Bialek – Schmidt 1

Introduction

Many people know that salt is used to preserve foods, but little know that sugar can do the

same. Salt, although effective, cam tarnish the taste of food. An example of this is in jams or jellies; salt

would make the spread taste bitter instead of sweet, which is not ideal for what people look for in fruit

spreads. Sugar preserves food by dehydration and the lack of water does not allow bacteria to grow.

This intrigued the researchers, which made them want to see if sugar substitutes can do the same thing.

The researchers carried out a research project in which different amounts of sugar and incubation

temperatures were combined to find out whether or not sugar substitutes can do the same thing that

normal sugar does to prevent bacterial growth. They hypothesized that if the high temperature and high

amount of sugar is combined, then the least amount of bacteria will grow.

Figure 1. Microorganisms Are Tiny Machines That Need Water.

Figure 1, above, compares bacteria and other microorganisms to tiny machines, which need

water to function. The bacteria in food, thrive on water and when too much bacteria is in the food, the

bacteria will spoil it. Sugar when applied on food takes the water away through the process called

osmosis. Osmosis is the process where water goes through a semi-permeable membrane from a less

concentrated solution to a more concentrated solution which equals the solutions out.

This experiment is very applicable to the community and to the real world because sugar

substitutes are a healthy alternative to sugar. For instance, sugar substitutes add sweetness to foods
 Bialek – Schmidt 2

and drinks without adding all the extra calories of sugar. This research gives a concise answer to

whether or not people can use sugar substitutes to preserve their foods. If sugar substitutes cannot

preserve foods, then it is important to inform people that they should continue using normal sugar and

salt.

Though this may sound like a simple topic, there are many complex concepts that go into food

preservation with sugar substitutes. The first scientific concept and principle that was researched was

how exactly sugar preserves foods in the first place, which would help determine whether or not the

first factor, which is the sugar substitute amount, will reduce the amount of bacteria. It turns out that

bacteria need a certain amount of water to survive, and sugar works to absorb enough of that water to

the point where it is very difficult for bacteria to grow and sustain life. This is shown when Scientific

American stated that, “Salt or sugar, whether in solid or aqueous form, attempts to reach equilibrium

with the salt or sugar content of the food product with which it is in contact. This has the effect of

drawing available water from within the food to the outside and inserting salt or sugar molecules into

the food interior. The result is a reduction of the so-called product water activity (a w), a measure of

unbound, free water molecules in the food that is necessary for microbial survival and growth” (Parish).

This evidence discusses how sugar actually preserves food, which if sugar substitutes have the same

properties, then this will help explain how sugar substitutes would reduce the amount of bacteria that

would grow on the Petri dish. So from this evidence, it can be inferred that the higher amount of sugar

substitute that is used, then the less amount of bacteria should grow.

The other factor that is in this experiment is incubation temperature. Of course, living things

need to live in a certain temperature so they can live and reproduce. For E. coli, as the University of

Nebraska-Lincoln states, “The optimal temperature to live in is 37 C°” (Albrecht). The degrees 36 and 38

were used because anything higher than 38 would be too hot for E. coli to grow, and anything below 36
 Bialek – Schmidt 3

would be too cold for E. coli to grow. The temperature is important for bacteria like E. coli because if it’s

too hot, it kills all of them, and if it’s too cold, it kills all of them.

The two factors of sugar substitute amount and incubation temperature when applied to E. coli

on agar inside of a Petri dish, will change the amount of bacteria that will grow, which is the response

variable. This response will help determine whether or not sugar substitutes and temperature effect

bacterial growth through if there is a varying number of percentages covered between the lows,

standards, and highs.

