Do More Expensive Hand Sanitizers Inhibit

Bacterial Proliferation Better Than Less Expensive Ones?

By: Henry MicrobeGrade 7

Question / Problem

To determine which hand sanitizer will inhibit the proliferation of Escherichia coli (E. coli ) for the lowest cost.

Background ResearchA. Bacteria

1. Kingdom a. Eubacteria2. Prokaryotic Cells3. Methods of Classifying Bacteria

a. Shape 1. Coccus - Round

2. Bacillus - Rod 3. Spirillum - Spiral

b. Gram Staining 1. Gram + = purple2. Gram - = red

c. Clustering / Grouping 1. Diplo= two bacteria2. Strepto= chain of bacteria3. Staphylo= cluster of bacteria

4. E. coli A. Bacillus bacteria B. Gram - C. Normally found in the intestines of animals D. Diseases Caused by E. coli 1. Urinary Tract Infection 2. Diarrhea 3. Nausea 4. Possible Death

HypothesisAs the price of the hand sanitizer increases its ability to inhibit bacterial proliferation will increase.

VariablesResponding / Dependent

Constants

•1- Pure slant of E. coli•21- Sterile Tryptic Soy Agar (TSA) plates•84 - Sterile blank discs •Incubated at 37° C •Incubator used •Method of determining the average zone of inhibition

Nuisance

• Minor fluctuations in temperature and humidity in the incubator•Age & viability of the E. coli•Irregular zones of inhibition

The hand sanitizers being tested

Swan, Germ-x Aloe, Purell Adv., Germ-x Foaming, Germ-x

The ability of the hand sanitizers to inhibit bacterial proliferation

Manipulated / Independent

Materials Used• Pen• Paper• Incubator• 21 Sterile Tryptic Soy Agar (TSA) Petri Dishes• 84 Sterile Blank Discs • Sterile Saline Water (H20)• Hydrogen Peroxide (H202)• Sterile Latex Gloves• Sterile Tweezers• Sterile Beakers• Black Sharpie Marker• .22µm Millex Millipore Filter• Goggles• Sterile Nutrient Broth• Hand Sanitizers Tested: Swan Brand $0.37/100mL Germ-x Aloe $0.55/100mL Purell Advance $1.13/100mL Germ-x Foam $1.19/100mL Germ-x $1.33/100mL

Experimental ProcedureStep 1

Obtain needed materials

Step 3Sterilize work area with Hydrogen Peroxide

Step 4 Inoculate 50mL of sterile nutrient broth solution with E. coli & incubate for 72 hours at 37 º C

Step 5Inoculate 21 Tryptic Soy agar plates from Step 2 with 1 mL of the nutrient broth/E. coli solution prepared in step 4

Step 6Dip a sterile disc into a sterile saline water solution(- Control)

Step 7 Place the sterile disc from Step 6 in the center of one of the quadrants on one of the TSA plates

Step 2Divide 21 Tryptic Soy agar (TSA) plates into 4 equal size quadrants

Step 8Repeat steps 6-7 using 3 more test discs

Step 9 Repeat steps 6-8 using 2 more TSA plates prepared in Step 2

Step 10Repeat Steps 6 – 9 for peroxide (+ Control) and each of the hand sanitizers being tested

Step 11Incubate all Petri dishes for 72 hrs. at 37 º C

Step 12Determine and record the average zone of inhibition for each of the sterile discs dipped in the solutions being tested

Determining the Average Zone of Inhibition for Hydrogen Peroxide (+

Control)

Sterile Disc Dipped in H₂O₂

Photo: A

Optional Slide

Which Hand Sanitizer Inhibits Bacteria the Best for the Lowest Cost?

