Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
LAB 13 - Nitrogen Cycling April 01-05, 2002 Purpose Learning Goal Students will gain an understanding of the significance of the nitrogen cycle in both human and ecological contexts. Overview (Strategy) Students will follow a schematic diagram of the nitrogen cycle as they complete activities that address sources of input (ammonia, urea, uric acid, fertilizers) and environmental response to varying levels of nitrogen products. Time One Lab period Level Introductory non-majors BIOL101/102/105 Prerequisites and Concurrent Lectures Human Nutrition and energy conversion Primary producers Agriculture and Sustainability Background The metabolic processes of animals produce wastes that are harmful and must be removed. In simple animals, this is accomplished by diffusion alone, but more complex animals use special organs or organ systems. The principal metabolic wastes are carbon dioxide (CO2), formed by cellular respiration, and nitrogenous wastes, produced by the breakdown of amino acids composing proteins. Most animals use respiratory organs to expedite the removal of CO2 and have special excretory organs to get rid of nitrogenous wastes. When amino acids are broken down, the amine groups tend to form toxic ammonia. This substance diffuses from small, simple animals, but diffusion is inadequate for removal of amine groups in larger, more complex forms. In aquatic vertebrates, nitrogenous wastes, including ammonia are flushed from the body with large amounts of water. Terrestrial forms, which must conserve water, convert ammonia into urea and uric acid. Urea, the primary nitrogenous waste in amphibians and mammals, is secreted in aqueous solution, but uric acid, the primary nitrogenous waste in insects, reptiles, and birds, is excreted as a semisolid mass with relatively little water. The process of urinary excretion not only removes the nitrogenous waste products of metabolism but also regulates the concentration of ions, water, and other substances in body fluids. The urinary systems of marine, freshwater, and terrestrial vertebrates are each uniquely adapted to perform these functions in radically different environments. What to Do and How to Do It (Students) PART I Nitrogen and the human perspective – Urinalysis In humans, the composition of urine is commonly used to assess the general functioning of the body. Diet, exercise, and stress may cause variations in composition and concentration, but significant deviations usually result from malfunctions of the body. Materials Needed: Biohazard bag Collecting cup, plastic Multi-test reagent strips Simulated urine samples Test tubes and holder In this section you will use urinalysis dip sticks to analyze several simulated urine samples to determine if they are normal or if their characteristics suggest Reformatted from Gunstream labs 22 and 39 Page 1 possible disease. Table 1 indicates normal values and some abnormal characteristics. Study it before proceeding. 4/8/2002 1. 2. 3. 4. 5. Obtain six reagent strips (dipsticks) and a color chart that indicates how to read the results. Study the color chart carefully to be sure that you understand the time requirements for reading the dipsticks and the location and interpretation of each reagent band. Obtain six numbered test tubes containing simulated urine samples. Place them in a test tube rack at your workstation. Analyze the urine samples one at a time by dipping a dipstick completely into the sample so that all reagent bands are immersed. Remove the dipstick and place it on a paper towel with the reagent bands facing upward. Read your results after one minute. OPTIONAL - If you wish to test your own urine, obtain a plastic collecting cup and a reagent strip from your TA and perform the test at home. Wash your hands with soap and water. Read your results. Record your results on the assessment sheet, page 6. Component Normal Abnormal (Not all possibilities are listed) Color Straw to amber Turbidity Clear to slightly turbid pH 4.8 to 8.0 (average 6.0) Specific Gravity 1.003 to 1.035 Blood or hemoglobin Absent Protein Absent or trace Glucose Ketones Absent or trace Absent or trace Bilirubin Small amounts Pink, red-brown, or smoky urine may indicate blood in the urine. The higher the specific gravity, the darker the color. Nearly colorless urine may result from excessive fluid intake, alcohol ingestion, diabetes insipidis, or chronic nephritis. Excess and persistent turbidity may indicate pus or blood in the urine. Acid urine may result from a diet high in protein (meats and cereals) or from a high fever; alkaline urine results from a vegetarian diet or bacterial infection of the urinary tract. Low values result from a deficiency of antidiuretic hormone or kidney damage that impairs water reabsorption. High values result from diabetes mellitus or kidney disease, allowing proteins to enter filtrate. Presence of intact RBCs may result from lower urinary tract infections, kidney disease allowing RBCs to enter filtrate, lupus, or severe hypertension. Presence of hemoglobin occurs in extensive burns and trauma, hemolytic anemia, malaria, and incomplete transfusions. Proteins are present in kidney diseases that allow proteins to enter the filtrate, and they may