The general method that was used to carry out the experiment was that a starter plate was

prepared through preparing agar on a hot plate, and then pouring that agar into a Petri dish, then

introducing E. coli to water and pouring that water into the Petri dish, and lastly the dish was placed into

a 37 degree incubator for 24 hours. Then every day after that, agar was prepared on a hot plate, and

sugar substitute would be measured on a thousand-gram scale, and 10 grams of agar would be poured

into a beaker and mixed with either .1, .2, or .3 grams of sugar substitute that was measured out. After

that, E. coli from the starter plate would be introduced to water in a test tube, which would then be

diluted twice, and then poured onto the correct Petri dish based on the trial being ran. Lastly, the Petri

dishes would be put into one of the three incubators, which are 36, 37, and 38 degrees, based on the

trial being ran, and wait 24 hours for results.

 Bialek – Schmidt 4

Problem Statement

Problem:

Can sugar substitutes and temperature (Celsius) affect microbial spoilage in agar over 30 days?

Hypothesis:

If the high concentration of sugar and high temperature is combined, then the least number of

bacteria will grow.

Data Measured:

The independent variables are temperature and sugar concentration. The dependent variable is

the Escherichia coli growth. The standard temperature, 37 degrees Celsius, was chosen because it is the

recommended temperature for E. coli bacteria growth, and the high and low, 38 and 36 respectively,

were one degree off the standard because if the temperature was more off it would be harder to

stimulate bacteria growth. The standard for sugar concentration would be 10 grams of sugar because in

jams that are sold, the average sugar concentration is 10 grams of sugar per 20 grams of jam to be the

best preserved while not having drastic health effects. The low which is 5 and the high which is 15 would

be used because then, our data that we will collect will show the effects of low sugar concentration and

high sugar concentration.

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Experimental Design

Materials:

nutrient agar Bunsen burner

Petri dishes (10 x 10 x 1 cm) sparker
artificial sweetener (Great Value Sweetener) hot mitt
sterilized water hot plate
Incubator (37, 38, 39 C°) 80 ml beaker
Escherichia coli gram scale
1 ml dropper pot of boiling water (at least more than 6 quarts)
Fridge (4 C°) beaker tongs
transfer loop wax paper
test tubes (10 ml)

Procedures:

1. Gather materials.

2. Prepare nutrient agar on a hot plate until the agar is clear (around 15 minutes).

3. Pour water into a pot, and put the pot onto a hot plate and wait until the water within the pot is
boiling (around 15-20 minutes).

4. After the water is to a boil, disinfect a beaker in the pot of boiling water by grabbing the beaker
with beaker tongs, then by submerging the beaker in the boiling water for around five seconds,
lastly pour out the boiling water.

5. Place the now disinfected beaker on the gram scale and zero the scale.

6. Take the liquid nutrient agar off the hot plate once it is ready, and pour it into the beaker on the
zeroed gram scale until there is 10 grams of agar in the beaker.

7. Measure out the correct amount of artificial sweetener (0.1, 0.2, and 0.3 grams) on sheets of
wax paper by using the gram scale.

8. Mix the correct amount of artificial sweetener depending on the run (0.1, 0.2, and 0.3 grams),
with the 10 grams of nutrient agar within the beaker.

9. Label a new Petri dish with the date, the researcher’s names, the class, the run, and the test that
is being ran.

10. Pour the nutrient agar and artificial sweetener mixture contained within the beaker into the
correct Petri dish.

11. Allow the nutrient agar and artificial sweetener mixture to dry, which should take around ten
minutes.
 Bialek – Schmidt 6

12. Use a dropper to gather sterilized water up to the 1 ml mark on a dropper, and drop that
amount of water into seven clean test tubes.

13. Sterilize the transfer loop by heating it up with the flame of a Bunsen burner.

14. Use the transfer loop to transfer Escherichia coli (E. coli) into one clean test tube with sterilized
water in it.

15. Sterilize the transfer loop again with the flame of a Bunsen burner, and then insert the transfer
loop into the first test tube with the E. coli in it, and then take the transfer loop out and place it
into a second test tube with sterilized water in it.

16. Sterilize the transfer loop again with the flame of a Bunsen burner, and then insert the transfer
loop into the second test tube with the first dilution of E. coli in it, and then take the transfer
loop out and place it into a third test tube with sterilized water in it.

