Lab 3 - Aerobic and Anaerobic Respiration PDF

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Aerobic and Anaerobic Respiration ...

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Cellular Respiration: Aerobic and Anaerobic Respiration of Yeast BY:

Hypothesis 1 - If a solution of yeast is prepared, added to a glucose solution, and incubated at different temperatures, then the yeast in the 45°C and the 60°C fermentation will be the most efficient as it wont allow the yeast to die from the heat or cold.

Hypothesis 2 - If a solution of yeast is prepared, added to an alternate carbohydrate source of: Galactose, Sucrose, Fructose, Maltose, Lactose, or Lactose + Lactaid, and incubated at 45°C, then the yeast with the maltose solution would produce the most CO2, as maltose is made up of the two monosaccharide’s glucose + glucose.

Hypothesis 3 – If a solution of mitochondrial suspension of lima beans is prepared, then the solution with more succinate in it will have a darker blue color and a higher percent transmittance (%T) than the others, as there are more electrons that have to be reduced resulting in less light being absorbed by the sample.

Results Table 1 Table 1 shows amount of CO2 that is produced by Glucose at different temperatures. Tube 1

Temperature (°C) 0

Initial Gas Volume (mL) 1.4

Final Gas Volume (mL) 1.4

Volume CO2 Produced (mL) 0.0

2

22

1.5

2.5

1.0

3

45

1.6

5.6

4.0

60

1.2

4.5

3.3

5

100

1.5

1.5

0.0

Volume Co2 Produced (mL)

4

4.5 4.0 3.5 3.0 2.5 2.0 1.5 1.0 0.5 0.0

4.0 3.3

1.0

0.0 0°C

22°C

45°C Tempreature

60°C

Volume Co2 Produced (mL) 0.0 100°C

Figure 1: CO2 Produced by Glucose Figure 1 shows a bar graph of the amount of CO2 that is produced by Glucose at different temperatures. The temperatures are all labeled on the x-axis, while the volume of CO2 emitted in mL are shown on the y-axis. In the first part of the experiment, fermentation, a yeast solution was made following the directions according to Table 4. Next these tubes were put into different temperatures to incubate. Table 1 and Table 2 were produced from these results. Looking at Table 1, it was found that at the temperature of 0°C, no CO2 was emitted. At 22°C or the room temperature at that moment in time, only 1mL of CO2 was emitted. At 45°C, 4.0 mL of CO2 was emitted. At 60°C, 3.3 mL of CO2 was emitted, and the boiled Yeast, at 100°C, had no CO2 emissions. Figure 1 is a graphical representation of this data.

Table 2: CO2 Produced by Different Carbohydrates Table 2 shows the amount of CO2 that is produced by different carbohydrates. Tube 3

Carbohydrate Glucose 45°C

Initial Gas Volume (mL) 1.6

Final Gas Volume (mL) 5.6

Volume CO2 Produced (mL) 4.0

6

Galactose

1.1

1.3

0.2

Sucrose

1.0

6.0

5.0

8

Fructose

1.2

7.4

6.2

9

Maltose

1.1

2.5

1.4

10 11

Lactose Lactose+Lactaid

1.6 1.0

1.9 6.0

0.3 4.0

Volume Co2 Produced (mL)

7

7.0 6.0 5.0 4.0 3.0 2.0 1.0 0.0

6.2 5.0 4.0

4.0

1.4 0.2

0.3

Carbohydrates

Figure 2: CO2 Produced by Different Carbohydrates Figure 2 shows a bar graph of the amount of CO2 that is produced by different carbohydrates. The different carbohydrates are labeled on the x-axis, while the volumes of CO2 emitted in mL are shown on the y-axis.

Looking at Table 2, it was found that Glucose had a CO2 emission of 4 mL. Galactose had a CO2 emission of 0.2 mL. Sucrose had a CO2 emission of 5.0 mL. Fructose had a CO2 emission of 6.2 mL. Maltose had a CO2 emission of 1.4 mL. Lactose had a CO2 emission of 0.3 mL, and Lactose+Lactaid had a CO2 emission of 4.0 mL. Figure 2 is a graphical representation of this data.

Table 3: Percent Transmittance of the Mitochondrial Suspension (600nm) Ta ble 3 shows the percent transmittance (%T) of the solutions at various times at 600 nm. Time (minutes) Tube

1

5

10

15

20

25

30

1

100

100

100

100

100

100

100

2

21.8

22.8

23.4

23.9

23.9

24.4

24.7

3

21.8

25.0

28.3

30.0

30.0

38.8

42.3

4

22.2

26.8

32.2

37.1

37.1

48.6

53.6

5

32.2

32.0

32.0

31.8

31.8

31.6

31.7

Percent Transmittance (%T)

Percent Transmittance of the Mitochondrial Suspension (600nm) 120.0 100.0 Tube 1 Tube 2 Tube 3 Tube 4 Tube 5

80.0 60.0 40.0 20.0 0.0 1

5

10

15

20

25

30

Time (Minutes)

Figure 3: Percent Transmittance of the Mitochondrial Suspension (600nm) Figure 3 shows a line graph of the percent transmittance (%T) of the solutions at various times. The time that has passed in minutes is labeled on the x-axis, while the %T is shown by the y-axis.

