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Laboratory Analysis of Cell Respiration

Explore three sets of experiments demonstrating glucose oxidation, fermentation, and Krebs cycle enzymatic reactions in cell respiration. Compare photosynthesis and respiration using CO2 detection with phenol red. Investigate fermentation in the absence of oxygen.

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Laboratory Analysis of Cell Respiration

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  1. Laboratory Analysis of Cell Respiration www.assignmentpoint.com

  2. CELL RESPIRATION Introduction to cell respiration laboratory This laboratory consists of three sets of experiments that illustrate the oxidation of glucose for the production of ATP. 1.) The photosynthesis vs. respiration experiment illustrates the importance of respiration to plants. 2.) Fermentation products will be tested for to demonstrate the importance of yeast in the fermentation process. 3.) An enzymatic reaction of the Krebs cycle will be studied to show how enzyme activity may be manipulated by changing substrates. www.assignmentpoint.com

  3. CELL RESPIRATION Introduction to comparison of photosynthesis and respiration It is well known that in the presence of light plants perform photosynthesis. It is less well understood that at the same time plants are also performing cell respiration. To demonstrate this we will determine whether CO2 is consumed or produced as Elodea is placed in either a light or dark environment. The change in CO2 will be detected by the pH indicator phenol red. Think about the chemical equations for respiration and photosynthesis. Which one releases CO2 causing an increase and which one uses up CO2 which will cause a decrease? www.assignmentpoint.com

  4. CELL RESPIRATION Introduction to comparison of photosynthesis and respiration (cont.) Phenol red is yellow under acidic conditions (high H ion concentration), pink under basic or alkalineconditions (low H ion concentration) and orange under neutral conditions. A change in CO2 will cause a directly proportional change in H ion. If the CO2 concentration decreases, the H ion concentration will also decrease and the solution will change to pink. If the CO2 concentration increases, the H ion concentration will also increase and the solution will change to yellow. www.assignmentpoint.com

  5. CELL RESPIRATION Experimental procedure for comparison of photosynthesis and respiration Step 1: Fill four test tubes approximately 2/3 full with tap water and add 15 drops of phenol red. What tap water should turn pink. Is Glendora tap water acidic or basic? Step 2: Adjust the pH of the tap water to neutral, orange by gently (remember this word gently) blowing into the solution with a straw. Repeat this procedure with all four tubes. It is important that all four tubes are the same orange color at the beginning of the experiment. www.assignmentpoint.com

  6. CELL RESPIRATION Experimental procedure for comparison of photosynthesis and respiration (cont.) Step 3: Place a piece of Elodea (cut end up) in two of the tubes. Each set of tubes will contain one with Elodea and a control without. www.assignmentpoint.com

  7. CELL RESPIRATION Experimental procedure for comparison of photosynthesis and respiration (cont.) Step 4: Place two tubes, one with Elodea and one without between the lights on the counter. www.assignmentpoint.com

  8. CELL RESPIRATION Introduction to fermentation Fermentation is a pathway for the oxidation of glucose that produces ATP in the absence of oxygen. Fermentation is similar to respiration because it begins with glycolysis. Fermentation differs from respiration because it does not involve the Krebs cycle or an electron transport chain. Fermentation yields waste products other than CO2 and H2O. In humans, fermentation yields lactic acid and in yeast fermentation yields ethyl alcohol in addition to CO2. Since fermentation lacks a Krebs cycle and an electron transport chain, the only energy products come from glycolysis. The 2 NADH are used to produce the alcohol or lactic acid. The total amount of ATP produced from one molecule of glucose is 2. www.assignmentpoint.com

  9. CELL RESPIRATION Experimental procedure for fermentation Tests performed by the Instructor: Test for carbon dioxide (CO2)production: The instructor will demonstrate the reaction between CO2 and barium carbonate by using a straw to breath into a tube of barium carbonate. Note the reaction when the instructor performs this activity. Students will not perform this activity because barium carbonate is toxic. The instructor will then place a tube from a beaker containing yeast in a sucrose solution into a second tube of barium carbonate to test for CO2 production by the yeast. www.assignmentpoint.com

