Unit 3 Activity Workbook

Unit 3 Activity Workbook - Unit Three Activity Cell...

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Unformatted text preview: Unit Three Activity Cell Structure ACTIVITY #3 Name: ________________________________________ The cell is the functional basic unit of life. It was discovered by Robert Hooke and is the functional unit of all known living organisms. It is the smallest unit of life that is classified as a living thing, and is often called the building block of life. Some organisms, such as most bacteria, are unicellular. Other organisms, such as humans, are multicellular. Humans have about 100 trillion or 1014 cells; a typical cell size is 10 µm and a typical cell mass is 1 nanogram. The longest human cells are about 135 µm and can reach from the toe to the lower brain stem in the anterior horn in the spinal cord, while granule cells in the cerebellum, the smallest, can be some 4 µm. The largest known cells are unfertilized ostrich eggs, which weigh 3.3 pounds. In 1835, before the final cell theory was developed, Jan Evangelista Purkyně observed small "granules" while looking at the plant tissue through a microscope. The cell theory, first developed in 1839 by Matthias Jakob Schleiden and Theodor Schwann, states that all organisms are composed of one or more cells, that all cells come from preexisting cells, that vital functions of an organism occur within cells, and that all cells contain the hereditary information necessary for regulating cell functions and for transmitting information to the next generation of cells. Modified from Wikipedia (http://en.wikipedia.org/wiki/Cell_(biology)) Goals: o o o o o o o o o Use a microscope properly in the investigation of cells Understand the process of transport through the cell membrane Understand the structure and function of the cell membrane Understand the limitations of cell size based on the ability to diffuse nutrients into the cell Understand the basic differences in the various cell types Understand the limitations of a light microscope as it regards viewing of cellular components Use dialysis tubing to model diffusion across the cell membrane Investigate the influence of solute concentrations on osmosis Investigate the concept of water potential in relation to water movement into or out of plant cells Unit Two Activity Cell Structure Page 1 Laboratory 3 – A: Surface Area to Volume Ratio: Cells are limited in how large they can be. This is because the surface area and volume ratio does not stay the same as their size increases. Because of this, it is harder for a large cell to pass materials in and out of the membrane, and to move materials through the cell. In this lab, you will evaluate how much fluid will infuse into two different sized cells. First you will calculate the volume of the model cells and the surface area of the model cells and determine the ratio of one to the other. Then you will immerse your model cells into a solution that will create a pink indicator in the model cells to indicate the amount it has diffused into the model cell. Materials: • • • 500 mL beaker Approximately 200 mL of alkaline solution large cell model • • • • Small cell model 1000 mL graduated cylinder Calculator Paper towels Cautions: • Alkaline solutions are caustic and can harm sensitive tissue Procedure: Obtain the surface area for the large cell model and record it in the table below Obtain the surface area for the small cell model and record it in the table below Using your 1000mL graduated cylinder and the displaced volume technique, obtain the volume of the large cell model and record that in the table below Using the same technique, obtain the volume of the small cell models and record that in table below Calculate the surface area to volume ratio for each by dividing the surface area by the volume and enter that value in the table below Unit Two Activity Cell Structure Page 2 1. Predict which model will allow the greater percentage of liquid to infuse in Put both cell models into the 500 mL beaker Pour alkaline solution into the beaker until both models are completely covered Allow the models to sit in the alkaline solution for 15 minutes exactly After 15 minutes remove both cell models, flush them with water for about a minute, and pat dry with a paper towel Cut both models in half Calculate the percentage of your model that the alkaline solution penetrated as follows: o To obtain the amount of the cell model that was not penetrated by the alkaline solution, subtract the amount of penetration (A’ + B) from the total length (C) o Divide the amount of the cell model not penetrated by the total length and multiply by 100. This gives you the percent of the cell model that was not penetrated by the alkaline solution o To calculate the percent of the cell model that was penetrated, subtract the percent of the cell model that was not penetrated by the alkaline solution from 100. Enter the percentage the alkaline solution penetrated into the table below MODEL SURFACE AREA SURFACE AREA/VOLUME (RATIO) VOLUME PERCENTAGE PENETRATION LARGE 2 cm mL SMALL 2 cm mL Unit Two Activity Cell Structure Page 3 2. What was the rate of penetration for each cell model? (Percentage Penetration/15 minutes) a. Rate of penetration for the large cell model _____ Percent per minute b. Rate of penetration for the small cell model _____ Percent per minute 3. Was the rate of penetration the same or different for both models? Explain. 