BIOL240_Plant&Animal_Energy_Acquisition_and_Use_Lab_Exercise_SP23

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Pennsylvania State University *

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240W

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Chemistry

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Jan 9, 2024

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BIOL240W: Plant and Animal Energy Acquisition and Use Lab Introduction Photosynthesis is a metabolic process that combines carbon dioxide (CO 2 ) and water (H 2 O) to form sugar (C 6 H 12 O 6 ) and oxygen (O 2 ). The process takes place within the chloroplasts of plants and algae. The chloroplasts contain the green pigment chlorophyll and the enzymes necessary for photosynthesis. The chlorophyll traps light, which serves as the energy source that allows the process to take place. A simplified equation for photosynthesis is. 6 H 2 O + 6 CO 2  C 6 H 12 O 6 + 6 O 2 All organisms require energy to sustain themselves. Nearly all organisms, including plants and animals, carry on aerobic respiration, in which sugar and oxygen react to form carbon dioxide, water, and a source of energy known as ATP (adenosine triphosphate). Mitochondria are cellular structures that contain enzymes necessary for the many individual steps or aerobic respiration. A simplified equation for aerobic respiration is: C 6 H 12 O 6 + 6 O 2  6 H 2 O + 6 CO 2 + energy (ATP) Look closely at the balanced equations for photosynthesis and respiration and notice that the end products of one reaction are the raw materials for the other. Only organisms containing chlorophyll can perform photosynthesis, where respiration can take place in virtually every organism. Many plants perform photosynthesis and respiration simultaneously. The P/R ratio (photosynthesis/respiration ratio) compares the rate of photosynthesis to the rate of respiration. Knowing this ration can help explain what happens in a plant at different times in its life. For instance, the P/R ratio is different for a corn plant during spring, summer, and fall. It is also different for day and night. Animals on the other hand can engage only in respiration. Because animals are incapable of converting inorganic raw materials into organic molecules, they must obtain energy-rich molecules be eating plants or other animals. They also need oxygen to allow them to release energy from organic molecules. Thus, animals are dependent on plants for the two end products of photosynthesis, organic molecules (sugar) and oxygen. Water has many gases dissolved in it, including oxygen and carbon dioxide. Aquatic organisms use these gases when they carry on photosynthesis and aerobic respiration and release gases into the water. This exercise allows you to make measurements of the rates of photosynthesis and aerobic respiration by measuring the amount of oxygen and carbon dioxide present in the water in which the organisms live. This laboratory activity gives you an opportunity to set up an experiment with proper controls, quantitatively test water samples for oxygen and carbon dioxide content and collect and analyze

