1. Disinfectant Wipe Extract Effects on Bacteria
Ethan Williams
Grade 9
Central Catholic High School

2. Rationale
Any disinfectant wipe should be able to significantly reduce survivorship of microbes
The more expensive Clorox wipes should be more effective in reducing survivorship so that they can make up for the price difference

3. Purpose
To determine if Clorox and Great Value wipes greatly reduce survivorship of gram positive and gram negative bacteria
If so, which is more effective and at what concentration?

4. Clorox Wipes
Higher-end, more expensive - $4.99 per canister – 78 wipes per character
Active Ingredient - Dimethyl Benzyl Ammonium Chloride
Common Household chemical used to clean surfaces
Clorox claims that these wipes will kill 99.9% of both Staph and E. coli on hard surfaces

5. Great Value Wipes
Lower-end wipes, cheaper - $1.99 per canister- 75 wipes per canister
Active Ingredient – Octyl Decyl Dimethyl Ammonium Chloride
Also used commonly in cleaning products
Claim that it kills 99.9% of bacteria

6. Comparison of Inactive Ingredients
Clorox
Great Value
Water
Substrate
Hexoxythenol
Citric Acid-Lemon scented
Fragrance
d-Limonene
Isopropanol
Sodium Bicarbonate
Water
Substrate
Citric Acid-Lemon scented
Fragrance

7. Escherichia coli (E. coli)
Gram (-)
Commonly found in the human gut
Most strains are not pathogenic
Most widely used bacteria model (represents other gram (-) bacteria)

8. Staphylococcus epidermidis (Staph)
Gram (+)
Normal human flora - typically skin flora
Can cause staph infections when it enters the body
Can cause staph food poisoning

9. Hypotheses
Null: Neither the Clorox nor the Great Value Wipes will have an effect on the survivorship of the bacteria
Alternative: The Clorox and Great Value wipes will significantly reduce survivorship of the E. coli and Staph
Alternative #2: The Clorox wipes will reduce survivorship more than Great Value

10. Materials 250 mL beaker 500 gram weight
Spreader bars
Ethanol
Sterile Water
Test Tubes + Rack
SDF (10 mM KH2PO4, 10 mM K2HPO4,1 mM MgSO4, 0.1 mM CaCl2, 100 mM NaCl)
Incubator
Forceps
Hole puncher
Clorox brand disinfectant wipes
Great Value brand disinfectant wipes
E. coli
Staph
YEPD Plates (0.5% yeast extract, 2% peptone, glucose
Micropipettes
Sterile Pipette Tips
Test Tubes
Sterile Tubes

11. Procedure: Stock Solution
45 mL of sterile water was placed in a sterile 50 mL conical tube
A wipe was placed in the tube for 5 seconds
The tube was inverted 10 times to ensure that the solution was evenly spread in the water
The wipe was removed with sterile forceps

12. Procedure: Liquid Pulse (Cont.)
0.1 mL of each concentration was pipetted onto 5 plates per concentration per brand resulting in 20 plates per brand of E. coli and 20 plates of staph per brand
The solutions were spread onto the plates with sterile spreader bars
The E. coli plates were incubated for 1 day at 37°C
The staph plates were incubated for 2 days at 37°C to allow for sufficient growth

13. Procedure: Liquid Pulse
E. coli and Staph were grown at a density of 10ˆ⁸ cells per mil
They were then diluted in SDF to a density of 10ˆ⁵ cells per mil
The following ingredients were mixed to create the following exposures
The solutions were incubated at room temperature for 10 minutes 0%
0.1% 1%
10%
SDF
9.9 mL
9.89 mL
9.8 mL
8.9 mL
Wipe Extract
0 mL
0.01 mL
0.1 mL
1 mL
Bacteria
Total Volume
10 mL

14. Procedure: Sterile Disk Assay
0.1 mL of bacteria was plated with a sterile spreader bar
A hole punch was used to create sterile disks from the wipe
4 sterile disks were placed in a cross shape on each plate
The plates were incubated for 1 day at 37°C for E. coli and 2 days at 37°C for staph
After incubation, the zone of inhibition was measured (mm)