17. Sterilize the transfer loop again with the flame of a Bunsen burner, and then insert the transfer
loop into the third test tube with the second dilution of E. coli in it, and then take the transfer
loop out and place it into a clean test tube with sterilized water in it.

18. Repeat step 17 until all remaining clean test tubes with sterilized water in them have been
introduced to the second dilution of E. coli within the third test tube.

19. Pour the sterilized water and the second dilution of E. coli mixture onto the dried nutrient agar
and artificial sweetener mixture in the Petri dish, and move the sterilized water and second
dilution of E. coli mixture around until it covers the surface area of the agar.

20. Pour any access sterilized water and second dilution of E. coli mixture out.

21. Turn the Petri dish upside down, and place it into the correct incubator depending on the trial
being ran.

22. Wait 24 hours and take the Petri dish out of the incubator and place it into a fridge.

23. Observe and gather data on how much bacteria has grown.

24. Repeat until all runs have been completed.

 Bialek – Schmidt 7

Diagrams:

Figure 1. Sterilizing the Transfer Loop

Figure 1, above, shows the loop of the transfer loop being heated up by the top of the Bunsen

burner flame, subsequently sterilizing the transfer loop.

Figure 2. Introducing Bacteria to The Agar

Figure 2, above, shows the Escherichia coli and sterilized water mixture being poured onto

the dried agar and artificial sweetener solution.

 Bialek – Schmidt 8

Figure 3. The Process of Preparing the Petri Dish

Figure 3 above shows the general materials being used to prepare the Petri dish and putting it

into the incubator.

 Bialek – Schmidt 9

Data and Observations

Data:
Table 1
Design of Experiment Values
Sugar Substitute Incubation Temperature
Concentration (grams) (Celsius)
- Standard + - Standard + Table 1, above, shows the
0.1 0.2 0.3 36 37 38
low, standard, and high levels for each factor in the experiment. The degrees 36, 37, and 38

were chosen because they are the recommended temperatures for bacteria to live in without

killing them. The amount of sugar 0.1, 0.2, and 0.3 were chosen because those amounts are

because they are the recommended amount of sugar without killing all of them (Albrecht).

Table 2
Percentage Covered of E. coli for each DOE
Percentage of Petri Dish Surface Covered
DOE
Sugar Substitute Concentration, Incubation Temperature
1st Run Standard (-,-) (+,+) Standard (+,-) (-,+) Standard
Percentage Covered 0.89 0.85 0.78 0.88 0.98 0.99 0.83
2nd Run Standard (-,-) (+,-) Standard (+,+) (-,+) Standard
Percentage Covered 0.78 0.78 0.85 0.95 0.80 0.71 0.62
3rd Run Standard (+,+) (-,+) Standard (+,-) (-,-) Standard
Percentage Covered 0.80 0.98 0.83 0.75 0.32 0.89 0.89
4th Run Standard (+,-) (-,-) Standard (+,+) (-,+) Standard
Percentage Covered 0.55 0.80 0.16 0.72 0.66 0.74 0.76
5th Run Standard (-,-) (-,+) Standard (+,+) (+,-) Standard
Percentage Covered 0.99 0.98 0.85 0.93 0.99 0.97 0.88
6th Run Standard (+,+) (-,-) Standard (+,-) (-,+) Standard
Percentage Covered 0.98 0.78 0.99 0.92 1.00 0.94 0.93

Table 2, above, shows how many DOE’s were completed during the testing period, and

it showed the results of each trial that was conducted within the run. There was a total of six

runs, and 7 trials within each.

Table 3
Average Percentage of Bacteria Covered for Each Type of Trial
  Average (percentage)
 Bialek – Schmidt 10

(+,+) 0.83
(+,-) 0.82
(-,+) 0.87
(-,-) 0.86
Standar
0.84
d

Table 3, above, shows the average percentages of each type of trial that was ran.

Observations:
Table 4
Observations
Date Observation

The percentage of bacteria covering the petri dish for the (+,-) in the
3/15/2017
third run was extremely low, compared to the average of standards.