4. Was the Percentage Penetration the same or different for both models? Explain. Unit Two Activity Cell Structure Page 4 5. Does the surface area to volume ratio correspond more closely with the rate of penetration or the Percentage Penetration? Explain. Laboratory 3 – B: Cell Observation: Staining is a technique used in microscopy to enhance contrast in the microscopic image. Stains and dyes are frequently used in to highlight cellular structures for viewing, often with the aid of different microscopes. Stains may be used to define and examine tissues like muscle fibers or connective tissue, cell populations, or organelles within individual cells. Staining and fluorescent tagging can serve similar purposes. Biological staining is also used to mark cells in flow cytometry, and to flag proteins or nucleic acids in gel electrophoresis. Observing biological specimens involves several techniques, not the least of which is microscopy itself. Proper knowledge, operation, and maintenance of your microscope is vital in microscopy. Microscopes are expensive and sensitive pieces of equipment that require careful handling and routine cleaning. In addition, knowledge of the stains and protocols used in microscopy is necessary. The first protocol in microscopy is mounting, a process that usually involves attaching the samples to a glass microscope slide for observation and analysis. In some cases, cells may be grown directly on a slide. For samples of loose cells (as with a blood smear or a pap smear) the sample can be directly applied to a slide. For larger pieces of tissue, thin sections (slices) are made using a microtome; these Specimens stained for microscopy slices can then be mounted and inspected. There are two main types of mounts, the wet mount and the fixed mount. Due to the poisonous Unit Two Activity Cell Structure Page 5 and disruptive nature of staining, a fixed mount is used for most staining protocols. This involves attaching your specimen to a slide and heat fixing the specimen either before or after immersing your specimen in stain. At its simplest, the actual staining process may involve immersing the sample (before or after fixation and mounting) in dye solution, followed by rinsing and observation. Many dyes, however, require the use of a mordant: a chemical compound which reacts with the stain to form an insoluble, colored precipitate. When excess dye solution is washed away, the mordanted stain remains. A simple staining method for bacteria which is usually successful even when the "positive staining" methods fail, is to employ a negative stain. This can be achieved simply by smearing the sample on to the slide, followed by an application of nigrosin (a black synthetic dye) or Indian ink (an aqueous suspension of carbon particles). After drying, the microorganisms may be viewed in bright field microscopy as lighter inclusions well‐contrasted against the dark environment surrounding them. Gram staining is used to determine gram status to classify bacteria broadly. It is based on the composition of their cell wall. Gram staining uses crystal violet to stain cell walls, iodine as a mordant, and a fuchsin or safranin counterstain to mark all bacteria. Gram status is important in medicine; the presence or absence of a cell wall will change the bacterium's susceptibility to some antibiotics. Gram‐positive bacteria stain dark blue or violet. Their cell wall is typically rich with peptidoglycan and lacks the secondary membrane and lipopolysaccharide layer found in Gram‐ negative bacteria. On most Gram‐stained preparations, Gram‐negative organisms will appear red or pink because they are counterstained. Due to presence of higher lipid content, after alcohol‐treatment, the porosity of the cell wall increases, hence the CVI complex (Crystal violet ‐Iodine) can pass through. Thus, the primary stain is not retained. Also, in contrast to most Gram‐positive bacteria, Gram‐negative bacteria have only a few layers of peptidoglycan and a secondary cell membrane made primarily of lipopolysaccharide. Types of stains and their use is varied. Some stains we may use include the Romanowsky stains, based on a combination of eosinate and methylene blue, which is used to examine blood or bone marrow