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Internal Assessment: Fermentation Biology Higher Level Examination session: May 2012

“The Effect of Different Concentrations of Glucose in the Anaerobic Respiration by

Yeast”

I.- Introduction

The world glycolysis (sugar splitting) is thought to have been one of the first

biochemical pathways to evolve. It uses oxygen and occurs in the cytosol of the cell.

The sugar splitting proceeds efficiently in aerobic or anaerobic environments. Glycolysis

is the metabolic pathway that is common to all organisms on Earth. (Miller, & Levine,

2006) However, when oxygen is not present, the process of glycolysis is combined with

different pathways. When combining these pathways with glycolysis, fermentation takes

place.

“Some organisms derive their ATP completely without the use of oxygen without the use

of oxygen and are referred to as anaerobic. The breakdown of organic molecules for

ATP production in anaerobic way is also called fermentation.” (Damon, McGonegal, &

Ward, 2009) “During fermentation, cells convert NADH to NAD+ by passing high energy

electrons back to pyruvic acid. This action converts NADH back into the electron carrier

NAD+, allowing glycolysis to continue producing a steady supply of ATP.” (Miller, &

Levine, 2006)

There exist two types of fermentation which are alcoholic fermentation and lactic acid

fermentation. However, this practice is focus on alcoholic fermentation due to the

production of carbon dioxide (CO2) gas.

Alcoholic fermentation. Yeast (single-celled fungus) uses this type of fermentation in

order to produce ATP molecules. Yeast cells take in glucose from the environment and

generate a net of gain of two ATP by way of glycolysis. (Damon, McGonegal, & Ward,

2009) Glycolysis produces organic products known as pyruvate molecules. The next

step is taken when yeast converts both of the 3-carbon pyruvate molecules to produce

“The Effect of Different Concentrations of Glucose in the Anaerobic Respiration by Yeast / Mayo 2012

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molecules of ethanol. “Ethanol is a 2-carbon molecule, which means that a carbon atom

was lost during the conversion. Therefore, the carbon atom is given off in a carbon

dioxide molecule. The waste products produced by yeast (ethanol and carbon dioxide)

are spread into the environment.” (Damon , McGonega l , & W ard , 2009 )

II.- Objective

The objective of this practice is to study the effect of different concentrations of glucose

on the rate of anaerobic respiration in yeast for the production of carbon dioxide gas

(CO2).

III.- Research Question

How will the effect of different concentrations of glucose (5%, 10%, 15%, 20%, 25%) be

on the production of carbon dioxide gas (CO2) by yeast fermentation at 37°C?

IV.- Hypothesis

If increasing the concentration of glucose then the rate of anaerobic respiration in yeast

will be faster. Therefore, the carbon dioxide (CO2) produced by the reaction will

increased at a higher concentration.

This will happen because for a great part of the reactions occurring in nature in our daily

life, increasing the concentration of one of the reactants increases the rate of the

reaction. For a reaction to occur, the particles of the reactants must collide. Therefore,

“at a higher concentration the collisions are greater.” (Clark, 2002) As yeast contains

certain types of enzymes, the effect of substrate concentration in enzymes is as follows:

“As the concentration of substrate increases, the rate of reaction will increase as well”

(Damon, McGonegal, & Ward, 2009). This is because there is an increase in collisions

between molecules so if there is a higher quantity of molecules, these will collide in a

greater rate. Increasing glucose concentration, there will be more molecules of glucose

to break down in order to produce more ethanol and CO2 gas. In breaking down

molecules, there are more heat energy released and, as a result more molecules will

collide with one another and with the enzymes in yeast. However, we cannot generalize


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the previous statement, because not in all the cases the following statement will have

the same effect. Enzymes have a certain rate at which they can work better.

V.- Variables

For anaerobic respiration in yeast, there is one main factor that affects directly the rate

of reaction between glucose and yeast: temperature. In order for the reaction to take

place, several factors must be met. This factor will be controlled as it can be appreciate

in the following table:

Table 1. Identification of Variables

Type of

variable

Variable Units Control Method

Independent

variable

Glucose

concentration

Percentage

(%)

The percentage of glucose solution

was controlled at the moment of

making the solution. Five different

concentrations of glucose solution were

used (5%, 10%, 15%, 20%, and 25%)

Dependent

variable

Carbon dioxide

(CO2) gas

production

Milliliters

(ml)

The amount of carbon dioxide gas

produced was monitored using a 250

ml graduated cylinder (±5%). It was

measured by means of volume of water

displaced.

(Continuation) Table 1. Identification of Variables

Type of

variable

Variable Units Control Method

Controlled Time Minutes Data will be collected at intervals of 5


4

variables (min) min. during a period of 25 min. (5, 10,

15, 20 and 25 min) by means of a

chronometer (± 0.5 s.).

Temperature Celsius

degrees

(°C)

An optimum temperature of 37°C was

used to carry out the reaction. The

temperature of the water bath was

monitored by means of a

thermometer (± 0.5 ˚C)

Yeast

concentration

Percentage

(%)

Concentration of yeast was controlled

at the moment of making the solution.

Yeast was used at a 10%

concentration.

Yeast volume Milliliters

(ml)

50 ml of yeast solution were required

for each trial throughout the

experiment.

Glucose volume Milliliters

(ml)

In each trial 50 ml of glucose solution

were used at five different concentrations

(5%, 10%, 15%, 20%, and 25%).