The percentage of bacteria covering the petri dish for one of the
3/21/2017 standards in the fifth run was 100 percent covered, which is the
highest out of any recorded.

When the (+,-) petri dish was prepared, the agar set in an abnormal
3/23/2017
pattern, probably formed from the temperature of the agar.

Table 4, above, shows the unusual observations that were recorded throughout our
trials.

Figure 1. Bubble Observed on agar.

 Bialek – Schmidt 11

In Figure 1, above, the trial for the fifth run is shown. Within the Petri dish in the picture,

there was most likely a bubble when it was going in the incubator. This was unusual because no

other results had this kind of “bubble” on the agar.

Data and Analysis Interpretation

 Bialek – Schmidt 12

Experiment: The Effect of Sugar Substitute Amount and Incubation Temperature on Bacterial Growth.

Hypothesis: If the high concentration of sugar substitute and high temperature are combined, then the

least amount of E. coli will grow.

Response Variable: Percentage of Bacteria Grown.

Predictor Variable: Sugar Substitute Amount.

Predictor Variable: Incubation Temperature.

Table 1
Variables Used in the Experiment
Sugar Concentration (grams) Incubation Temperature (Celsius)
(-) Standard (+) (-) Standard (+)
0.1 0.2 0.3 36 37 38

Table 1 shows the factors that were chosen for the experiment to see how they effected the

amount of E. coli grown on a Petri dish. The amount of sugar in grams, and the incubation temperature

in Celsius, were used as the two factors. The table above shows the lows, the standards, and the highs

for the two factors. The standard for the sugar concentration was chosen because .2 grams of sugar to

the amount of agar used in the Petri dish is proportionate to the amount of sugar used in jams to

preserve them. The standard temperature was chosen because 37 degrees Celsius is the optimal

temperature to grow bacteria in.

Table 2
 Bialek – Schmidt 13

Data Gathered for Six Runs

Percentage of Petri Dish Surface Covered
DOE
Sugar Substitute Concentration, Incubation Temperature
1st Run Standard (-,-) (+,+) Standard (+,-) (-,+) Standard
Percentage Covered 0.89 0.85 0.78 0.88 0.98 0.99 0.83
2nd Run Standard (-,-) (+,-) Standard (+,+) (-,+) Standard
Percentage Covered 0.78 0.78 0.85 0.95 0.80 0.71 0.62
3rd Run Standard (+,+) (-,+) Standard (+,-) (-,-) Standard
Percentage Covered 0.80 0.98 0.83 0.75 0.32 0.89 0.89
4th Run Standard (+,-) (-,-) Standard (+,+) (-,+) Standard
Percentage Covered 0.55 0.80 0.16 0.72 0.66 0.74 0.76
5th Run Standard (-,-) (-,+) Standard (+,+) (+,-) Standard
Percentage Covered 0.99 0.98 0.85 0.93 0.99 0.97 0.88
6th Run Standard (+,+) (-,-) Standard (+,-) (-,+) Standard
Percentage Covered 0.98 0.78 0.99 0.92 1.00 0.94 0.93

Table 2 above shows the results of how much E. coli covered agar in a Petri dish (%) after it was

subjected to E. coli, and was effected by various temperatures and sugar substitute concentrations.

After twenty-four hours in one of the three different temperature incubators, the data was collected.

The runs also show the randomization in the experiment, which was done by using the “randint”

function on the TI-Inspire CX. The percentages were calculated by placing a tested Petri dish on a grid,

and counting how many squares were filled by bacteria, and dividing that number by the total number

of boxes on the grid. Most of the percentages landed around 80, but a few were really low, such as the

low sugar substitute concentration and low incubation temperature trial in the fourth run. This can show

that there was a lurking variable in the experiment or that somewhere, the researcher’s made a mistake

in their experimental design when testing this.