samples. This technique is used to examine blood to detect malaria, a blood‐borne parasite. Sudan staining uses Sudan dyes to stain lipids. This technique is used to examine fecal fat to diagnose steatorrhea. Crystal violet when combined with a suitable mordant stains cell walls (an important component in Gram stains). Acid fuchsine is used to stain collagen, smooth muscles, or mitochondria. Iodine is used as an indicator for starch. Starch is a substance common to most plant cells, so a weak iodine solution will stain the presence of starch in these cells. Unit Two Activity Cell Structure Page 6 Lugol’s iodine (IKI) is a solution that is used to enhance the nuclei of cells (used as a mordant in Gram stain). Malachite green is used as a counterstain to safranin in the Gimenez staining technique for bacteria and can also be used to stain spores. Methylene blue is used to stain animal cells like human cheek cells making their nuclei more observable. Neutral red is used as a counterstain with other dyes. Safranin is also a nuclear stain producing red nuclei it also stains collagen yellow (used as a counterstain in Gram stain). Gram Stain Gram staining (or Gram's method) is a method of differentiating bacterial species into two large groups (Gram‐positive and Gram‐negative) based on the chemical and physical properties of their cell walls. The Gram stain is almost always the first step in the identification of a bacterial organism. The word Gram is always spelled with a capital, referring to Hans Christian Gram, the inventor of Gram staining. Gram stains are performed on body fluid or biopsies when bacterial infection is suspected. It is a relatively quick way to begin the process of differentiation, a step by step process that separates a bacteria sample into increasingly smaller groups until the specific species can be identified. Cerebrospinal fluid can be examined to check for meningitis, synovial fluid, or septic arthritis using this method. After identifying the bacteria’s Gram stain reaction, a physician would use other special chemicals and growth media to further differentiate the species. Gram‐positive Gram‐ i Gram‐positive bacteria have a thick mesh‐like cell wall made of peptidoglycan which stains purple while Gram‐negative have a thinner layer of peptidoglycan which stains pink. Gram‐ negative bacteria also have an additional outer membrane which contains lipids, and is separated from the cell wall by the periplasmic space. Gram positive bacteria include the phyla Firmicutes which includes many well‐ known genera such as Bacillus, Listeria, Staphylococcus, Streptococcus, En terococcus, and Clostridium. Gram negative bacteria include proteobacteriam, cyanobacteria, spirochaetes, green sulfur and green non‐sulfur bacteria. Gram‐negative bacteria also include many medically relevant Gram‐ negative cocci, bacilli and many bacteria associated with nosocomial infections. Unit Two Activity Cell Structure Page 7 Materials: • • • • • • • Microscope Lens paper and cleaning solution 3 microscope slides Lens oil Inoculation loop Cloths pin Abiotic solution • • • • Gram stain reagents (dyes‐ crystal violet, Gram’s iodine, 95% ethyl alcohol, and safranin) Bunsen Burner, hose, and striker Bacteria cultures Paper towel Cautions: • • • • The stains you will use will stain skin and clothing The alcohol you will use can burn sensitive tissues You will use a Bunsen burner, handle the flame with caution Bacterial cultures require following aseptic techniques Procedure: Obtain a microscope and gently clean the microscope. Obtain your slides and clean them if necessary (preferable to have new slides) Clean your work area with abiotic solution then wash your hands Set up your Bunsen burner, light it, and adjust the gas and air to achieve a blue flame Sterilize your inoculation loop and allow to cool, about 5 seconds. Place one loop full of distilled water on your slide Sterilize your loop again before obtaining the bacteria sample Gently obtain a sample of bacteria from your culture using your inoculation loop Place your sample on your microscope slide Allow your sample to air dry When air dried, fix the sample by passing your sample over the Bunsen burner flame three or four times. Use the clothes pin to hold your slide so as not to get burned. Cover with CRYSTAL VIOLET for 20 seconds. (PRIMARY STAIN) Gently rinse off the stain with water and shake off the excess under the faucet at low pressure. Cover with GRAM'S IODINE (MORDANT) for one minute then pour off the Gram's iodine. Angle your slide over the sink and run 95% ETHYL ALCOHOL (DECOLORIZING AGENT)down the slide until the solvent runs clear (about 10‐20 seconds). THIS STEP IS CRITICAL! THICK SMEARS REQUIRE MORE TIME THAN THIN ONES. Rinse with water to stop the action of the alcohol. Cover with SAFRANIN (COUNTER STAIN) for 20 seconds. Unit Two Activity Cell Structure Page 8 Gently rinse off the stain with water. Gently blot with paper towel and clean off the bottom of the slide with 95% alcohol. o HELPFUL SUGGESTIONS: DO NOT make your smears too thick! Be very careful when you decolorize. Be sure your cultures are young, preferably less than 24 hours old. Older cultures tend to lose the ability to retain stains. 