  10. CELL RESPIRATION Experimental procedure for fermentation (cont.) Tests performed by the Instructor (cont.): Test for ethyl alcohol production: The instructor will assemble a distillation apparatus to collect alcohol from both flasks. The flasks will be heated to separate the alcohol from the water. The distillate will be tested for the presence of alcohol as described in the lab manual. This flask contains yeast and sucrose. <===== This flask contains sucrose alone. =====> www.assignmentpoint.com

  11. CELL RESPIRATION Experimental procedure for fermentation (cont.) Tests performed by the Student: Controls for alcohol production: You will prepare two tubes (Tube 1 and Tube 2) that will be used as controls for the presence of alcohol. Tube 1 will be a negative control because it will not contain alcohol. This tube will give a negative result for the presence of alcohol. Tube 2 will contain alcohol and will give a positive result for the presence of alcohol. You will use these tubes to interpret the results of the alcohol tests performed by the instructor. www.assignmentpoint.com

  12. CELL RESPIRATION Introduction to the Krebs Cycle and the Action of Succinic (Acid) Dehydrogenase. Succinic acid dehydrogenase is and enzyme that oxidizes succinic acid into fumaric acid as part of the Krebs cycle. During this reaction the coenzyme FAD is reduced to FADH2. This reaction is shown as Figure 7.4 in your lab manual. This reaction represents a typical enzymatic reaction that involves a substrate (succinic acid), an enzyme (succinic acid dehydrogenase), and a product (fumaric acid). We will use methylene blue as an indicator that this reaction is taking place. Methylene blue will remove the electrons from FADH2 and turn from blue to colorless. Thus we will be able to determine the rate of the reaction by noting how long it takes the methylene blue to turn from blue to clear. www.assignmentpoint.com

  13. CELL RESPIRATION Introduction to the Krebs Cycle and the Action of Succinic (Acid) Dehydrogenase. We will use methylene blue as an indicator that this reaction is taking place. Methylene blue will remove the electrons from FADH2 and turn from blue to colorless. Thus we will be able to determine the rate of the reaction by noting how long it takes the methylene blue to turn from blue to clear. The methylene blue will clear as FADH2 is produced. <==== The methylene blue will start out blue. ===> www.assignmentpoint.com

  14. CELL RESPIRATION Experimental procedure for The Krebs Cycle and the Action of Succinic (Acid) Dehydrogenase. Tube 1: Place approximately 2 cm of liquid hamburger into a test tube. Then add 10 drops of succinic acid and 3 drops of methylene blue to the hamburger and mix. Note the time as you mix. This is a timed experiment and it is important to know the time the reaction started. Once the reagents are mixed do not mix them again (Your instructor will explain why). Not the time when the only blue remaining is a thin band at the surface of the reagents. Tube 2: Place approximately 2 cm of liquid hamburger into a test tube. Then add 10 drops of distilled water instead of succinic acid and 3 drops of methylene blue to the hamburger and mix. Note the time as before and watch for the color to change. www.assignmentpoint.com

  15. CELL RESPIRATION Experimental procedure for The Krebs Cycle and the Action of Succinic (Acid) Dehydrogenase (cont.). Tube 3: Place approximately 2 cm of liquid hamburger into a test tube. Then add 10 drops of malonic acid instead of succinic acid and 3 drops of methylene blue to the hamburger and mix. Note the time as before and watch for the color to change. Tube 4: Add 5 ml of water to the test tube instead of liquid hamburger. Then add 10 drops of succinic acid and 3 drops of methylene blue to the hamburger and mix. Note the time as before and watch for the color to change. Suggestions: Rinse the mixing rod between tubes. Mix all the tubes at the same time. Do wait for tube 1 to finish reacting before starting tube 2, tube 3, or tube 4. www.assignmentpoint.com

  16. CELL RESPIRATION Experimental procedure for The Krebs Cycle and the Action of Succinic (Acid) Dehydrogenase (cont.). Results: Three of the four tubes should clear. The time required for each of the three tubes should be different. To determine why the times are different you should consider the only variable used in this part of the experiment. One of the tubes should not change. Can you guess which one and why? Your results should look like this picture. www.assignmentpoint.com

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