data. You may also learn that experimental work sometimes yields results that are difficult to interpret. Elodea (commonly known as water weed or pond weed) is a genus of a freshwater plant native to the Americas but is widely introduced across the globe (WCSP 2021). Beds of Elodea are often considered noxious for boaters and anglers, but serves as an important contributor to the balance of gases in freshwater ecosystems. Many other organisms (fishes, macroinvertebrates, bacteria) rely on the balance of these gases. One such organism is the aquatic freshwater snail (Class: Gastropoda). Freshwater snails are benthic macroinvertebrates most commonly found in slow moving sections of lakes, ponds, springs, streams, rivers and man-made bodies of water that are highly oxygenated (Johnson et al ., 2013). Benthic macroinvertebrates, such as snails, can be used as indicators of water quality because of their specific sensitivities to water quality changes such as temperature and oxygen. The two major types of freshwater snails are prosobranchs (gilled snails) and pulmonata (lunged snails). While gilled snails solely obtain oxygen from dissolved O 2 in water, lunged snails can absorb O 2 from air or, for certain species of lunged snails, from water stored in their lung-like cavity beneath the shell (Johnson et al . 2013). Therefore, gilled snails tend to be more susceptible to pollution in bodies of fresh water than their lunged counterparts. Ramshorn snails are part of the pulmonata group, while pond snail species are either part of the pulmonata or prosobranchs groups (Nordsieck). Purpose During this exercise you will allow organisms to engage in their normal biochemical processes. Evidence that the organisms carried out photosynthesis or respiration will be revealed by sampling the oxygen and carbon dioxide content of the water in which they live. The tests for measuring the oxygen and carbon dioxide content appear later on. Follow these test procedures carefully, because you will be measuring very small quantities -- parts per million (ppm) -- of oxygen and carbon dioxide. If your water sample contains 8 ppm of oxygen, it means that there are eight oxygen molecules dissolved in every 1 million molecules of your sample, During this lab exercise, your group will: 1. determine dissolved oxygen and dissolved carbon dioxide concentrations in aged tap water. (The purpose of doing this is to get baseline data as well as to give you practice with these complex tests.) 2. set up controls of aged water and three experimental situations: Plants in light, plants in darkness, and a snail. 3. determine dissolved oxygen and dissolved carbon dioxide concentrations of the controls and three experimental situations at the end of an hour incubation. 4. use the data collected to answer questions. Procedure Fill a large beaker with ~675 mL from the container labeled aged water. The aged water is simply tap water that has been sitting open to the atmosphere overnight. Therefore, the aged water has the same concentration of carbon dioxide and oxygen dissolved throughout the container. In addition, it is equilibrated to room temperature. Because everyone in class will use this aged water to setup their experiment, everyone will start with water that contains the same amount of carbon dioxide and oxygen and has the same temperature. Test this aged water for dissolved O 2 and CO 2 . The directions for the dissolved oxygen test and the dissolved carbon

dioxide test are found later in the exercise. Record the results in the proper column of Table 1 (use the separate lab notebook/data collection sheet provided). You will prepare two test tubes per condition, one of which you will test for dissolved oxygen and the other for dissolved carbon dioxide. Controls Fill four large test tubes with aged water; cap the tubes in such a way that no air is trapped. Label these tubes Controls with your table number (e.g., Control Table 2) and place two of them in a test tube rack marked Light. Place the other two control tubes in a test tube rack in the dark. Your instructor will designate these locations. Experimental Tubes Plant in Light Place one healthy sprig of Elodea , approximately the length of the 50mL tube, into each of two tubes. Then, fill the test tubes with aged water. There should be plant material from the top to the bottom of the test tube, but the plants should not be jammed together in a clump. Cap the tubes without trapping air. Label these tubes Plant in Light with your table number (e.g., Plant in Light Table 6) and place in the test tube rack underneath of a florescent light source for 1 hour. It is best to use florescent lights because incandescent lights tend to heat up the water in the tubes and change the amount of gases that can remain dissolved in the water. Plant in Darkness Fill two more test large tubes with Elodea and aged water as you did above; cap tube as before. Label these tubes Plant in Dark with your table number (e.g., Plant in Dark Table 4) and place in the designated dark area for 1 hour. Snail Fill two more large test tubes with aged water and place a snail in the test tube. Cap the tube without trapping air bubbles. Label this tube Snail with your table number (e.g., Snail Table 5) and place them with the plant in the light for 1 hour. Be sure to note on the data collection sheet if you chose a pond or ramshorn snail for these samples. Measuring Dissolved Gasses The following procedures contain chemicals that are potential harmful to human health if used incorrectly. Dedicated outerwear (i.e. a sweatshirt or long-sleeved shirt), gloves, and eye goggles must be worn until the completion of testing. It is important to take your time and follow directions closely. You should notify your TA of any spills. Waste must be disposed of in separately labeled containers for the dissolved CO 2 and O 2 tests. Gently rinse your glassware between tests at any of the sinks. Dissolved Oxygen Test 1. Fill large glass Water Sampling Bottle to neck of bottle (with water from both tubes if necessary). 2. Add 8 drops of Manganous Sulfate Solution. 3. Add 8 drops of Alkaline Potassium Iodide Azide. 4. Cap Water Sampling Bottle and mix (invert 3-4 times). *A dark orange/brown precipitate should form. 5. Allow precipitate to settle (~2 minutes). 6. Add 8 drops of Sulfuric Acid to Water Sampling Bottle.