  Average Zone of Inhibition (mm)

Disc Tested Hydrogen

Peroxide (+ Control)

Sterile Saline (- Control)

Swan Brand ($0.37

/100 mL)

Germ X Aloe

($0.55 /100mL)

Purell Adv. ($1.13 /100mL)

Germ X Foaming ($1.19 /100mL)

Germ X ($1.33/ 100mL)

1 7.5 0.0 5.0 4.0 6.0 4.0 6.0

2 10.5 0.0 8.0 4.5 1.0 3.5 6.0

3 10.0 0.0 3.5 5.0 13.5 3.5 5.5

4 11.0 0.0 6.5 4.5 11.0 3.5 4.5

5 10.5 0.0 8.5 5.0 7.0 3.5 4.5

6 6.0 0.0 10.0 5.0 7.5 4.0 4.0

7 9.0 0.0 1.0 1.0 7.0 3.5 5.0

8 10.0 0.0 8.0 5.0 4.5 4.0 5.0

9 10.5 0.0 1.0 4.5 5.5 3.5 4.5

10 10.5 0.0 4.0 4.5 6.0 4.0 5.0

11 10.0 0.0 5.5 5.0 4.5 3.5 5.0

12 9.5 0.0 6.0 5.5 9.0 1.0 6.0

Average 9.58 0.00 5.58 4.46 6.88 3.46 5.50

Hydro

gen

Pero

xide

(+

Con

trol)

Ster

ile S

aline

(- Con

trol)

Swan

Bra

nd

($0.

37

/

100

ml)

Germ

X A

loe

($0.

55

/100

ml

Pure

ll Adv

. ($1

.13

/1

00m

l)

Germ

Foa

ming

($1.

19

/100

ml)

Germ

X (

$1.3

3/10

0ml)

0123456789

1011 9.58

0.00

5.584.46

6.88

3.46

5.50

Which Hand Sanitizer Inhibits Bacteria the Best for the Lowest Cost?

Type and Price of Hand Santizer Tested

Avera

ge Z

on

e o

f In

hib

itio

n (

mm

)

Two Sample Independent T-Test Sterile Saline H

2O (- Control)

Significance Level = 0.05Claim: µ1 = µ2

Confidence Level = 95%

- Control vs. P-value Result

Swan Brand ($0.37 / 100 mL) 0.0023

Hypothesis Accepted

Germ X Aloe ($0.55 / 100 mL) 0.0450

Hypothesis Accepted

Purell Adv. ($1.13 / 100 mL) 0.0031

Hypothesis Accepted

Germ Foaming ($1.19 / 100 mL) 0.0420

Hypothesis Accepted

Germ X ($1.33 / 100 mL) 0.038

Hypothesis Accepted

Optional Slide

Two Sample Independent T-Test Hydrogen Peroxide (+ Control)

Significance Level = 0.05Claim: µ1 = µ2

Confidence Level = 95%

+ Control vs. P-value Result

Swan Brand ($0.37 / 100 mL) 0.0510

Hypothesis Rejected

Germ X Aloe ($0.55 / 100 mL) 0.0400

Hypothesis Accepted

Purell Adv. ($1.13 / 100 mL) 0.0148

Hypothesis Accepted

Germ Foaming ($1.19 / 100 mL) 0.0550

Hypothesis Rejected

Germ X ($1.33 / 100 mL) 0.0410

Hypothesis Accepted

Optional Slide

Based on the data I collected from my experiment I concluded that the data partially supported my hypothesis.

As the price of the hand sanitizer increases its ability to inhibit bacterial proliferation will increase.

Hand Sanitizer Cost/100 mL Average Zone of Tested Inhibition (mm)

Swan Brand $0.37 5.58 Germ-x Aloe $0.55 4.46 Purell Advance $1.13 6.88 Germ-x Foam $1.19 3.46 Germ-x $1.33 5.50

Conclusion

Actual Results

Hypothesis

H2O2 (+ Control) $0.35 9.58 Saline (- Control) $0.22 0.00

ConclusionBased on the Two Sample Independent T-Test that I performed on the data that I collected from my experiment I concluded that the data partially supported my hypothesis.

HypothesisAs the price of the hand sanitizer increases its ability to inhibit bacterial proliferation will increase.

