YEAST Number of colonies Average Yeast Survivorship 120 Overall P-value: 0. 017711 100 80 0% Oil 60 0. 10% 1% 40 10% 20 0 0 Seconds Exposure 45 90 130

Did olive oil have an effect by itself? P-Value 4. 24 E-6 Significant Did cigarette smoke reduce yeast survivorship? P-Value 0. 00171 Significant Did oil increase survivorship in smoke stressed yeast? P-Value 0. 017711 Significant

CONCLUSION The null hypothesis was rejected. Both variables had individual effects as well as a synergistic effect. Both oil and smoke harmed the yeast, however oil created a shielding affect at high concentrations on smoke stressed yeast.

LIMITATIONS/EXTENSIONS Limitations Lag time between smoke exposures could have created unwanted variance Yeast should have been exposed more immediately after plating Extensions More replicates More exposure times and concentrations of oil A more sophisticated smoke chamber

REFERENCES [http: //atvb. ahajournals. org/content/21/6/1029. long] [http: //www. ncbi. nlm. nih. gov/pubmed/10618003] [http: //cancergrace. org/lung/2007/11/27/cooking-oilfumes-and-lcins/]

SINGLE FACTOR ANOVA FORSMOKE EFFECTS Groups Count Sum Average Variance Column 1 6 481 80. 16667 47. 76667 Column 2 6 455 75. 83333 52. 56667 Column 3 6 425 70. 83333 42. 56667 Column 4 6 381 63. 5 25. 5 ANOVA Source of Variation SS df MS F Between Groups 921. 8333 3 307. 2778 7. 29876 Within Groups 842 20 42. 1 1763. 833 23 Total P-value F crit 0. 00171 3. 098391

SINGLE FACTOR ANOVA FOROLIVE OIL EFFECTS Groups Count Sum Average Variance Column 1 6 481 80. 16667 47. 76667 Column 2 6 581 96. 83333 282. 5667 Column 3 6 482 80. 33333 223. 4667 Column 4 6 278 46. 33333 9. 866667 ANOVA Source of Variation SS Between Groups 8101. 5 3 2700. 5 Within Groups 2818. 333 20 140. 9167 Total 10919. 83 23 df MS F P-value 19. 16381 F crit 4. 24 E-06 3. 098391

2 FACTOR ANOVA SUMMARY Source of Variation SS df MS F 63. 92777 P-value F crit Sample 14810. 86 3 4936. 955 3. 46 E-21 2. 718785 Columns 1598. 615 3 532. 8715 6. 90006 0. 000343 2. 718785 Interaction 1677. 594 9 186. 3993 2. 413652 0. 017711 1. 999115 Within 6178. 167 80 77. 22708