6. What specimen did you receive? 7. Observe your smears in the microscope under oil immersion. Draw a few representative organisms from your smear below. 8. Is your specimen a Gram‐negative or a Gram‐positive species? Explain how you know. Unit Two Activity Cell Structure Page 9 9. Choose a Gram‐negative bacteria that is a human pathogen and list the name, disease caused, and symptoms below: Laboratory 3 – C: Osmosis Lab Preparation: To conduct the following laboratories we must prepare a 1M solution of sucrose and 0.2 M, 0.4 M, 0.6 M, and 0.8 M dilutions of that solution. Begin by answering the following questions. 10. What is the molecular formula for sucrose? (remember sucrose is a disaccharide formed by a dehydration reaction between glucose and fructose) Unit Two Activity Cell Structure Page 10 11. What is the formula weight of sucrose? (Review the on‐line tutorial at http://www.wisc‐online.com/objects/ViewObject.aspx?ID=GCH7004. We will calculate carbon in class) Atoms atomic mass x the number of atoms of that element in the molecule = mass for that element a. b. Hydrogen ‐ _____ x _____ = _____ c. Oxygen ‐ _____ x _____ = _____ d. 12. Carbon ‐ _____ x _____ = _____ Formula weight of sucrose = Define Avogrado’s number 13. Define a mole 14. Define molar mass 15. What is the molecular mass of sucrose? Unit Two Activity Cell Structure Page 11 Materials: • • • Sucrose Large Erlenmeyer flask Funnel • • • 1000 mL graduated cylinder Balance 5 jars with caps Cautions: There are no cautions in this portion of the lab, always use proper precautions in a laboratory Procedure: Using the information you obtained above, dissolve the appropriate amount of sucrose (the grams indicated in the molecular mass g/mol) into 500 mL of distilled water in Erlenmeyer flask and mix it well Use a funnel and pour the solution into a 1000 mL graduated cylinder Add distilled water to bring the volume in the cylinder up to 1000 mL or 1 liter. Now you have a 1 molar sucrose solution. 16. Mathematically figure the following dilutions of the 1 M solution: (X mL of solution diluted with distilled water) 1000 ml X ‐‐‐‐‐‐‐‐‐‐ = ‐‐‐ 1 M 0.5 M (Put X ml in a 1000 mL graduated cylinder and fill to 1000 mL with distilled water. Substitute the 0.5 M with the molar concentration you are making.) a. 0.0 M – b. 0.2 M – Unit Two Activity Cell Structure Page 12 c. 0.4 M – d. 0.6 M – e. 0.8 M – Check your calculations with the instructor If the calculations are correct, create the dilutions and store them in a jar with the lid secured Write your name and the molarity on each jar and put the jar in the refrigerator Laboratory 3 – D: Diffusion: The cell membrane is a cell’s interface with its surroundings. In one sense, this membrane must function as a barrier: it must keep together in one bundle the enzymes, DNA, and metabolic pathways that make life possible. The cell membrane must also function as a gateway: waste products must be discharged through it and essential materials (oxygen, water, etc.) must enter through it. A membrane that allows some molecules to pass through while blocking the passage of others is said to be semipermeable. Molecules pass through the cell membrane either through processes that require the cell to expend energy (active transport), or through processes driven by the kinetic (thermal) energy of molecules (passive transport). In these lab activities, you will investigate the passage of materials through a semipermeable membrane by passive transport. The membrane you will use, dialysis tubing, is semipermeable because it has submicroscopic holes through it. Molecules are in constant random motion. By chance, a molecule’s motion may move it toward the membrane (Figure 1). If it collides with the Unit Two Activity Cell Structure Page 13 membrane wall, it rebounds. If its motion takes it toward a pore, it may either pass through the pore, or it may rebound, depending upon the size of the molecule relative to the diameter of the pore. Molecules that are small enough to pass through the pores can pass through in either direction. Notice that on one side of the membrane solute molecules have displaced some of the water molecules. Thus, there is a higher concentration of water molecules on the opposite side of the membrane. More water molecules are available to collide with the membrane on the side having the higher concentration of water. Thus, although water molecules will move in both directions across the membrane, more will move from the side having