Do More Expensive Hand
Sanitizers Inhibit
Bacterial Proliferation
Better Than Less
Expensive Ones?
By: Henry Microbe
Grade 7
Question / Problem
To determine which hand
sanitizer will inhibit the
proliferation of Escherichia coli
(E. coli ) for the lowest cost.
Background Research
A. Bacteria
1. Kingdom
a. Eubacteria
2. Prokaryotic Cells
3. Methods of Classifying Bacteria
a. Shape
1. Coccus - Round
2. Bacillus - Rod
3. Spirillum - Spiral
b. Gram Staining
1. Gram + = purple
2. Gram - = red
c. Clustering / Grouping
1. Diplo= two bacteria
2. Strepto= chain of bacteria
3. Staphylo= cluster of bacteria
4. E. coli
A.
B.
C.
D.
Bacillus bacteria
Gram Normally found in the intestines of animals
Diseases Caused by E. coli
1. Urinary Tract Infection
2. Diarrhea
3. Nausea
4. Possible Death
Hypothesis
As the price of the hand
sanitizer increases its
ability to inhibit bacterial
proliferation will increase.
Variables
Manipulated / Independent
The hand sanitizers being tested
Swan, Germ-x Aloe, Purell Adv.,
Germ-x Foaming, Germ-x
Responding / Dependent
The ability of the hand
sanitizers to inhibit
bacterial proliferation
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
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 Procedure
Step 1
Obtain
needed
materials
Step 8
Repeat
steps 6-7
using 3
more test
discs
Step 9
Repeat steps
6-8 using 2
more TSA
plates
prepared in
Step 2
Step 2
Divide 21
Tryptic Soy
agar (TSA)
plates into 4
equal size
quadrants
Step 7
Place the sterile
disc from Step 6 in
the center of one
of the quadrants
on one of the TSA
plates
Step 10
Repeat Steps 6
– 9 for peroxide
(+ Control) and
each of the
hand sanitizers
being tested
Step 3
Sterilize work
area with
Hydrogen
Peroxide
Step 6
Dip a sterile
disc into a
sterile saline
water
solution
(- Control)
Step 11
Incubate
all Petri
dishes for
72 hrs. at
37 º C
Step 4
Inoculate 50mL of
sterile nutrient broth
solution with E. coli &
incubate for 72 hours
at 37 º C
Step 5
Inoculate 21 Tryptic
Soy agar plates from
Step 2 with 1 mL of
the nutrient broth/E.
coli solution
prepared in step 4
Step 12
Determine 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
Optional Slide
Photo: A
Which Hand Sanitizer Inhibits Bacteria the Best for the Lowest Cost?
Average Zone of Inhibition (mm)
Purell Adv.
($1.13
/100mL)
Germ X
Foaming
($1.19
/100mL)
Germ X
($1.33/
100mL)
4.0
6.0
4.0
6.0
8.0
4.5
1.0
3.5
6.0
0.0
3.5
5.0
13.5
3.5
5.5
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
Disc
Tested
Hydrogen
Peroxide
(+ Control)
Sterile Saline
(- Control)
Swan Brand Germ X Aloe
($0.37
($0.55
/100 mL)
/100mL)
1
7.5
0.0
5.0
2
10.5
0.0
3
10.0
4
Which Hand Sanitizer Inhibits Bacteria the Best for
the Lowest Cost?
Average Zone of Inhibition (mm)
11
10
9.58
9
8
6.88
7
5.58
6
5.50
4.46
5
3.46
4
3
2
1
0
0.00
Hydrogen Sterile Saline Swan Brand Germ X Aloe
Peroxide
(- Control)
($0.37
($0.55
(+ Control)
/100 ml)
/100ml
Purell Adv.
($1.13
/100ml)
Germ
Foaming
($1.19
/100ml)
Type and Price of Hand Santizer Tested
Germ X
($1.33/100ml)
Two Sample Independent T-Test
Sterile Saline H O (- Control)
2
Significance Level = 0.05
Claim: µ1 = µ2
Confidence Level = 95%
Optional
Slide
- 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
Two Sample Independent T-Test
Hydrogen Peroxide (+ Control)
Significance Level = 0.05
Claim: µ1 = µ2
Confidence Level = 95%
Optional
Slide
+ 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
Conclusion
Based on the data I collected from my experiment I concluded
that the data partially supported my hypothesis.
Hypothesis
As the price of the hand sanitizer increases its ability
to inhibit bacterial proliferation will increase.
Actual Results
Hand Sanitizer
Tested
Cost/100 mL
Average Zone of
Inhibition (mm)
Swan Brand
Germ-x Aloe
Purell Advance
Germ-x Foam
Germ-x
$0.37
$0.55
$1.13
$1.19
$1.33
5.58
4.46
6.88
3.46
5.50
H2O2 (+ Control)
Saline (- Control)
$0.35
$0.22
9.58
0.00
Conclusion
Based 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.
Hypothesis
As the price of the hand sanitizer increases its ability to
inhibit bacterial proliferation will increase.
Significance Level = 0.05
Results
Confidence Level = 95%
Saline Vs Sanitizer
Hand Sanitizer
Tested
H2O2 Vs Samitizer
Cost/
(P Value) (Hypothesis) (P Value) (Hypothesis)
100mL
Swan Brand
Germ-x Aloe
Purell Advance
Germ-x Foam
Germ-x
$0.37
$0.55
$1.13
$1.19
$1.33
H2O2 (+Control)
Saline (-Control)
$0.35
$0.22
0.0023
0.0450
0.0031
0.0420
0.0380
Accepted
Accepted
Accepted
Accepted
Accepted
0.0510
0.0400
0.0148
0.0550
0.0410
Rejected
Accepted
Accepted
Rejected
Accepted
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 Procedure
Procedure:
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
Germ-x($1.33/100mL)
Ethyl Alcohol
Optional Slide
0.13%
63%
Sterile Saline Water (- Control)
Optional Slide
Photo: B