be present in fever, trauma, anemia, leukemia, hypertension, and other nonrenal disorders. They may occur due to excessive exercise and high-protein diets. Presence usually indicates diabetes mellitus. Presence results from excessive fat metabolism, as in diabetes mellitus and starvation. Excess may indicate liver disease (hepatitis or cirrhosis) or blockage of bile ducts. Presence indicates a bacterial urinary tract infection. Presence indicates a urinary tract infection. Nitrite (bacterial) Absent Leukocytes (pus) Absent Table 1. Urine Components Evaluated by Urinalysis PART II Nitrogen and an environmental perspective Aquatic microecosystems Pollution often results from the by-products of human activities accumulating in the environment at levels that are harmful to living organisms. Sources of pollution are varied. For example, air pollution is usually caused by submicroscopic particles, sulfur oxides and nitrogen oxides produced by the burning Reformatted from Gunstream labs 22 and 39 Page 2 of coal or petroleum products. Lakes and rivers may be polluted by pesticides and fertilizers in water runoff from agricultural land, contaminated wastewater from mining operations or chemical plants, and sewage from cities. Whatever the type and source of pollution, it overloads the natural 4/8/2002 processes of an ecosystem and damages its delicate balance by altering one or more populations within the community of organisms. Its effect may be sudden and dramatic or gradual and subtle. In this section you will analyze aquatic microecosystems set up in the laboratory to simulate three lake environments: (1) Normal, (2) polluted with acid rain, and (3) polluted with organic materials. Your task is to compare these ecosystems to ascertain the effect of acid and organic pollution on the diversity and density of organisms. Acid rain is a side effect of air pollution. Sulfur and nitrogen oxides react with water in the atmosphere to form sulfuric acid and nitric acid, respectively, which are returned to Earth in rain. Rainwater with a pH of less than 5.6 is considered to be acid rain. Lakes and ponds exposed to acid rain have a lowered pH, which adversely alters their biogeochemical cycles, decreasing the availability of nutrients. Lakes and ponds normally undergo a process called eutrophication, which gradually increases the available nutrients. Eutrophication over many years, leads to changes in the community of organisms within an ecosystem and, ultimately, leads to the conversion of a pond to a meadow. However, the process is so slow that it is seldom noticed within a person’s lifetime. In contrast to the slow process of eutrophication, organic pollution of lakes and ponds by fertilizers or sewage causes a sudden increase in the available nutrients, which alters the ecosystem by favoring some members of the community while harming others. A polluted aquatic ecosystem tends to have a reduced concentration of oxygen, which may kill off desirable species such as gamefish. Materials Needed: Aquatic ecosystems in small aquaria simulating (1) normal, (2) acid pollution, and (3) organic pollution. Small beaker Algae key Pipettes Slides with cover slips 1. Observe the three microecosystems macroscopically to determine the relative density of autotrophs. Do this by noting the degree of clarity or greening color of the water. Record your observations on the assessment sheet, page 7. 2. Use figure 1 to learn the recognition characteristics of the crustaceans; Gammarus, Cyclops, and Daphnia. Gammarus is the largest form and tends to stay near the bottom of the container. Daphnia is intermediate in size and swims in the water by pulsating beats of the antennae, as it engulfs microscopic photosynthetic organisms. Cyclops is the smallest and will be either swimming in the water or attached to the filamentous algae or to the sides of the container; all these crustaceans can be located with the unaided eye or with a hand lens. 3. Locate the crustaceans in the microecosystem. Note the type and relative density of the crustaceans in each system. Record your observations on the assessment sheet, page 7. 4. Learn the recognition characteristics of the autotrophs in the microsystems by studying the diagrams and the demonstration slides provided. 5. Determine the autotrophs in each microecosystem by making slides of water from Figure 1. Heterotrophs Reformatted from Gunstream labs 22 and 39 Page 3 4/8/2002 each microecosystem and observing them with your microscope. If filamentous algae are present, remove a small amount with forceps and mount it on a slide for observation. Most of the autotrophs are very small. Use the 10X objective to locate a specimen, then switch to the 40X objective to identify it. Use reduced illumination for best results. It may be necessary to male many slides to locate specimens in the microecosystem with the lowest density of autotrophs. Record your results on the assessment sheet, page 7. Figure 2. Autotrophs Reformatted from Gunstream labs 22 and 39 Page 4 4/8/2002 PART III Testing for ammonia, nitrates and nitrites (Standardized test kits using specific reagents and color keys) Instructions will be provided in class. The Nitrogen Cycle Vocabulary and Glossary Acidic Cellular respiration Alkaline