Table 3
 Bialek – Schmidt 14

Average and Grand Average Percentage of E. Coli on Petri Dish

(SA,IT)  Average (percentage)
(+,+) 83.16́
(+,-) 82
(-,+) 84.3́
(-,-) 77.5
Standar
83.61́
d
Grand 81.75

Table 3, above, shows the averages and the grand average of each combination of factors. The

averages were calculated by adding all runs under the specific condition, and dividing it by six because

there were six runs that were completed. For the standards, it was divided by eighteen because for each

run, there were three standards that were ran, and there were six runs in total. The grand average was

calculated by adding the four averages and dividing it by four. All the averages are within 6.8 3́ of each

other, so that could mean that the effects did not have much of an effect on the amount of bacteria that

grew. The grand average is calculated so that when there are future experiments being done, the

amount of bacteria (percentage) should be around that grand average. As seen in rows one and three in

Table 3, the high incubation temperatures produced higher percentages of bacteria covering the petri

dish, than the low incubation temperatures in row two and four.

Table 4
Effect of Sugar Substitute Amount
Sugar
Sugar Substitute
Substitute EffectEffect
(grams)
.1 ( - ) .3 ( + ) 85
Percentage (%)

83
77.5 81 6́
83.1
84.3́ 7982
77
75 3́
80.9167 82.58
-1 82.583́ – 80.9167 = 1.16́
Sugar Substitute

Figure 1. Effect of Sugar Substitute Amount

Table 4 above shows how much the Petri dish was covered when different amounts of sugar

substitute were given. To find these numbers, Table 3 is was used because it gives the averages for all
 Bialek – Schmidt 15

factors. When the low sugar substitute amount was applied, the average coverage of the Petri dish of E.

coli was 80.9167 %. When the high sugar substitute amount was applied, the average coverage of the

Petri dish of E. coli was 82.5833 %. The effect was found by calculating the averages for both the low and

high percentages, then by subtracting the low average from the high average. The high and low average

for sugar substitute concentration were very close, which can mean that it did not have much of an

effect on the growth of bacteria. Plus, the high average of sugar substitute amount had a greater

amount of bacteria that grew as compared to the low average.

Figure 1 above shows on graph, the effect of sugar substitute amount. On average, as the

amount of sugar increased, the percentage of the Petri dish covered changed by 1.6 %, and had a

positive slope. This was calculated by subtracting the high average amount and the low average amount.

This also shows that the sugar substitute concentration did not have too big of an effect on the amount

of bacterial growth. This effect that was found is important because it shows how the sugar substitute

played a role in the experiment.

Table 5
Effect of Incubation Temperature
Incubation
Incubation Temperature
Temperature Effect
Effect (Celsius)
36 ( - ) 38 ( + ) 85
Percentage (%)

83
77.5 83.1 6́
81
79 3́
82 84.
79.75 83.75
77
83.75 – 79.7575
=4
-1 1
Incubation Temperature

Figure 2. Effect of Incubation Temperature

Table 5 above shows how much the Petri dish was covered when different temperatures were

applied. To find these numbers, Table 3 was used because it gives the averages for all factors. When the

low temperature was applied, the average coverage of a Petri dish that was covered with E. coli was
 Bialek – Schmidt 16

79.75 %. When the high temperature was applied, the average coverage of a Petri dish that was covered

with E. coli was 83.75 %. This shows that there was not too big of an impact on the amount of bacteria

that grew when subjected to different temperatures, but there still was one.

Figure 2, above, shows the effect of temperature on a graph. On average, as the temperature

increased, the percentage of bacteria that covered the Petri dish went up by 4 %, and the line had a

positive slope. This was calculated by subtracting the high temperature average and the low

temperature average. This shows that the incubation temperature had an effect on the amount of

bacteria that grew that was greater than the effect of sugar substitute concentration, but still relatively

low. This effect that was found is important because it shows how the incubation temperature played a

role in the experiment.