the higher concentration to the side having the lower concentration. The movement of molecules from areas of higher concentration to areas of lower concentration is called diffusion. The diffusion of water molecules across a semipermeable membrane is termed osmosis. A process that depends upon random motion might seem inefficient, but so many water molecules are involved and they move so fast, that it is estimated that a red blood cell floating in blood plasma gains an amount of water equal to 125 times its own volume every second. It also loses the same amount of water each second, all by osmosis. This occurs because the concentration of solutes in the blood plasma is the same as the concentration of solutes in red blood cells. Solutions that have the same solute concentration are isotonic. If we took a sample of whole blood and added salt to the plasma, increasing its solute concentration, the plasma becomes hypertonic to the solution in the red blood cells, and the cells lose water and shrink. If we add water to the blood plasma, decreasing its solute concentration, the plasma becomes hypotonic to the solution in the red blood cells. The cells gain water, swell, and may even burst. In this laboratory you will explore the diffusion of different molecules through dialysis tubing, a semipermeable membrane. You will use glucose test strips to check for the presence of glucose and I2KI solution to test for the presence of starch. As you probably know, I2KI reacts with starch to give a dark blue, almost black color. Materials: • • • • Dialysis tubing Plastic cup Glucose/starch solution Distilled water • • • • Iodine‐potassium iodide (I2KI)solution Dropping pipet Glucose test strips Funnel Caution: I2KI solution can irritate the skin, mouth, and eyes, and can stain skin or clothing. Unit Two Activity Cell Structure Page 14 Procedure: Pour 160–170 mL of distilled water into a plastic cup. Add approximately 4 mL of I2KI solution to the water and mix well. Record the initial solution color in Table 1 below. Dip a glucose test strip into the solution and record the initial glucose test results in Table 1 below. Use the + symbol to indicate a positive test result for glucose and the – symbol to indicate a negative result. Discard the used glucose test strip. Dip a fresh glucose test strip into the glucose/starch solution. Record the initial results in Table 1. Discard the used glucose test strip. Obtain a piece of dialysis tubing that has been soaked in water. The tubing should be soft and pliable. Roll the tubing between your thumb and index finger to open it. Close one end of the tube by knotting it or tying it off with string. This will form a bag. Using a small funnel, pour 15 mL of glucose/starch solution in the dialysis bag. Smooth out the top of the bag by running it between your thumb and index finger to expel the air. Tie off the open end of the bag making sure to leave enough room in the bag to allow for expansion. Record the initial color of the glucose/starch solution in Table 1. Immerse the dialysis bag in the solution in the cup. Make sure that the portion of the bag that contains the glucose/starch solution is completely covered by the solution in the cup at all times. Wait 30 minutes 17. Indicate on the figure below the initial locations (inside or outside of the bag) of all the kinds of molecules that are available for diffusion through the dialysis membrane. 18. For each of the molecules you list on this figure, predict their direction of net (overall) diffusion and give a reason for each prediction [Into the bag, out of the bag, both into and out of the bag equally, or none (will not diffuse across the dialysis membrane)] Unit Two Activity Cell Structure Page 15 After 30 minutes, remove the bag from the cup, blot it on paper towel, and cut a slit in the bag large enough to insert a glucose test strip. Fill in the final columns of Table 1. 19. Compare your results with your predictions. Do you find any conflicts that would cause you to revise your predictions? If so, explain. 20. Does this activity account for the diffusion of all the molecules that you listed on the figure above and in your predictions? If not, what data could have been collected to show the net direction of diffusion of this molecule or molecules? Unit Two Activity Cell Structure Page 16 21. What does your data tell you about the sizes of the molecules relative to the pore size of the dialysis tubing? Laboratory 3 – E: Osmosis: In this portion of the laboratory you will investigate the influence (if any) of solute concentration on the net movement of water molecules through a semipermeable membrane. The solute you will use is sucrose (cane or table sugar) in the following molar concentrations: 0.0 M (distilled water), 0.2 M, 0.4 M, 0.6 M, 0.8 M, and 1.0 M Materials • • • • Dialysis tubing Plastic cups Distilled