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7. Cap and mix until reagent and precipitate dissolves. Continue inverting the bottle, this may take 3-4 minutes. *Solution should be a clear yellow/orange color at end of this step. 8. Fill glass test tube to the 20 mL line with yellow/orange solution. 9. Fill Titrator (syringe with pink tip) with Sodium Thiosulfate, 0.025N, until the large ring on the Titrator meets the 0 line at the top of the Titration barrel. 10. Titrate, carefully swirling tube in between every few drops, until sample color is pale yellow. DO NOT DISTURB SOLUTION IN TITRATOR. 11. Add 8 drops of Starch Indicator (4170WT) to the pale-yellow solution. Swirl to mix. 12. Continue titration, with gentle swirling, until blue color just disappears and solution is colorless. 13. Read result in ppm Dissolved Oxygen. 14. When finished, dispose of solutions in labeled O 2 waste bin, rinse Water Sampling Bottle and glass test tube with tap water, and wipe tip of Titrator. The Water Sampling Bottle and glass test tube can be immediately reused. Dissolved Carbon Dioxide Test 1. Fill glass test tube to 20 mL line with sample water. 2. Add 2 drops of Phenolphthalein Indicator, 1%. If solution remains colorless, proceed to Step 3. If solution turns pink or red, no free carbon dioxide is present. 3. Fill Direct Reading Titrator with Carbon Dioxide Reagent B. 4. Insert Titrator into center hole of glass titration tube cap. 5. While gently swirling the tube, add Carbon Dioxide Reagent B, one drop at a time, until a faint pink color is produced and persists for 30 seconds. *Note: place glass test tube on white paper towel to more clearly see faint pink color during this step. 6. Read test result directly from the scale where the large ring on the Titrator meets the Titrator barrel. Record as ppm Carbon Dioxide. 7. Carefully return remaining titration solution back to the carbon dioxide B bottle. 8. When finished, dispose of solutions in labeled waste bin. When you have finished all testing, return Elodea to “Used” container and Snails back to their containers. Table 1. Test Tube Dissolved O2 (ppm) Dissolved CO2 (ppm) Initial Aged Water Control in Light Control in Dark Plant in Light Plant in Dark Ramshorn or Pond Snail in Light (Circle snail used) Analysis of Results Often, a visual presentation of data helps one see patterns or trends and makes interpretation easier. You will need to use the aggregated class data to construct a bar graph including means and standard errors. Use different colors to distinguish between dissolved oxygen and dissolved

carbon dioxide measurements. Additionally, you will perform a statistical analysis to observe differences among measurements.

Assignment As part of this lab series, you will write a formal lab report including background information, methods (laboratory and statistical), results, discussion, and references. You will be provided with more details and expectations in a future lab session and in the assignment posted on Canvas. References Johnson, P. D., Bogan, A. E., Brown, K. M., Burkhead, N. M., Cordeiro, J. R., Garner, J. T., Hartfield, P. D., Lepitzki, D. A., Mackie, G. L., Pip, E., Tarpley, T. A., Tiemann, J. S., Whelan, N. V., & Strong, E. E. (2013). Conservation status of freshwater gastropods of Canada and the United States. Fisheries , 38(6), 247–282. https://doi.org/10.1080/03632415.2013.785396 Nordsieck, R. (n.d.). Fresh Water Snails. Snails and Slugs (Gastropoda). Retrieved January 6, 2023, from https://www.molluscs.at/gastropoda/index.html%2Fgastropoda%2Fmorphology%2Frespirati on.html WCSP ( R.Govaerts ). 'World Checklist of Selected Plant Families. Facilitated by the Royal Botanic Gardens, Kew. Published on the Internet; http://wcsp.science.kew.org/ Retrieved December 8, 2021.'

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