Table 6
Interaction Effect of Incubation Temperature and Sugar
Interaction Effect Substitute Amount
85Incubation Temp.
Percentage (%)

83
81 (Celsius)
  7936 ( - ) 38 ( + )
77
Solid Segment 75
-1 1 82 83.16́
Amt. of Sugar .1 (+)
Incubation Temperature
Substitute (grams) Dashed
77.5 84.3́
Segment .3 (-)

Figure 3. Interaction Effect of Incubation

Temperature and Sugar Substitute Amount

Table 6 above compares the total averages between the amount of sugar substitute and the

incubation temperature when they interact with one another. These numbers were taken from Table 3,

because this table just shows the averages in a different order.

Figure 3 above shows the interaction effect between the amount of sugar substitute and the

temperature. The solid segment represents the high amount for sugar substitute amount and the dotted

segment represents the low amount of sugar substitute. The slope of the solid segment is 0.58 3́ and the
 Bialek – Schmidt 17

slope of the dotted segment is 3.416́ . The slopes were calculated by subtracting the high amount and

the low amount and then dividing it by two. With these values, the interaction effect can be calculated.

Use the high sugar substitute amount and subtract the low sugar substitute amount to get -2.8 3́. When

the two factors interact, their effect on the response variable will cover the Petri dish less by -2.8 3́ %.

Since the two slopes are different, there is probably an interaction, and if there was one, it would be

small. When looking at the high temperature average, it was 83.75 %. When graphing the high

temperature, the values used were 82 and 83.1 6́ %. These two numbers have a small difference, so it

can show that there is no too much of an effect from this factor. When looking at the low temperature

average, it was 79.75 %. When graphing the low temperature, the values used were 77.5 and 84. 3́ %.

There is a bigger difference in these numbers as compared to the high, which can show that this had a

bigger effect on the amount of bacteria that grew.

Table 7
Standards of Data
Standards Data (Percentage Covered)
89 88 83 78 95 62 80 75 89
55 72 76 99 93 88 98 92 93

Table 7 above shows how much of the Petri dish was covered by bacteria under the standard

conditions. The range of standards was from 55 % to 99 %, giving a range of standards of 44 %. The

overall range of the data is 84% so it makes sense that the range of standards is lower than the overall

data range by a half almost. It should be less than the overall range of the data because the standard

conditions is only under one condition while the overall data range is under all conditions. With these

standards, they could have been better because with range of standards, it’s a big gap especially

because one was 55% while the highest was 99%. These values do influence the range of standards and

it affected it by making it too big. The range of standards doubled is 88%, which is almost impossible for

a Petri dish covered with E. coli to be statistically significant.

 Bialek – Schmidt 18

Standards

100
Percentage Covered (%)

80

60

40

20

Run 1 Run 2 Run 3 Run 4 Run 5 Run 6

Figure 4. Dot Plot of Standards

Figure 4 above shows how much the Petri dishes were covered under standard conditions. The

range of all the standards was 44 and the overall range was 84 which is to be expected. Double the

range of standards was 88, which is quite large and makes it hard for things to be statistically significant.

The pattern in this graph is that the standards start out high, then they go down near run 4, then they

pick back up again near the end. This suggests that there was possibly a lurking variable which might

have affected the data. One of those lurking variables could have been that when transferring E. coli

from the starter plate to the other test plates, the incorrect amount of E. coli could have been

transferred.
 Bialek – Schmidt 19

Figure 5. Test of Significance

Figure 5 above shows the dot plot for the test of significance. The three effect values are

compared to the range of standards doubled to see if anything is deemed significant. If any value is

outside the vertical lines (which are also just known as fences), it is deemed significant. Since no value is

outside the lines (fences), these effects are not significant. Although none are significant, the incubation

temperature is the most significant by being the farthest away from zero.

Y = Grand Average + Effect of First Predictor Variable/2 * First Variable + Effect of Second Predictor
Variable/2 * Second Variable + Effect of the Interaction/2 * First Variable * Second Variable + “noise”

Y = 81.75 + 1.6́ /2 * (S) + 4/2 * (T) + (-2.83́/2) * (S)(T) + “noise”

(S) = Sugar Substitute Amount

(T) = Incubation Temperature
Figure 6. Prediction Equation

Figure 6 above shows the prediction equation. The prediction equation can be used to help

determine the outcome of future experiments. Noise is all the factors that can influence the results for

instance, not doing the procedures correctly.