VI.- Materials:

250 ml Graduated cylinder (±5 %)

100 ml Graduated cylinder (±5 %)

125 ml Flask (±5 %)

500 ml Beaker (±10 %)

Thermometer (±0.5 °C)

Chronometer (± 0.5 s)

Rubber stopper

Water bath

Support stand rod (base, support and clamp/holder)

Container

Incubator


5

Test tube rack

Reagents

250 ml of 5% Glucose solution





1. 25 L of 10% Yeast solution

Tap water

VII.- Procedure

Focusing on glucose and yeast reaction, the following method was designed with the

objective to measure the production of carbon dioxide (CO2) gas. It must be pointed that

the amount of CO2 was measured by means of the volume of water displaced in the 250

ml graduated cylinder (±5%). Five repetitions were carried out for each of the different

concentrations of glucose solution at intervals of 5 min (5, 10, 15, 20 and 25 min). The

following procedure was used for the development of the experiment:

1. Each flask and beaker used was labelled in order to avoid any confusion.

2. Then the 10% yeast solution was incubated at 37 °C inside the incubator, for a

period of one hour. In addition, each of the five glucose concentrations was

incubated for a period of one hour (at the same temperature), in order to fast the

experiment.

3. While yeast and glucose were being incubated; the electric water bath was filled

with 2 liters1 of tap water and it was turned on.

4. Then the temperature of the water bath was monitored by means of a

thermometer (±0.5 °C) until it reached an optimum temperature of 37 °C.

5. After setting the water bath, a container was filled with approximately 2 liters2 of

tap water.

1 The amount of water tap does affect neither the experiment nor the results.

2 The amount of water tap does affect neither the experiment nor the results.


6

6. A support stand rod was set up besides the container with tap water in order to

hold the 250 ml graduated cylinder (±5%) by means of a clamp/holder attached to

the rod.

7. Before holding the 250 ml graduated cylinder (±5%) by means of the support

stand rod, the graduated cylinder was submerged inside the container in order to

fill the graduated cylinder with tap water.

8. Once the 250 ml graduated cylinder (±5%) was filled with tap water, it was placed

in the support stand.

9. The initial volume of the 250 ml graduated cylinder (±5%) was collected before

initiating the fermentation process.

10. Then the hose of the rod stopper was placed underneath the 250 ml graduated

cylinder (±5%).

11. Once the one hour period was completed, the yeast solution and each of the five

concentrations of glucose solution were taken out from the incubator.

12. Then 50 ml of 5% glucose solution were measured by means of the 100 ml

graduated cylinder (±5 %).

13. After that, the 50 ml of 5% glucose solution were placed into a 125 ml flask (±5

%) in order to introduce it into the electric water bath.

14. In addition, 50 ml of 10% yeast solution were measured by means of the 100 ml

graduated cylinder (±5 %).

15. Then the 50 ml 10% yeast solution were placed into another 125 ml flask (±5 %)

labelled in order to introduce it into the electric water bath together with the flask

containing the 50 ml of 5% glucose solution.

16. The temperature of both solutions was regulated by means of a thermometer

(±0.5 °C), until they reach the optimum temperature of 37 °C.

17. Once the temperature was reached, the reaction was carried out. The flask

containing the 5% glucose solution was mixed with the flask containing 10%

yeast solution.

18. The flask was closed as fast as possible using the rod stopper. This step was

taken into action as fast as possible to prevent a loss of CO2.

19. Then the chronometer (±0.5 s) was activated once the reaction had begun.


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20. In order to measure the volume of water displaced from the 250 ml graduated

cylinder (±5%) to determine the production of CO2 gas delivered by the reaction,

the stopwatch was stopped in intervals of 5 minutes during a 25 min period.

21. Then the water displaced from the graduated cylinder was recorded, by

measuring the final volume in milliliters.

22. The data was collected.

23. For each of the five trials of the same glucose concentration, the same procedure

was followed. After ending with one complete concentration, the same process

was repeated for each of the five glucose concentrations.

VIII.- Set-up of apparatus

Figure 1. Glucose and yeast reaction producing CO2 Figure 2. Temperature controlled by the thermometer

IX.- Safety

During the development of the experiment, neither dangerous materials nor hazard

reactants were used, so the need of gloves and safety goggles was not necessary. In

addition, it was not used a dangerous temperature for the electric water bath.

X.- Data collection

The raw data obtained from the experiment is presented in the following tables. The

tables contained the data of the initial volume3 of the 250 mL graduated cylinder with an

3 The initial volume is represented in the table as V0.


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uncertainty of ±5 % as well as the final volume4 of water displaced by CO2 gas

production, after a period of 25 min, for each of the five different concentrations of

glucose (5%, 10%, 15%, 20% and 25%).

Table 2. Volume of water displaced to determine CO2 (ml) production at 5% glucose concentration.