Table 3 shows the percent transmittance (%T) of the 5 different solutions at 600 nm. From the table it was found that tubes 2,3, and 4 %T went up, while tube 5 %T decreased. Tube 1 is the blank and has a constant %T of 100. Figure 3 is a graphical representation of this data.

Discussion Fermentation occurs when there is no oxygen present during glycolysis. When yeast is fermented, the byproducts of the reaction are CO2 and ethanol. (Alba-Lois, 2010) The experiments aim was to recreate Aerobic Respiration by testing the effects of temperature on the yeast. Looking at Table 1 it was found that there were emissions of CO2 at 22, 45, and 60°C. The yeast solutions at 0°C and 100°C had no CO2 emissions. At those temperatures, the enzymes in the yeast were moving to fast, causing the enzymes to denature (100°C), or moving to slowly (0°C). From the data in Table 1 and Figure 1, it can be inferred that the yeast ferments the best in between the temperatures of 45 and 60°C. These observations agree with the hypothesis, as it was stated that the most efficient temperature would be between 45 and 60°C. The next part of the experiment was to see the effects of using different carbohydrates had on the fermentation process. In order for fermentation to take place, a sugar must gain a phosphate group. This sugar can be one of many carbohydrates. Different Carbohydrates will produce a different amount of CO2 emissions. (Cap, 2006) The carbohydrates used in this experiment were: Glucose, Galactose, Sucrose, Fructose, Maltose, Lactose, and Lactose+Lactaid. It was found that fructose had the highest CO2 emission with sucrose having the second highest. Fructose, a monosaccharide can be converted to glucose, using an enzyme called isomerase. Therefore, all the Fructose became glucose. Similarly Sucrose is a disaccharide made up of two monosaccharaides, glucose and fructose. The fructose would again be converted to glucose, and the added glucose, producing more CO2 emissions. (Hewitson, 2015) Yeast did ferment in all the cases, however when galactose and lactose were used, the CO2 emission was minimal. This is because the enzyme lactase splits lactose into glucose and galactose. Yeast however is not able to process lactase, causing very minimal CO2 emissions. From this data it can also be concluded that disaccharides are a better nutrient source. The addition of Lactaid in the Lactose caused the CO2 emission to go drastically. The lactaid breaks down the lactose into glucose and galactose, allowing for better fermentation. These conclusions go against the stated hypothesis as it was originally thought maltose

would produce the most CO2. Though maltose is made up of two glucoses, it needs the enzyme maltase to break apart, however this enzyme is not present within aerobic respiration. In the last part of the experiment, the %T of 5 different solutions DCIPI ‘s were tested. Specifically the conversion of succinate to fumarate was observed. The reaction reduces FAD to FADH2, producing free electrons. Then DCIPI is used in order to view this reaction. (Lab Manuel) It was found that the tube with more succinct in it, (tube 4) %T increased dramatically. This is because the increased DCIPI allowed for more electrons to bind with it causing less light to be absorbed, leading to a higher %T. This proves the hypothesis as correct because the tube with more succinct in it did indeed have a darker color than the rest and a higher %T. Error is likely to happen in this experiment. During the incubation period, the tubes could have been taken out to early or late, or some of the CO2 could have been lost ending up with incorrect data. Another place error could have occurred is with the yeast itself. If the yeast was bad or contaminated, along with any of the other chemicals, then data could have been incorrect.

Appendix Table 4: Sample Preparation of Aerobic Respiration Table 1 shows the amount of: Phosphate Buffer, Mitochondrial Suspension, DCIPIP, and Succinate needed in each test tube in order to test the %T. Tube

Phosphate Buffer (mL)

Mitochondrial Suspension (mL)

1 mM DCIPIP (mL)

Succinate (mL)

1

4.4

0.5

0.0

0.1

2

4.2

0.5

0.3

0.0

3

4.1

0.5

0.3

0.1

4 5

4.0 4.6

0.5 0.0

0.3 0.3

0.2 0.1

Reference Alba-Lois, L., & Segal-Kischinevzky, C. (2010). Yeast Fermentation and the Making of Beer and Wine. Retrieved February 23, 2016, from http://www.nature.com/scitable/topicpage/yeast-fermentation-and-themaking-of-beer-14372813 Cap, A. (2006, October 2). Yeast Lab – The Catabolization of Glucose, Fructose, Mannose, and Galactose. Retrieved February 23, 2016, from http://adamcap.com/schoolwork/yeast-lab-thecatabolization-of-glucose-fructose-mannose-and-galactose/ Hewitson, J., & Hill, C. (2015). How does sugar affect yeast growth? Retrieved February 23, 2016, from http://www.saps.org.uk/saps-associates/browse-q-and-a/169-q-a-a-how-does-sugar-affect-yeast-growth Lab Manuel...