water Funnel • • • • Sucrose solutions Paper towels Balance Computer Cautions: • There are no cautions for this lab, as always; follow proper lab protocol Procedure Complete the following steps for each sucrose solution that you are assigned to test. Pour 160–170 mL of distilled water into a plastic cup. Label the cup with the concentration of sucrose that you will test. Unit Two Activity Cell Structure Page 17 Obtain a piece of dialysis tubing that has been soaked in water. The tubing should be soft and pliable. Roll the tubing between your thumb and index finger to open it. Close one end of the tube by knotting it or tying it off with string. This will form a bag. Using a small funnel, pour 25 mL of sucrose solution into the dialysis bag. Smooth out the top of the bag, running it between your thumb and index finger to expel the air. Tie off the open end of the bag. Leave enough room in the bag to allow for expansion. Dry the bag on paper towels and then determine the mass. Record this as the initial mass in the Biology Unit 3 Activity Workbook – EXCEL spreadsheet on the class website and enter the data in Laboratory 3 – E. Immerse the dialysis bag in the water in the cup. Make sure that the portion of the bag that contains the sucrose solution is completely covered by the water in the cup at all times. Wait 30 minutes before continuing to the next step. After 30 minutes, remove the bag from the cup and dry it on paper towels. Mass the bag and record the final mass in the Biology Unit 3 Activity Workbook – EXCEL spreadsheet on the class website and enter the data in Laboratory 3 – E. Obtain and record the percent change in mass for all of the lab groups as well as your own and enter that into in the Biology Unit 3 Activity Workbook – EXCEL spreadsheet on the class website and enter the data in Laboratory 3 – E. 22. Save your AP Biology Unit 3 Workbook – EXCEL using the following convention: File name: //213x/apbio/AP Biology Unit 3 Activity Workbook – EXCEL – insert your name here 23. What does the change in mass indicate? 24. Write a hypothesis that this experiment is designed to test. Unit Two Activity Cell Structure Page 18 25. What variable is being tested in this experiment? 26. List at least three variables (other than the tested variable) that could influence the outcome of this experiment. Briefly describe the method of control used for each of these variables. 27. The Excel program created two graphs, the percent change in mass for your group and for the class. Based on the graphs created answer the following: a. The independent variable is: b. The dependent variable is: 28. Explain any differences between the graph for your group and the graph of class averages. Unit Two Activity Cell Structure Page 19 29. On the basis of your data and graph, has this experiment adequately tested the experimental variable you listed above as the variable you were testing? 30. Do your results support your hypothesis, refute it, or require that you modify it? 31. On the basis of your results, write a statement that expresses the relationship of solute concentration and direction of net movement of water molecules in osmosis. 32. In which, if any, of the experimental setups were the solutions in the bag and outside of the bag isotonic to each other? 33. If the experimental setup specified that only distilled water be used to fill the dialysis bag and that the sucrose solutions be used to fill the cup, how would that change your results? Unit Two Activity Cell Structure Page 20 34. When you drink a glass of water, most of it is absorbed by osmosis through cells lining your small intestine. Drinking seawater can actually dehydrate the body. How? Laboratory 3 – F: Water Potential: As you have seen, in animal cells, movement of H2O into and out of a cell is influenced by the relative concentration of solute on either side of the cell membrane. If water moves out of the cell, the cell will shrink or crenate. If water moves into the cell it will swell and may even burst, or cytolyze. In plants, the situation is complicated by the presence of a rigid or semi‐rigid cell wall. Consider a potato cell suspended in pure water. As water enters the cell, the cell membrane is pressed against the cell wall; thus, a physical or hydrostatic pressure develops that opposes the entrance of additional water. When the pressure inside the cell becomes great enough, no additional water can accumulate in the cell, even though the cell still has a higher solute concentration than does pure water. Thus, movement of water through plant tissue cannot be predicted simply by knowing the relative solute concentrations on either side of the plant cell membrane. It is necessary to also account for any differences in pressure between the two sides. The concept of water potential is used to combine the differences in solute concentration