Volume of water displaced by CO2 gas (ml) at 5% glucose concentration

Trial 1 Trail 2 Trial 3 Trial 4 Trial 5

Time

(min)

±0.5

V0 (ml)

±5%

Vf (ml) by

CO2

±5%

V0 (ml)

±5%

Vf (ml) by

CO2

±5%

V0 (ml)

±5%

Vf (ml) by

CO2

±5%

V0 (ml)

±5%

Vf (ml) by

CO2

±5%

V0 (ml)

±5%

Vf (ml) by

CO2

±5%

5 13.00 26.00 83.00 97.00 15.00 25.00 90.00 102.00 16.00 29.00

10 13.00 47.00 83.00 121.00 15.00 52.00 90.00 123.00 16.00 53.00

15 13.00 64.00 83.00 141.00 15.00 72.00 90.00 143.00 16.00 72.00

20 13.00 71.00 83.00 151.00 15.00 80.00 90.00 159.00 16.00 80.00

25 13.00 81.00 83.00 161.00 15.00 92.00 90.00 164.00 16.00 89.00


Volume of water displaced by CO2 gas (ml) at 10% glucose solution

Trial 1 Trial 2 Trial 3 Trial 4 Trial 5

Time

(min)

±0.5

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

5 26.00 55.00 34.00 62.00 33.00 57.00 16.00 36.00 36.00 65.00

10 26.00 108.00 34.00 125.00 33.00 120.00 16.00 94.00 36.00 124.00

15 26.00 172.00 34.00 177.00 33.00 174.00 16.00 162.00 36.00 179.00

20 26.00 184.00 34.00 190.00 33.00 191.00 16.00 173.00 36.00 192.00

25 26.00 194.00 34.00 206.00 33.00 200.00 16.00 181.00 36.00 200.00




Time

(min)

±0.5

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

4 The final volume displaced after each 5 min interval is represented in the table as Vf.


9

5 12.00 38.00 14.00 36.00 14.00 28.00 12.00 28.00 13.00 37.00

10 12.00 85.00 14.00 78.00 14.00 70.00 12.00 74.00 13.00 75.00

15 12.00 136.00 14.00 134.00 14.00 134.00 12.00 130.00 13.00 138.00

20 12.00 186.00 14.00 200.00 14.00 187.00 12.00 184.00 13.00 189.00

25 12.00 236.00 14.00 230.00 14.00 233.00 12.00 232.00 13.00 246.00




Time

(min)

±0.5

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO (ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

5 32.00 40.00 40.00 51.00 16.00 29.00 26.00 38.00 26.00 39.00

10 32.00 86.00 40.00 92.00 16.00 72.00 26.00 80.00 26.00 83.00

15 32.00 132.00 40.00 138.00 16.00 118.00 26.00 126.00 26.00 132.00

20 32.00 182.00 40.00 184.00 16.00 165.00 26.00 178.00 26.00 180.00

25 32.00 223.00 40.00 230.00 16.00 214.00 26.00 215.00 26.00 221.00

Table 6. Volume displaced of CO2 (cm3) production at 25% glucose concentration.


Trial1 Trial 2 Trial 3 Trial 4 Trial 5

Time

(min)

±0.5 s

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

VO

(ml)

±5%

Vf (ml) by

CO2

±5%

5 12.00 35.00 32.00 48.00 54.00 70.00 34.00 60.00 14.00 34.00

10 12.00 64.00 32.00 88.00 54.00 106.00 34.00 92.00 14.00 73.00

15 12.00 108.00 32.00 124.00 54.00 145.00 34.00 126.00 14.00 104.00

20 12.00 134.00 32.00 144.00 54.00 174.00 34.00 153.00 14.00 133.00

25 12.00 160.00 32.00 175.00 54.00 203.00 34.00 179.00 14.00 161.00

XI.- Data processing

a) Calculating the production of CO2


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In order to obtain the amount of carbon dioxide (CO2) gas produced from the experiment

(which will be measured as volume), we must use the volume of water displaced during

the 25 min period each 5 min.

In the SI, the unit used to measure the volume of any substance is the cm3. As a result,

the volume of carbon dioxide (CO2) gas recorded from the experiment must be

converted to cm3. The following equivalence must be used:

1 Cubic Centimeter (cm3) = 1 Milliliter

Example: For 5% glucose solution initial volume:

13.00 ml = 13.00 cm3

The volume of water displaced represents the amount of CO2 gas produced by the

fermentation of each of the five concentrations of glucose solution and yeast. First of all,

it must be calculated the difference between the volumes of CO2 gas displaced each 5

min interval during the 25 min period. Therefore, we must subtract the final volume of

water displaced by CO2 at each 5 min interval, from the initial volume recorded by the

250 ml graduated cylinder (±5%). This procedure must be used for each of the samples

collected during the experiment. In order to obtain the difference between the volume

displaced each time interval and the initial volume, the following formula must be used:

d ( )

Where:

d = Difference

= Initial volume

= Volume displaced at y- time interval

Example: For 5% glucose solution

d ( )

d = 13.00 cm3

Table 7. Volume of CO2 (cm3) produced during a 25 min period at 5% glucose solution.

Volume of CO2 (cm3) produced during a 25 min period at 5% glucose solution.


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TIME (min)

Trial 1

Trial 2

Trial 3

Trial 4

Trial 5

0 0.00 0.00 0.00 0.00 0.00

5 13.00 14.00 10.00 12.00 13.00

10 34.00 38.00 37.00 33.00 37.00

15 51.00 58.00 57.00 53.00 56.00

20 58.00 68.00 65.00 69.00 64.00

25 68.00 78.00 77.00 74.00 73.00

Table 8. Volume of CO2 (cm

3) produced during a 25 min period at 10% glucose concentration.