Diffusion Ammonia Diuretic Autotroph Eutrophication Bilirubin Filtrate Heterotroph Ketones Metabolic Nitrate Nitrite Reagent Turbidity Urea Uric acid Suggested Reading Campbell Chapter 20: Pages 460-470 References and Resources Gunstream, Stanley E. 1996. Explorations in Basic Biology. Prentice Hall, New Jersey. Harmful Algal Blooms Event around the globe http://www.redtide.whoi.edu/hab/notedevents/notedevents.html Marine Notes Uncommon Blooms: The Nitrogen Factor ftp://ftp.mdsg.umd.edu/pub/MDSG/MarNotes/MN16_3.PDF Red Tide Maps http://www.redtide.whoi.edu/hab/ Reformatted from Gunstream labs 22 and 39 Page 5 4/8/2002 Lab 13 – Nitrogen Cycling Assessment Sheets Name ______________________ Section _____________________ PART I - Urinalysis Record the results of your analysis of the simulated urine samples in the table below. Use an X to indicate the presence of a component and record the color and specific gravity. Tubes Characteristic 1 2 3 4 5 6 Glucose Bilirubin Ketone Specific Gravity Blood PH Protein Nitrite Color Indicate for each urine sample the health condition of the “patient” based on your urinalysis. Select the health condition from the list that follows: Tube 1 ____________________________________________________________________ Tube 2 ____________________________________________________________________ Tube 3 ____________________________________________________________________ Tube 4 ____________________________________________________________________ Tube 5 ____________________________________________________________________ Tube 6 ____________________________________________________________________ Diabetes insipidis This disorder is caused by a deficiency of antidiuretic hormone, which results in the inability of kidneys to reabsorb water. Symptoms are constant thirst, weight loss, weakness, and production of 4-10 liters of urine each day. The urine is very dilute (a low specific gravity) and is nearly colorless. Diabetes mellitus This disorder is caused by a deficiency of insulin, which results in a decreased ability for glucose to enter cells. Therefore, glucose accumulates in the blood and fats are used excessively in cellular respiration, producing ketones as a waste product. Symptoms include weakness, fatigue, weight loss, and excessive blood glucose levels. In uncontrolled diabetes, a urine sample contains excess glucose and ketones and has a low pH. Hepatitis Hepatitis is usually caused by a viral infection. In type A hepatitis, symptoms may include fatigue, fever, generalized aching, abdominal pain, and jaundice (yellowing tinge to the whites of the eyes due to excessive blood levels of bilirubin). Urine is dark amber in color due to excessive bilirubin. Glomerulonephritis In this kidney disease, excessive permeability allows proteins and red blood cells to enter the filtrate. They are present in the urine since they cannot be reabsorbed. Hemolytic anemia A number of conditions cause the destruction of red blood cells, thereby releasing hemoglobin into the blood plasma. Such conditions include malaria and incompatible blood transfusions. The urine of such patients contains hemoglobin and may be red-brown or smoky in color. Reformatted from Gunstream labs 22 and 39 Page 6 4/8/2002 Normal Concentrations of dissolved substances in normal urine (mg/100ml) Urea 2000 Uric Acid 50 Inorganic salts 1500 Protein 0 Amino Acids 0 Glucose 0 Strenuous exercise and a high-protein diet Athletes in excellent health may produce urine with an acid pH, a small amount of protein, and an elevated specific gravity due to the presence of proteins. Urinary tract infection Bacterial urinary tract infections may involve structures of the urinary tract including the kidneys. Production of alkaline urine promotes the growth of infectious bacteria. Urethritis (infection of the urethra) often causes a burning pain during urination. Symptoms may include a low-grade fever and discomfort of the affected region. Urine will test positive for nitrite because bacteria in the urine convert nitrate, a normal component, into nitrite. Severe infections may also cause the urine to contain blood and pus (leukocytes), producing cloudy urine. PART II - Aquatic Microecosystems Based on macroscopic observations, which microecosystem seems to have: The greatest density of autotrophs? (darkest green) _____________________________________________ The lowest density of autotrophs? (lightest green) ______________________________________________ The greatest density of heterotrophs? ________________________________________________________ The lowest density of heterotrophs? _________________________________________________________ Based on macroscopic observations, record the relative abundance of each kind of crustacean in the microecosystems in the table below. Use this scale: 0=none; 5=most abundant. Based on examination of your slides, record the presence of autotrophs in the Microsystems by placing an “x” in the appropriate spaces in the table below. Compare your data with the class data. Organism Acid Pollution Normal Organic Pollution Cyanobacteria Oscillatoria Protista Euglena Peridinium Green Algae Microspora Pandorina Spirogyra Crustacea Cyclops Daphnia Gammarus Reformatted from Gunstream labs 22 and 39 Page 7 4/8/2002