Combustion Products
Effects on Yeast Population
Growth
Jack Leech
Pittsburgh Central Catholic High School
Grade 9
Car exhaust
• Car exhaust was used because it is a very
common source of gaseous pollution, and
contains elements that are known to cause
health problems.
• The internal combustion engine gives off
carbon monoxide, carbon dioxide, oxides
of nitrogen, hydrocarbons, sulfur dioxide,
and micro-particles.
Cigarette Smoke
• Cigarette smoke also has harmful effects
through all of the thousands of poisons it
releases when burned.
• The main pollutants are acetone, carbon
monoxide, arsenic, benzene, butane,
cyanide, nicotine, lead, formaldehyde, and
many others.
Charcoal Smoke
• Charcoal smoke was used because it is
also a common fuel source.
• Burned charcoal creates hydrocarbons
and soot particles. In addition, the
charcoal used in the experiment was
soaked in kerosene, which when burned
produces carbon monoxide.
Background Information
• The smoke sources were chosen because
they are known to have an effect on the
environment, and now the effect on living
cells would be tested.
Background Information (cont.)
• The cells that would be tested on was a
species of yeast known as
Saccharomyces cerevisiae. This cell was
used because it is very similar in its
structure to the human cell, and so the
results that would be drawn could be
associated with human cells.
Purpose:
• To find out if common combustion
products have an effect on the growth of
yeast colonies.
• If smoke does have an effect on yeast,
which specific smoke source has the
greatest effect on yeast growth; car
exhaust, cigarette smoke, or charcoal
smoke.
Hypothesis
• Null- the smoke sources will have no
significant effect on yeast survivorship.
• Alternative- based on the excessive
amounts of toxins, smoke will have a
significant negative effect in the
destruction of yeast populations.
Materials
• 48 YEPD agar plates (1%
yeast extract, 2% peptone,
2% glucose (dextrose), 1.5%
agar)
• YEPD media (1% yeast
extract, 2% peptone, 2%
glucose (dextrose))
• Sterile capped test tube with
Sterile Dilution Fluid (SDF)
(10 mM KH2PO4, 10 mM
K2HPO4, 1 mM MgSO4, 0.1
mM CaCl2, 100 mM NaCl)
• Cigarettes
• Charcoal
• Access to car for exhaust
(car used was a 2003 Honda
CR-V)
• Butane lighter
• Shoebox with a hole cut in
the side (with plates in box,
attach to exhaust pipe)
• Micropipette
• Permanent marker
• Aluminum foil
• Plate spreader
• Ethanol (used to sterilize
plate spreader)
Procedure (Indirect exposure)
• S.c. Yeast was grown overnight in sterile YEPD Media.
• A sample of the overnight culture was added to fresh
media in a sterile sidearm flask.
• The culture was placed in a shaking water bath (30 C)
until a density of 50 Klett spectrophotometer units was
reached. This represents a cell density of approximately
107 cells/ml.
• An aluminum foil chimney was created and placed over
the smoking sources for intervals of 1 minute and 2
minutes.
• After vortexing to evenly suspend cells, 0.1 ml. aliquots
were removed from the tubes and spread on 24 plates
• The cell culture was diluted in sterile dilution fluid to a
concentration of approximately 103 cells/ml.
• The plates were incubated at 30 C for 48 hours.
• The resulting colonies were counted. Each colony is
assumed to have arisen from one cell.
Procedure (Direct exposure)
• S.c. Yeast was grown overnight in sterile YEPD Media.
• A sample of the overnight culture was added to fresh
YEPD media in a sterile sidearm flask.
• The culture was placed in a shaking water bath (30 C)
until a density of 50 Klett spectrophotometer units was
reached. This represents a cell density of approximately
107 cells/ml.
• The cell culture was diluted in sterile dilution fluid to a
concentration of approximately 103 cells/ml.
• After vortexing to evenly suspend cells, 0.1 ml. aliquots
were removed from the tubes and spread on 24 plates
• An aluminum foil chimney was created and placed over
smoking sources for 1 minute and 2 minutes. The plates
were incubated at 30 C for 48 hours.
• The resulting colonies were counted. Each colony is
assumed to have arisen from one cell.
Indirect Exposure
P=0.2
.2
.06
1 Minute
Exposure
.09
.003
.04
Car
Exhaust
Charcoal
.24
Resulting
Colonies
350
300
250
200
150
100
50
0
P=0.02
2 Minute
Exposure
Time Exposed to Smoke Sources
Cigarette
Smoke
Control
Direct Exposures
400
P=2.6*10-5
Car
Exhaust
Charcoal
300
.0005
.006
.0001
100
.003
200
.002
.0006
Resulting
Colonies
P=0.0003
0
1 Minute
Exposure
2 Minute
Exposure
Time Exposed to Smoke Sources
Cigarette
Smoke
Control
Indirect 2 min. exposure Indirect 1 Min. exposure
Smoke source
Results
Car exhaust
P-value (.24) > than .05 (Insufficient data to
have effect on control)
Charcoal Smoke
P-value (.04) < than .05 (Significant effect
on control)
Cigarette Smoke
P-value (.003) < than .05 (Significant effect
on control)
Car exhaust
P-value (.09) > than .05 (Insufficient data to
have effect on control)
Charcoal Smoke
P-value (.06) > than .05 (Insufficient data to
have effect on control)
Cigarette Smoke
P-value (.2) > than .05 (Insufficient data to
have effect on control)
Direct 1 Min. exposure
Direct 2 min. exposure
Smoke source
Results
Car exhaust
P-value (.002) < than .05 (Significant effect
on control)
Charcoal Smoke
P-value (.0006) < than .05 (Significant
effect on control)
Cigarette Smoke
P-value (.003) < than .05 (Significant effect
on control)
Car exhaust
P-value (.0005) < than .05 (Significant
effect on control)
Charcoal Smoke
P-value (.006) < than .05 (Significant effect
on control)
Cigarette Smoke
P-value (.0001) < than .05 (Significant
effect on control)
Interpretation
• After reviewing results, a trend was
discovered.
• The direct exposure had a much more
destructing effect on the yeast.
• In addition, the time exposed had little to
do with the yeast destruction, but instead
the exposure was what determined the
survivorship.
Conclusions
• After reviewing results, the null hypothesis was
partially rejected. There was sufficient evidence to
see that the direct exposure smoke did have a
negative effect on the yeast cells, but the indirect
was not as effective.
• Based on the p-values of the smokes compared to
the control, the conclusions were that cigarette
smoke is the most effective smoke out of the others
in destroying yeast cells.
• In addition, although the indirect exposure did have
an effect it was not enough to prove that the smoke
was effecting the yeast cells
• The most effective form of cell destruction resulted
from the direct exposure.
Limitations
• Such error could have been not exposing the
yeast completely to the smoke source if smoke
escaped through the chimney or if wind was a
factor.
• Because the plates were open, possible
contamination could have been a factor.
• Other reasons cold have been not vortexing the
yeast before each trial, therefore pipetting a
wrong concentration.
Extensions
• Further extensions on the experiment
would be to use different kinds of smoke to
expose to the cells.
• Also, the individual ingredients of products
could be used to see which is the most
effective. For example, use the separate
poisons of a cigarette instead of the entire
cigarette.
Works Cited
• “Modern Day Pollutants” Robert Ingald
• www.bbc.co.uk
• www.epa.gov
• www.sciencedirect.com
• www.tobaccoscam.uscf.edu