TIME (min)

Trial 1

Trial 2

Trial 3

Trial 4

Trial 5

0 0.00 0.00 0.00 0.00 0.00

5 19.00 28.00 24.00 20.00 29.00

10 72.00 91.00 87.00 78.00 88.00

15 126.00 143.00 141.00 126.00 143.00

20 148.00 156.00 158.00 146.00 156.00

25 158.00 167.00 167.00 155.00 164.00

Table 9. Volume of CO2 (cm



TIME (min)

Trial 1

Trial 2

Trial 3

Trial 4

Trial 5

0 0.00 0.00 0.00 0.00 0.00

5 26.00 22.00 14.00 16.00 24.00

10 73.00 64.00 56.00 62.00 62.00

15 124.00 120.00 120.00 118.00 125.00

20 174.00 186.00 173.00 172.00 176.00

25 224.00 216.00 219.00 220.00 233.00

Table 10. Volume of C02 (cm



TIME

Trial 1

Trial 3

Trial 4

Trial 4

Trial 5


12

(min)

0 0.00 0.00 0.00 0.00 0.00

5 8.00 11.00 13.00 12.00 13.00

10 54.00 52.00 56.00 54.00 57.00

15 100.00 98.00 102.00 100.00 106.00

20 150.00 144.00 149.00 152.00 154.00

25 191.00 190.00 198.00 189.00 195.00

Table 11. Volume of CO2 (cm3) produced during a 25 min period at 25% glucose concentration.


TIME (min)

Trial 1

Trial 2

Trial 3

Trial 4

Trial 5

0 0.00 0.00 0.00 0.00 0.00

5 23.00 16.00 16.00 26.00 20.00

10 52.00 56.00 52.00 58.00 59.00

15 96.00 92.00 91.00 92.00 90.00

20 122.00 112.00 120.00 119.00 119.00

25 148.00 143.00 149.00 145.00 147.00

The following table shows the total final volume of CO2 gas displaced for each of the five

different concentrations of glucose solution.

Table 12. Final volume displaced of CO2 (cm3) production of each of glucose concentrations.

Final volumes of CO2 gas (cm3) of each of glucose concentrations

GLUCOSE CONCENTRATION

TRIAL 1

TRIAL 2

TRIAL 3

TRIAL 4

TRIAL 5

5% 68.00 78.00 77.00 74.00 73.00 10% 168.00 172.00 167.00 165.00 164.00

15% 224.00 216.00 219.00 220.00 233.00

20% 191.00 190.00 198.00 189.00 195.00

25% 148.00 143.00 149.00 145.00 147.00

b) Calculating mean and standard deviation


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In order to analyze and to graph the data to discuss the hypothesis previously stated,

the mean and the standard deviation of each of the samples must be obtained. To

obtain the mean of the samples, the following formula must be used:

represents the mean of the values.

is all the values collected through the experiment.-

represents the number of items used in the sample.

Example: Taking the data collected for 5% glucose concentration at 5 min interval:

cm3

The reason of using standard deviation is because in the laymen’s terms, the standard

deviation represents a number which tells how far from the mean a data value is with

respect to how far the other data values are from the mean.

S= √∑ ((∑ ) )

Example: Taking the data collected for 5% glucose concentration at 5 min interval.

S= √∑ (( ) )

S = 1.52

c) Dispersion graphs

n x 2

1 13.00 169.00

2 14.00 196.00

3 10.00 100.00

4 12.00 144.00

5 13.00 169.00

62.00 778.00


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In order to analyze the data obtained by the standard deviation values of each of the five

different sugar concentrations (5%, 10%, 15%, 20% and 25%), it is necessary to plot a

dispersion graph. These graphs will show the correlation between each time (min)

period interval and the mean volumes of CO2 gas (cm3) produced during yeast

fermentation. Furthermore, the graphs will show the error bars which are helpful to

appreciate the standard deviation calculated for each of the five sugar concentration in a

graphical form. The error bars state how far from the mean a value is with respect to

how far the other values are from the mean.

In addition, the line of logarithmic regression can be appreciated from the dispersion

graph. This line shows the logarithmic relationship between each mean value obtained

at each 5 min. interval. From the graphs, the correlation coefficient of Pearson (R2) can

be appreciated. This coefficient is necessary to determine in a mathematical way the

correlation strength between the mean volumes of CO2 gas displaced.

In order to have an accurate analysis form the graphs, the maximum value of 250 cm3

was chosen to be plot in the y-axis. The following tables and graphs show the standard

deviation, the mean values and the regression line for each glucose concentration (5%,

10%, 15%, 20% and 25%):

Table 13. Mean and standard deviation of the data collected of CO2 gas produced at 5% glucose concentration

TIME (min)

TRIAL 1 Volume of

CO2 (cm

3)

TRAIL 2 Volume of

CO2

(cm3)

TRAIL 3 Volume of

CO2 (cm

3)

TRIAL 4 Volume of

CO2 (cm

3)

TRAIL 5 Volume of

CO2 (cm

3)

S

0 0.00 0.00 0.00 0.00 0.00 0.00 0.00

5 13.00 14.00 10.00 12.00 13.00 12.40 1.52

10 34.00 38.00 37.00 33.00 37.00 35.80 2.17

15 51.00 58.00 57.00 53.00 56.00 55.00 2.92

20 58.00 68.00 65.00 69.00 64.00 64.80 4.32

25 68.00 78.00 77.00 74.00 73.00 74.00 3.94


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Graph 1. Mean and standard deviation of the data collected for 5% glucose concentration

From the previous graph it can be observed the rate of CO2 gas released in fermentation

for the 5% glucose solution with yeast. The graph shows the mean values of each trial

performed for the 5% glucose concentration.