15. Clorox vs Great Value E. coli Survivorship

16. Dunnet’s Test Evaluation
T Value
T crit
Significant/ Not Significant
Clorox 0.1%
10.98
2.77
Sig
Clorox 1%
17.86
Clorox 10%
24.23
Great Value 0.1%
9.97
Great Value 1%
15.21
Great Value 10%
22.23

17. Clorox v. Great Value Staph Survivorship

18. Dunnet’s Test Evaluation
T Value
T-crit
Significant/ Not Significant
Clorox 0.1%
1.95
2.81
Not Sig
Clorox 1%
26.24
Sig
Clorox 10%
39.67
Great Value 0.1%
11.92
Great Value 1%
19.87
Great Value 10%
25.05

19. E. coli Zone Inhibition

20. Staph Zone Inhibition

21. Conclusions
The null can be rejected, as the effects on the bacterial survivorship were very significant, and the p-values were far below the alpha of .05
The alternative hypothesis was supported, as the Clorox wipes had far less bacterial survivorship than the Great Value wipes

22. Limitations and Extensions
Test more brands of wipes
More replicates for each concentration
Use a lower concentration than 10% as the high, possibly 5%
Use different types of bacteria
Limitations:
Possible error in pipette technique
Possible error in spreading on the plates
Possible error in counting of colonies
Only 5 replicates were used
Study does not reveal other effects on bacterial health

23. Works Cited
“Ammonium Chloride.” National Center for Biotechnology Information. PubChem Compound Database, U.S. National Library of Medicine, pubchem.ncbi.nlm.nih.gov/compound/ammonium_chloride#section=Top.
“E. Coli (Escherichia Coli) | E. Coli | CDC.” Centers for Disease Control and Prevention, Centers for Disease Control and Prevention,
Foodsafety.gov. “E. Coli.” FoodSafety.gov, U.S. Department of Health and Human Services, 24 Aug ,
“Gateway on Pesticide Hazards and Safe Pest Management.” Beyond Pesticides,
“Healthcare-Associated Infections.” Centers for Disease Control and Prevention, Centers for Disease Control and Prevention, 17 Jan. 2011,
“Ingredients Inside.” The Clorox Company, 9 Dec. 2016, were-made-of/ingredients-inside/clorox/clorox-disinfecting-wipes1-crisp-lemon /.
“Staphylococcal (Staph) Food Poisoning | Food Safety | CDC.” Centers for Disease Control and Prevention, Centers for Disease Control and Prevention,

24. Raw Data- Clorox E. coli SUMMARY Groups Count Sum Average Variance 0%5
1470
294
509
0.10%
697
139.4
662.8 1%
471
94.2
187.7
10%
ANOVA
Source of Variation SS df MS F
P-value
F crit
Between Groups 3
3.0948E-13
Within Groups
5438 16
Total 19

25. Raw Data- Great Value E. Coli
SUMMARY
Groups
Count
Sum
Average
Variance 0% 5
2943
588.6
340.3
0.10%
2084
416.8
339.7 1%
1433
286.6
387.3
10% 60 12
139.5
ANOVA
Source of Variation SS df MS F
P-value
F crit
Between Groups 3
E-18
Within Groups
4827.2 16
301.7
Total
891586 19

26. Raw Data – Clorox Staph SUMMARY Groups Count Sum Average Variance 0% 5
1447
289.4
449.3
0.10%
719
143.8
328.7 1%
511
102.2
10% 1
0.2
ANOVA
Source of Variation SS df MS F
P-value
F crit
Between Groups 3
E-14
Within Groups
4427.6 16
Total 19

27. Raw Data – Great Value Staph
SUMMARY
Groups
Count
Sum
Average
Variance 0% 5
2401
480.2
226.7
0.10%
1722
344.4
332.8 1%
941
188.2
482.2
10% 56
11.2
84.2
ANOVA
Source of Variation SS df MS F
P-value
F crit
Between Groups 3
E-17
Within Groups
4503.6 16
Total
617524 19