David McFall
How caffeine
interacts with the
body
The binding of the
hormone adenosine to an
adenosine receptor in the
brain brings about sleep.
Caffeine blocks adenosine
from being connected to
the adenosine receptor.
The hormone epinephrine
is then produced. This
creates the notable effects
of caffeine.
Coffee
•Coffee has a high
caffeine content due
to its high
concentration in its
endosperm.
•Coffee can be
decaffeinated by a
treatment of
methylene chloride.
The Effects of Coffee
Coffee’s side effects are
drawn from the release of the
hormone epinephrine, shown
here.
The most notable effects
include:
high heart beat
high blood pressure
 higher blood sugar
content
 tense muscles
Epinephrine
Starbucks X-bold Sumatra Roast (Coffee A)
20 mg of Caffeine per ounce
Starbucks Decaf House Blend (Coffee B)
0.6 to 1.4 mg of Caffeine per ounce
Escherichia coli
The model for this
experiment is
Escherichia coli (E.
coli).
It is found in the
intestinal tract of most
animals, including
humans.
Most importantly, E.
coli has been tested in
conjunction with
caffeine before in
previous studies.
A
study from the Lithuanian State Science and Studies
Foundation concluded that when in high concentration, caffeine
lowers the population of E. coli.
Another study from the Indiana University School of Medicine
supported the previous study.
To determine the effect of coffee on human coli form
populations.
Null Hypothesis: Neither caffeinated nor decaffeinated coffee will have an effect on the
population of E. coli at any concentration.
Alternative Hypothesis: Caffeinated and decaffeinated coffee will significantly effect
survivorship of E. coli
E. coli
Permanent Marker
Latex Gloves
Micropipettes
Microtubes
Caffeinated Coffee
Decaffeinated Coffee
Sterile Distillation Fluid
58 LB agar plates (Per Liter:1% Tryptone, 0.5% Yeast Extract, and 1% NaCl)
LB media
Klett Spectrophotometer
0.22 micron syringe filters
2 Sterile Filters
Bunsen Burners
Spread Bar
Incubator
Ethanol
1. E. coli was grown overnight in sterile LB media.
2. A sample of the overnight culture was added to fresh media in a sterile sidearm flask.
3. The culture was placed in an incubator (37°C) until a density of 50 Klett
spectrophotometer units was reached. This represents a cell density of approximately
108 cells/mL.
4. The culture was diluted in sterile dilution fluid to a concentration of approximately 105
cells/mL.
5. Caffeinated and decaffeinated coffee was mixed with the appropriate amount of
SDFto create coffee concentrations of 10%, 1%, and 0.1%.
6. 100 µL of cell culture was then added to the solutions, yielding a final volume of 10 mL
and a cell density of approximately 103 cells/mL.
7. The solutions were vortexed and allowed to sit at room temperature for 15 minutes.
8. 100 µL aliquots were removed from the tubes and spread on LB plates.
9. The plates were incubated at 37 degrees for 24 hours.
10. The resulting colonies were counted visually. Each colony was assumed to have arisen
from one cell.
11. The appropriate statistical analyses were performed to adequately assess the
hypothesis.
1.
2.
3.
4.
Repeat steps 1-6 in the Preliminary Procedure.
Both brands of coffee were brewed as directed.
The coffee was then sterile filtered.
200 µL of sterile coffee were removed from the tubes and
spread on LB plates.
5. The plates were incubated at 37 degrees for 24 hours.
6. 100uL of bacteria (control tube) was spread onto each plate.
7. The resulting colonies were counted visually. Each colony
was assumed to have arisen from one cell.
8. The appropriate statistical analyses were performed to
adequately assess the hypothesis.
Chart of Concentration
Table 1
Control (x2) .01x (x4)
.1x (x4)
x (x4)
E. coli
.1 ml
.1 ml
.1 ml
.1 ml
Sterile Distillation
Fluid
9.9 ml
9.89 ml
9.8 ml
8.9 ml
Coffee
0 ml
.01 ml
.1 ml
1 ml
Total Volume
10 ml
10 ml
10 ml
10 ml
Average Effect of Coffee on E. coli
700
Colonies of E. coli
600
P<0.05
500
P<0.05
P<0.05
400
300
P>0.05
P>0.05
P<0.05
P>0.05
Control
Decaffeinated
200
Caffeinated
Infused Plates
100
0
Concentrations
T Critical= 3.49 (Significant)
Variable
Concentration
.01x CAFF
.1x CAFF
x CAFF
.01x DCAFF
.1x DCAFF
x DCAFF
T Value
2.2
3.46
4.78
7.63
4.54
3.69
Interpretation
Insignificant
Insignificant
Significant
Significant
Significant
Significant
 Caffeinated coffee significantly increased the
population of E. coli, but only in the 10%
concentration.
Decaffeinated coffee significantly increased the
population of E. coli in all concentrations.
Test more brands of coffee
Use a different model
Test coffee in conjunction with other caffeinated
beverages, such as tea or energy drinks
Plating
was not exactly synchronized, which
could have resulted in extra time for bacterial
replication. A team of students could remedy
this technical problem.