Results

Hand Sanitizer Cost/ (P Value) (Hypothesis) (P Value) (Hypothesis) Tested 100mL

Swan Brand $0.37 0.0023 Accepted 0.0510 RejectedGerm-x Aloe $0.55 0.0450 Accepted 0.0400 AcceptedPurell Advance $1.13 0.0031 Accepted 0.0148 AcceptedGerm-x Foam $1.19 0.0420 Accepted 0.0550 RejectedGerm-x $1.33 0.0380 Accepted 0.0410 Accepted

H2O2 (+Control) $0.35 Saline (-Control) $0.22

Confidence Level = 95%

Saline Vs Sanitizer

Significance Level = 0.05

H2O2 Vs Samitizer

Conclusion Based on Statistical Analysis Optional Slide

Future Improvements / Changes

• Test more discs for each of the hand sanitizers being tested. (≥ 30 discs)

• Perform more trials (3 – 10 trials)

• Use a different protocol for determining the ability of hand sanitizers to inhibit bacterial proliferation.

• Purchase a greater number of hand sanitizers

Future Experiments / Extensions

• Design an experiment in order to determine the effect of hand sanitizers on a variety of different types of bacteria.

• Perform an experiment in order to determine the ability of natural herbs and spices to inhibit bacterial proliferation.

• Design and perform an experiment in order to determine whether bacteria will develop a resistance to various antibacterial chemicals.

• Perform an experiment in order to test the ability of different types of anti bacterial hand soaps to inhibit bacterial proliferation.

• Design and perform a similar experiment with hand soap

Real Life Applications

• The results obtained from my experiment could be used by cafeterias, restaurants, and hospitals to purchase the least expensive but most effective hand sanitizer.

• The results obtained from my experiment could assist my family to purchase the least expensive and most effective hand sanitizer for home use.

• This information could assist retailers in determining the most effective hand sanitizer to purchase for the least cost.

Bibliography of Sources "Clinical Infectious Diseases." Consumer Antibacterial Soaps: Effective or Just Risky? N.p., n.d. Web. 05 Dec. 2013.

"On the Structure of Eukaryotic, Prokaryotic, and Viral Chromatin." On the Structure of Eukaryotic, Prokaryotic, and Viral Chromatin. N.p., n.d. Web. 05 Dec. 2013.

"Zone of Inhibition." - Definition from Biology-Online.org. N.p., n.d. Web. 04 Feb. 2014.

"10-2 The Antibiotic Disk Sensitivity Test." Microbiology Labs. N.p., n.d. Web. 05 Feb. 2014.

"Escherichia Coli." Wikipedia. Wikimedia Foundation, 27 Jan. 2014. Web. 05 Feb. 2014.

"Wash Your Hands." Centers for Disease Control and Prevention. Centers for Disease Control and Prevention, 11 Dec. 2013. Web. 03 Feb. 2014.

Gram Staining ProcedureProcedure:

A. Slant Cultures

 1. Prepare and heat-fix smears.  2. Prepare the smears of S. epidermidis and N. sicca on a second slide.  Heat-fix.  3. Stain the slides as follows:       a. Flood the crystal violet for one minute.       b. Pour off excess dye and wash gently in tap water and drain the slide against a paper towel.       c. Expose the smears to Gram's iodine for one minute by washing with iodine, then adding more iodine and leaving it

on the smear until the minute is over.    d. Wash with tap water and drain carefully.  (Do not blot.)    e. Wash with 95% alcohol for 30 seconds.    f.  Wash with tap water at the end of the 30 seconds to stop the decolorization.  Drain.   g. Counterstain with 0.25% safranin for 30 seconds.   h. Wash, drain, blot, and examine under oil.   i. Draw the cells showing morphology, grouping, and relative

sizes.  Color a few of the cells of each bacterial species to show the Gram reaction.     j. Save these slides and the ones from parts B & C of this exercise to use at the next lab period.  

Optional Slide

Active Ingredients

Hand Sanitizer Active Ingredient Percentage

Swan Brand($0.37/100mL) Ethyl Alcohol 65% Germ-x Aloe($0.55/100mL) Ethyl Alcohol 62%

Purell Advance($1.13/100mL) Ethyl Alcohol 70%

Germ-x Foaming($1.19/100mL) Benzalkonium Chloride 0.13%

Germ-x($1.33/100mL) Ethyl Alcohol 63%

Optional Slide

Sterile Saline Water (- Control)

Optional Slide Photo: B