When using the 5% glucose concentration, the error bars show that there was a very

small difference between the production of CO2 gas collected in each trial. The

fermentation rate in each trial for the 5% glucose concentration did not differ

significantly. The standard deviation error bars show that the difference in the values

obtained for the production of CO2 gas collected in each trial was very small. This

means that the results of obtained were very near between them which gives a more

accurate information.

Furthermore, the tendency line shows the correlation between the values obtained in

each 5 min interval. As time increases, there is an increased in the production of CO2

gas. Visually, it can be observed that there is a very strong positive correlation between

y = 38.66ln(x) - 50.84 R² = 0.99

0

50

100

150

200

250

0 5 10 15 20 25 30

CO

2 P

RO

DU

CTI

ON

(cm

3 )

TIME (min)

GLUCOSE 5% - CO2 PRODUCTION

Glucose 5%

Tendency line


16

the means. From the graph, it can pointed that the correlation coefficient (R2) is 0.99,

which means that as the coefficient of correlation obtained is approaching to 1, there is a

very strong correlation, and there is not any outlier. The production of CO2 gas released

during fermentation was almost the same in each trial. There was not a significant

difference in the rate of CO2 released in each trial for 5% glucose concentration.

Table 14. Mean and the standard deviation of the data collected for the CO2 gas produced at 10% glucose concentration

TIME (min)

TRIAL 1

Volume of CO2 (cm3)

TRIAL 2

Volume of CO2

(cm3)

TRAIL 3

Volume of CO2 (cm

3)

TRIAL 4

Volume of CO2

(cm3)

TRIAL 5

Volume of CO2

(cm3)

S

0 0.00 0.00 0.00 0.00 0.00 0.00 0.00

5 29.00 28.00 24.00 20.00 29.00 26.00 3.94

10 82.00 91.00 87.00 78.00 88.00 85.20 5.17

15 146.00 143.00 141.00 146.00 143.00 143.80 2.17

20 158.00 156.00 158.00 157.00 156.00 157.00 1.00

25 168.00 172.00 167.00 165.00 164.00 167.20 3.11


y = 92.36ln(x) - 121.23 R² = 0.97

0

50

100

150

200

250

0 5 10 15 20 25 30

CO

2 P

RO

DU

CTI

ON

(cm

3)

TIME (min)


Glucose 10%

Tendency line


17

The previous graph shows the rate of fermentation process for the 10% glucose solution

with yeast. It shows the mean values of each trial performed for the 10% glucose

concentration in order to make a comparison between the production CO2 released in

each of the trials.

From the graph it can be appreciated that the standard deviation error bars show no

significant difference. The fermentation rate in each trial of 10% glucose concentration

did not differ significantly. This means that the results of obtained are more accurate.

Visually, the tendency line of the graph shows the correlation between the values

obtained in each 5 min interval during a period of 25 min. It can be observed that there

is a strong positive correlation between the means from each trial in respect to the

tendency line.

For 10% glucose solution there was a mean between 150 cm3 and 200 cm3 of CO2

released. As time increases, there is an increased in the production of CO2 gas.

According to correlation coefficient (R2) given, which is 0.97; this means that as the

coefficient of correlation obtained is approaching to 1, there is a very strong correlation.

In addition there is not the presence of any outlier. There was not a significant difference

in the rate of CO2 released in each trial for 10% glucose concentration.

Table 15. Mean and the standard deviation of the data collected for the production of CO2 at 15% glucose concentration

TIME (min)

TRIAL 1

Volume of CO2 (cm3)

TRIAL 2

Volume of CO2 (cm3)

TRAIL 3

Volume of CO2 (cm3)

TRIAL 4

Volume of CO2 (cm3)

TRIAL 5

Volume of CO2

(cm3)

S

0 0.00 0.00 0.00 0.00 0.00 0.00 0.00

5 26.00 22.00 14.00 16.00 24.00 20.40 5.18

10 73.00 64.00 56.00 62.00 62.00 63.40 6.15

15 124.00 120.00 120.00 118.00 125.00 121.40 2.97

20 174.00 186.00 173.00 172.00 176.00 176.20 5.67

25 224.00 216.00 219.00 220.00 233.00 222.40 6.58


18


The graph presents the rate of fermentation process for the 15% glucose solution with

yeast. It shows the mean values of each trial performed for the 15% glucose

concentration in relation to the production CO2 released in each trial.

Visually, the graph shows that the standard deviation error bars show a very small

difference between each trial. The fermentation rate in each trial of 15% glucose

concentration did not differ significantly. In addition, the tendency line shows the

correlation between the values obtained during a period of 25 min. There is a strong

positive correlation between the means obtained from each trial and the tendency line.

The 15% glucose solution shows a mean between 200 cm3 and 250 cm3 of CO2 gas

released. Moreover, the as there is a strong correlation, it can be observed that as time

increases, there is an increased in the production of CO2 gas. From the graph, it can be

remarked that the correlation coefficient (R2) is 0.94; the coefficient of correlation

obtained is approaching to 1, so there is a very strong correlation. There was not a

y = 124.66ln(x) - 199.23 R² = 0.9

0

50

100

150

200

250

0 5 10 15 20 25 30

CO

2 P

RO

DU

CTI

ON

(cm

3 )

TIME (min)


Glucose 15%

Tendency line


19

significant difference in the rate of CO2 released in each trial for 10% glucose

concentration.