Y = 81.75 + 1.6́ /2 * (1) + 4/2 * (1) + (-2.83́/2) * (1)(1) + “noise”

Y = 83.16́

Figure 7. Check of Prediction Equation

Figure 7 above shows the checked prediction equation. The values one and one were used for

the variables because the condition for the high amount of sugar substitute and the high temperature

was used and the high amount of sugar substitute and the high temperature is one. When the prediction

equation is solved, the answer is 83.16́ , which was the average for the high amount of sugar substitute

and the high temperature condition in the experiment. This means that the math was calculated
 Bialek – Schmidt 20

correctly and when using different interpolated numbers, it will show how much the Petri dish was

covered.

Y = Grand Average + Statistically Significant Data + “noise”

Y = 81.75 + “noise”
Figure 8. Parsimonious Prediction Equation

Figure 8 above shows the parsimonious prediction equation. Only statically significant data can

be used in the equation, and there was none, so there is only the grand average and “noise” in the

equation. With this equation, people who want to predict how much the E. coli will cover the Petri dish

can use values between the low and high factors used for this experiment.

Using 37.5 degrees Celsius and using 0.25 grams of Sugar Substitute

Y = 81.75 + “noise”
Figure 9. Interpolated Parsimonious Prediction Equation

Figure 9 above shows an interpolated parsimonious prediction equation. The researchers used

these factors to see how much the E. coli will cover the Petri dish. Since none of the predictor variables

were significant, that means it would just be 81.75 + “noise”. This means that when the researchers do

this experiment again, they should get an E. coli coverage of around 81.75 %.+

Looking at all of the data, no effects were statistically significant. The researchers would like to

talk about the Petri dish that only 16% of it was covered in the 4 th run. This, compared to the other Petri

dishes that were under the same condition was extremely low. The average for the Petri dishes under

the low factors was 77.5 % while this one Petri dish was 16 %. This can show that somewhere in the

experimental design, the researchers either did not pour the right amount of sugar or there was some

lurking variable that affected this Petri dish that badly. Though out of the data that was collected, the

incubation temperature had the greatest effect on the amount of bacteria that grew, and the sugar

substitute amount had the least effect on the amount of bacteria that grew. Also, all of this data may be
 Bialek – Schmidt 21

inconclusive and could have happened by chance. This is due to the fact that the outcome when working

with bacteria may be influenced by many different things. Another possibility is that the incubation

temperature and sugar substitute amount just plainly had no effect on the amount of bacteria that

grew.

Conclusion

It was hypothesized that if the high sugar substitute concentration and high incubation

temperature were combined, then the least number of bacteria would grow. This hypothesis was

rejected because the high sugar substitute concentration and high incubation trials produced on average

83.16́ percentage of bacteria covering the Petri dish, while the low sugar substitute concentration and

low incubation temperature trials produced on average 77.5 percentage of bacteria covering the Petri
 Bialek – Schmidt 22

dish. This shows that the hypothesis was wrong because the high sugar substitute concentration and

high incubation temperature trials did not have the least number of bacteria grown. This is

potentially due to the fact that the researchers were wrong in their thinking that the sugar substitutes

would actually influence the number of bacteria that would grow. 

 The researchers wanted to test to see how temperature and sugar substitute would affect the

growth of E. coli. The reason the researchers wanted to investigate this topic was because food is a

necessity for survival, and the longer that food stays edible, the longer it can be used. So, it is important

to know if sugar substitutes can prevent bacterial growth, to make sure that foods can be good for

longer periods of time. E. coli,  one very popular bacteria in food spoiling, can also cause terrible side

effects, such as when consumed, it can give the host bloody diarrhea, urinary tract infections, or even

death. Therefore, this research is also important so that those problems could be prevented.  