TIME (min)

TRIAL 1

Volume of CO2 (cm3)

TRIAL 2

Volume of CO2 (cm3)

TRAIL 3

Volume of CO2 (cm3)

TRIAL 4

Volume of CO2 (cm3)

TRIAL 5

Volume of CO2 (cm3)

S

0 0.00 0.00 0.00 0.00 0.00 0.00 0.00

5 8.00 11.00 13.00 12.00 13.00 11.40 2.07

10 54.00 52.00 56.00 54.00 57.00 54.60 1.95

15 100.00 98.00 102.00 100.00 106.00 101.20 3.03

20 150.00 144.00 149.00 152.00 154.00 149.80 3.77

25 191.00 190.00 198.00 189.00 195.00 192.60 3.78


y = 110.63ln(x) - 182.07 R² = 0.94

0

50

100

150

200

250

0 5 10 15 20 25 30

CO

2 P

RO

DU

CTI

ON

(cm

3 )

TIME (min)

GLUCOSE 20% -CO2 PRODUCTION

Glucose 20%

Tendency line


20

The graph shows the rate of fermentation process for 20% glucose solution with yeast. It

shows the mean values of each trial performed for the 20% glucose concentration in

relation to the production CO2 released in each trial.

The graph shows that from the standard deviation error bars there is a very small

difference in CO2 gas production in fermentation process of each trial. The fermentation

rate in each trial of 20% glucose concentration did not differ significantly. From the

tendency line shows the correlation between the values obtained, meaning that there is

a strong positive correlation between the means obtained from each trial. This

correlation observed means that as time increases, there is an increased in the

production of CO2 gas; and such increased was not very different in the rate of reaction

of each of the trials that were carried out.

Mathematically, the correlation coefficient obtained was 0.94; which is a value

approaching to 1, so there is a very strong correlation. There was not a significant

difference in the rate of CO2 released in each trial for 20% glucose concentration.


TIME (min)

TRIAL 1

Volume of CO2 (cm3)

TRIAL 2

Volume of CO2 (cm3)

TRAIL 3

Volume of CO2 (cm3)

TRIAL 4

Volume of CO2 (cm3)

TRIAL 5

Volume of CO2 (cm3)

S

0 0.00 0.00 0.00 0.00 0.00 0.00 0.00

5 23.00 16.00 16.00 26.00 20.00 20.20 4.38

10 52.00 56.00 52.00 58.00 59.00 55.40 3.29

15 96.00 92.00 91.00 92.00 90.00 92.20 2.28

20 122.00 112.00 120.00 119.00 119.00 118.40 3.78

25 148.00 143.00 149.00 145.00 147.00 146.40 2.41


21


The previous graph represents the rate of fermentation process for 25% glucose

solution concentration with yeast. It shows the mean values of each trial performed for

the 25% glucose concentration in relation to the production CO2 released in each trial.

The standard deviation error bars presented in the graph show that there was no

significant difference in CO2 gas production in fermentation process of each trial.

The tendency line shows the correlation between the values obtained. It can be

observed that there is strong positive correlation between the means obtained from each

trial of 25% glucose concentration. Mathematically, the correlation coefficient was 0.971;

a value approaching to the value of 1, meaning that there is a very strong correlation.

There was no significant difference in the rate of CO2 released in each trial carried out

for 25% glucose concentration. It can be observed that as time increases, the production

of CO2 gas increases as well; and such increased was not very different in the rate of

reaction of each of the trials that were carried out.

y = 77.52ln(x) - 112.48 R² = 0.97

0

50

100

150

200

250

0 5 10 15 20 25 30

CO

2 P

RO

DU

CTI

ON

(cm

3 )

TIME (min)


Glucose 25%

Tendency line


22

d) Calculating mean values and standard deviation for final production of CO2 of each

glucose concentration.

In order to compare the effect of the five different concentrations of glucose solution

(5%, 10%, 15%, 20% and 25%) in the fermentation process of yeast, it is necessary to

calculate and to graph the mean values of the final volume of CO2 released for each of

the five concentrations.

Table 18. Values of mean and the standard deviation of final production of CO2 for each glucose concentration.

Glucose Concentration

TRIAL 1

Volume of CO2 (cm3)

TRIAL 2

Volume of CO2 (cm3)

TRAIL 3

Volume of CO2 (cm3)

TRIAL 4

Volume of CO2 (cm3)

TRIAL 5

Volume of CO2 (cm3)

S

5% 68.00 78.00 77.00 74.00 73.00 74.00 3.94

10% 168.00 172.00 167.00 165.00 164.00 167.20 3.11

15% 224.00 216.00 219.00 220.00 233.00 222.40 6.58

20% 191.00 190.00 198.00 189.00 195.00 192.60 3.78

25% 148.00 143.00 149.00 145.00 147.00 146.40 2.41

Graph 6. Mean and standard deviation of the final amount CO2 gas released for each glucose concentration.

74.00

167.20

222.40

192.60

146.40

0

50

100

150

200

250

5% 10% 15% 20% 25%

CO

2 P

RO

DU

CTI

ON

(cm

3 )

GLUCOSE CONCENTRATION

CO2 PRODUCTION

5%

10%

15%

20%

25%


23

As it can be seen from the previous graph, the 5% glucose concentration was the

concentration which produced the less amount of carbon dioxide gas (CO2) during the

experiment, with 74.00 cm3. In addition, it can be observed that there was an increased

in the production of CO2 as the concentration of glucose increased until the 15% glucose

concentration, which showed the maximum production of CO2.

The greatest production of CO2 gas released was presented at 15% glucose

concentration. However, from 15% glucose concentration it was a significant change in

the production of CO2 gas for 20% and 25% glucose concentration; for such

concentrations of glucose, the volume of CO2 gas released started decreasing in a

significant way. From 5% to 15% glucose concentration, there was an increased in the

volume of CO2 production as the concentration increased. However, as the

concentration of glucose was increasing from 20% to 25%, the CO2 gas released was

decreasing.

In addition, from the standard deviation error bars, we can observe that for each of the

different concentrations of glucose (5%, 10%, 15%, 20%, 25%) there was a relatively

small difference from the data collected for the five trials carried out for each sugar

concentrations. This means that the data collected of CO2 gas released for each of the

five concentrations and their respectively trials were constant during fermentation

process.

e) Calculating ANOVA-test.

Visually, we can observe from graph 7 that the means of the final volume of CO2 gas

released from each glucose concentration are different to one another. However, in

order to state whether or not the means of the final volumes of CO2 gas released for

each concentration differ to one another, it is necessary to calculate ANOVA – test5.

5 In order to carry out ANOVA – test, a normality test was previously carried out in order to ensure that the set of data

chosen for ANOVA-test was well modeled by a normal distribution.


24

In order to perform ANOVA – test, it is necessary to group the data in a separate table.

This data refers to the final volume displaced of CO2 gas after the 25 min. period for

each sugar concentration.

Table 19. Final volume of CO2 gas displaced for each glucose concentration after a 25 min. period.

Final volume displaced of CO2 gas (cm3) after 25 min period

Groups Trial 1 Trial 2 Trial 3 Trial 4 Trial 5

5% glucose 68.00 78.00 77.00 74.00 73.00

10% glucose 168.00 172.00 167.00 165.00 164.00

15% glucose 224.00 216.00 219.00 220.00 233.00

20% glucose 191.00 190.00 198.00 189.00 195.00

25% glucose 148.00 143.00 149.00 145.00 147.00

The null hypothesis (H0) as well as the alternative hypothesis (H1) must be stated. After

calculating ANOVA – test:

If F < FC then H0 is accepted.

Hypothesis:

H0= There is no difference between the volume of CO2 gas released in each sample

treated with each glucose concentration.

H1= There is a difference between the volume of CO2 gas released in each sample

treated with each glucose concentration.


25

SUMMARY

Table 20. ANOVA – test summary.

Groups

N0. of measurements

Sum Mean Varience

5% Glucose

5.00 370.00 74.00 15.50

10% Glucose

5.00 836.00 167.20 9.70

15% Glucose

5.00 1112.00 222.40 43.30

20% Glucose

5.00 963.00 192.60 14.30

25% Glucose

5.00 732.00 146.40 5.80

ANOVA

Table 21. ANOVA – test.

Source of variation

Sum of squares

Freedom Degrees

Mean of squares

F P-value F – crit.

Between groups

62939.84 4.00 15734.96 887.98 3.31E-22 2.87

Within groups

354.40 20.00 17.72

Total 63294.24 24.00

As F is equal to 887.98 and Fc equals to 2.87, the value of F is greater than Fc then H0 is

not accepted, 887.7 >2.87. So, there is a difference between the volumes of CO2 gas

released in each sample treated with each glucose concentration.

XII.- Conclusion

As it could be observed in the information presented in the previous tables and graphs,

the hypothesis previously stated was not correct. At the moment of analyzing the raw

data, it could be noticed that the production of carbon dioxide gas by the fermentation of

glucose was increasing while the glucose concentration was increasing. However, this

increase in CO2 gas production was present only when using 5%, 10% and 15% glucose

concentrations; but when fermenting yeast at 20% and 25% glucose concentrations, the

production of carbon dioxide gas started to decrease considerably.


26

When using 5% glucose concentration, there was a mean of 74.00 cm3 of carbon

dioxide (CO2) gas production. This concentration of glucose sugar was the one

producing the less volume of CO2 gas released. Furthermore, analyzing graph 1 with

each of the values collected from the five trials of sample concentration; it could be

observed by means of the standard deviation error bars that there was a very small

difference between the production of CO2 gas collected for each trial, giving a more

accurate information. In addition, the R2 coefficient, 0.99, approached to 1, there was a

very strong correlation, and there were not any outlier. So the production of CO2 gas

released during fermentation was almost the same in each trial.

For the 10% glucose concentration, it could be observed a significant difference

between the mean of CO2 of 5% glucose concentration, releasing a mean volume of

167.20 cm3. The amount of such gas increased at a 10% concentration. In addition, from

graph 2 it could be appreciated that the standard deviation error bars show no significant

difference. The fermentation rate in each trial of 10% glucose concentration did not differ

significantly. With a correlation coefficient (R2) of 0.97, there was a very strong positive

correlation.