This research was conducted by preparing Petri dishes with agar and different amounts of sugar

substitutes, and then introducing E. coli to the dish, and lastly placing the dish in one of three

incubators. After six runs of this, with 7 trials in each run, the results yielded in data that either shows

that sugar substitutes and temperature do not have much of an effect on bacterial growth, the sugar

substitute could have no effect on bacterial growth, or data that could have all happened by chance.

This is because there was no statistically significant data, and because all the averages were close

in percentage of bacteria covered. Though the incubation temperature did have the greatest overall

effect on the amount of bacteria that grew, while the sugar substitute barely had any effect. This could

be because sugar substitutes are too different in their makeup to be able to have the same food

preserving properties as normal sugar. There were not many researchers who went into this topic, and

the researchers were not able to find a study involving sugar substitutes and temperature affecting E.

coli growth. Which means that the researchers will not be able to compare their findings to other

people’s findings. Though the other researchers that have not went with sugar substitutes, they
 Bialek – Schmidt 23

experimented with sugar and found that sugar prevents microbial spoilage by going through osmosis.

The osmosis makes the bacteria dehydrated and so that means that they can’t reproduce and they die

off (Rettner). 

The researchers may potentially have had a design flaw. The researchers used square grid and

counted how many squares it filled divided by the total number of squares in the square grid. As the

readers can see, the squares on the boundary of the Petri dish can be counted as half a square or a

quarter of a square which is not accurate. This could have led to inaccurate percentages in

the amount of bacteria covering the petri dish. Also, the researchers potentially messed up in the

testing. In the fifth run, testing for the low sugar substitute and the low temperature, the result was 16

%. Now 16 % for that factor is low because the average for the low sugar substitute amount and the low

temperature was 77.5 %. The researchers could have messed up in the amount of sugar substitute that

was put in or not the right amount of E. coli was put on it, and this could have been the case for other

dishes. 

Suggestions to further improve the understanding in this topic is to also test how much E.

coli  would grow with no sugar substitutes. The reason why is so that the researcher(s) can compare the

results with the Petri dishes that have the sugar substitute in them to see if it influences it. This

experiment can benefit society because people always look for heathy alternatives, and sugar

substitutes are a great alternative to sugar for people who cannot normally consume

it. Specifically, people with diabetes will benefit from this research, because they will know If they will

be able to preserve foods without sugar. Lastly, it’s also good to inform people that are using sugar

substitutes to try to preserve foods, if the method is effective or not. The artificial sweetener has also

become a very popular choice for sweetening stuff because it gives the same taste of sweetness, but it

gives fewer calories (Selim). It also benefits the society through the way of preserving jams and

jellies ("Sugar in Food Preserving."). 

 Bialek – Schmidt 24

The lessons that the researchers learned is to always make sure you follow your procedures

correctly and make sure to always try to have the correct amount of bacteria on each Petri dish because

if the Petri dishes don’t have the correct amount of bacteria on it, that can majorly influence the results

thus ruining your experiment.

Works Cited

Albrecht, Julie A. "Escherichinia coli O157:H7 (E Coli)." Escherichinia coli O157:H7 (E Coli) | UNL Food.

n.p., n.d. Web. 10 May 2017.

Parish, Mickey. "How Do Salt and Sugar Prevent Microbial Spoilage?" Scientific American. n.p., 17 Feb.

2006. Web. 30 Jan. 2017.

 Bialek – Schmidt 25

Rettner, Rachael. "Sugar Helps Antibiotics Trick and Kill Deadly Bacteria." Live Science. n.p., 12 May 2011.

Web. 30 Jan. 2017.

Selim, Jocelyn "The Chemistry of . . . Artificial Sweeteners." Discover Magazine. Kalmbach Publishing

Co., n.d. Web. 31 Jan. 2017.

"Sugar in Food Preserving." Sugar in Food Preserving -. ACS Distance Education, n.d. Web. 30 Jan. 2017.