Analyzing the data provided by the samples of 15 % glucose concentration, it could be

seen that at this concentration, there was a greater production of carbon dioxide with a

mean of 226.40 cm3 released. At this concentration, glucose fermented yeast at a higher

rate. Furthermore, it was showed in graph 3, by means of the standard deviation, that

there was a small difference on the amount of CO2 released between the data values

collected from each trial. Moreover, the correlation coefficient was 0.94; giving a very

strong positive correlation. There was not a significant difference in the rate of CO2

released in each trial for 10% glucose concentration.

However, the refutation of the hypothesis stated is presented when analyzing the data

from the 20% glucose concentration. At this concentration, the volume of CO2 released

decreased in a significant way. For this concentration was expected a greater volume

released of such gas; however, it happens the other way around, with a decrease in the

released of CO2. Furthermore, there is present the greater difference in the volume

produce of CO2 of each of the trials of the sample, with a mean of 192.70 cm3.


27

When analyzing the results of the 25% glucose concentration, it was a greater

decreased on the production of CO2. For this concentration, and based on the

hypothesis, it was expected the major production of CO2; however, there was a greater

decreased on CO2 with reference to the 20% concentration. In addition to these results,

the standard deviation error bars presented in the graph 5 showed that there was no

significant difference in CO2 gas production in fermentation process of each individual

trial. Mathematically, the correlation coefficient was 0.97, meaning that there was a very

strong positive correlation. There was no significant difference in the rate of CO2

released in each trial carried out for 25% glucose concentration.

As it was shown in the data collected from the experiment the effect of increasing the

concentration of glucose with respect to the volume of CO2 released was not as it was

expected from the hypothesis stated. At the beginning, as the concentration from 5%

increased to a 10% glucose concentration; however as the concentration increased from

15% to 25%, there was a decreased in the volume of carbon dioxide gas released.

According to Damon, McGonegal, and Ward, (2009) a reason for the decreased on

carbon dioxide volume released from yeast respiration, it could be because when

increasing the concentration of a substance, there is an increased of molecular

collisions; however, as yeast contains certain types of enzymes, such enzymes have a

certain limit to which they can work at a maximum rate. As a result, if we continue

increasing the concentration of substrate, there will be a point in which the enzymes will

be working as far as possible until they cannot work efficiently, so the rate of reaction

starts decreasing.

Another reason for the decreased of the volume of CO2 released as the concentration of

glucose was increasing is because as the experiment was carried out within a closed

system, the production of carbon dioxide gas as the concentration of glucose was

increasing, it could have killed the yeast inside the flask.


28

XIII.- Evaluation

Along the experiment it was possible to obtain reliable data when following cautiously

the method previously designed step by step with the objective of avoiding any possible

problem in terms of the control of variables. In addition, a set of five trials were

performed for each sugar concentration (5%, 10%, 15%, 20% and 25%) in order to

gather current and reliable data for the analysis. However, there will be always problems

that can arise when performing the experiment which can be improved for future

repetitions of the experiment.

The data gathered from the production of carbon dioxide (CO2) by means of the final

water displacement of the 250 ml graduated cylinder (±5 %), it can be 80% accurate due

to the lost of CO2 present when mixing the substances and at the moment of closing the

flasks by means of the rubber stopper; a reason why this happened is because when

mixing the glucose solution with yeast, the reaction was very fast and there is a loss in

carbon dioxide gas from the closed system to the environment. However, the

fermentation process was carried out in a way in which it was able to obtain quantitative

data with respect to the volume of carbon dioxide gas released.

In addition, when performing this experiment a problem aroused related with

temperature. As it was pointed out, the optimum temperature to carry out the

fermentation reaction was 37°C; however, the room temperature form the laboratory

affected the optimum temperature set on the electric water bath. Even though

precautions were taken in terms of maintaining the optimum temperature as the use of

the incubator and the use of the electric bath, the control of the temperature was difficult

to maintain due to the opening of the electric water bath.

Besides that, another possible weakness of the experiment was the period of time

considered for the collection of data during the fermentation process. As the data was

collected at 5 min intervals during a 25 min period due to the laboratory conditions, the

data gathered was limited to a fraction of the process. As a result, this can be

considered as a limitation for the overall analysis since it can probably affect the

accuracy of the data for the approval of the hypothesis.


29

XIV.- Improvements

An improvement for the experiment can be the fact of increasing the period of time for

fermentation. As the data was collected during a 25 min period, the data collected

corresponds only to a fraction of the complete fermentation process. It would be best to

record the data until the fermentation process has ended, in order to have more

accurate information about the rate of CO2 released.

Another improvement in order to enhance the experiment could be to have a better

control over the room temperature from the laboratory where the experiment was

performed. Even though some precautions were taken with respect to maintain the

optimum temperature of 37°C as the use of the incubator and the electric water bath, the

opening of the water bath was a limitation because the 125 ml flasks (±5 %) in which the

fermentation process was taking place were exposed to the room temperature from the

laboratory.

XV.- Bibliography

Clark, Jim. (2002). The effect of concentration on reaction rates. Web. 24 Oct 2011.

<http://www.chemguide.co.uk/physical/basicrates/concentration.html

Damon, Alan, McGonegal, Randy, Tosto, Patricia, & Ward, William. (2009). Higher level

Biology developed specifically for the IB diploma. Pearson Education, Inc.

Miller, K. R., & Levine, Joseph S. (2006). Biology. Boston, Massachusetts: Pearson

Education, Inc.

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