and pressure to predict the direction in which water will diffuse through living plant tissues. In a general sense, water potential is a quantification of the tendency of water to diffuse from one area to another under a given set of perimeters. Traditionally, water potential is expressed in bars, a metric unit of pressure equal to 10 newtons per cm2 or about 1 atmosphere. Water potential is abbreviated by the Greek letter psi (Ψ) and has two major components: solute potential (ΨS), which is dependent on solute concentration, and pressure potential (Ψp), which results from the exertion of pressure—either positive or negative—on a solution. Thus: Ψ = Ψp + ΨS Water Potential = Pressure Potential + Solute Potential The water potential of pure water at 1 atmosphere is by definition zero. If we dissolve a substance in the water, the water potential drops below zero. This may seem a bit counterintuitive, but remember that we are talking about the tendency of water molecules to diffuse. This is driven by the kinetic energy of free water molecules. As solute concentration goes up, water concentration goes down, there are fewer water molecules available to diffuse, so water potential must go down. Notice that solute potential must be either zero (for pure water), or negative, if the water contains a solute. (As solute concentration increases, solute potential decreases.) Pressure potential can be Unit Two Activity Cell Structure Page 21 positive, negative, or zero. Although water diffuses in all directions, the net movement of water will always be from an area of higher water potential to an area of lower water potential. Returning to our example of a potato cell suspended in pure water: the system is open to the atmosphere, so Ψp = 0. We will assign a value of –9 forΨS. The initial conditions can then be described as follows: Pure Water Ψ = Ψp + ΨS or Ψ = 0 + 0 = 0 Potato Cell Ψ = Ψp + ΨS or Ψ = 0 + (–9) = –9 Since –9 is less than 0, water will diffuse into the cell. At equilibrium, the following condition exists: Pure Water Ψ = Ψp + ΨS or ψ = 0 + 0 = 0 Potato Cell ψ = ψp + ψS or ψ = 9 + (–9) = 0 Since 0 = 0, there will be no net movement of water into or out of the cell. Suppose we add solute to the water in the cup until we produce the following initial conditions: Water + Solute Ψ = Ψp + ΨS or Ψ = 0 + (–15) = –15 Potato Cell ψ = ψp + ψS or ψ = 0 + (–9) = –9 Since –15 is less than –9, water will diffuse out of the cell. If this continues, the cell will shrink and the cell membrane may pull away from the cell wall, a condition known as plasmolysis. This often injures the cell and may kill it. Unit Two Activity Cell Structure Page 22 If we know the molarity of a sucrose solution that will produce equilibrium between the solution and the contents of the potato cell, we can determine the solute potential by using the following formula. ΨS = –iCRT In the formula ΨS = –iCRT, the variables are: • • • • i = ionization constant (for sucrose, this value is 1) C = molar concentration of sucrose per liter at equilibrium (must be determined experimentally) R = pressure constant (0.0831 liter bar/mole K) T = temperature of solution in kelvins (K = °Celsius + 273) Example: If C is experimentally determined to be 0.300 and T is 293 K, then ΨS = –(1)(0.300 mole/liter)(0.0831 liter bar/mole K)(293 K) ΨS = –7.304 bars In this activity, you will investigate water potential by immersing potato cores in sucrose solutions and determining the change in mass, if any, of the cores. You will graph your data and use the graph to determine a value for C. Using the experimentally determined value for C, you will then calculate a value for ΨS. Your group will be assigned one or two sucrose solutions to test. Materials • • • • • • Plastic cups Distilled water Sucrose solutions Cork borer Knife or scalpel Potato • • • • • Plastic wrap Paper towels Balance Computer Thermometer (optional) Caution • The cork borer is sharp Unit Two Activity Cell Structure Page 23 Procedure Label a cup with the concentration of sucrose that you will test. Use a cork borer to cut four cylinders of potato tissue from the potato. Do not stab your hand with the cork borer. Trim both ends of each cylinder to remove the skin. Cut each cylinder into sections that are approximately 3 cm in length. Use caution when slicing the cylinders so you do not injure yourself or others. If you cannot immediately use a balance to mass your cores, place them in a beaker or cup and cover them with a lid or with plastic wrap. Why do you think this is necessary? Use a balance to determine the total mass of all the potato sections. Record this as the initial mass in the Biology Unit 3 Activity Workbook – EXCEL spreadsheet on the class website, enter the data in Laboratory 3 – F, Individual Data for Potato Cores section. Place all of the potato sections in the labeled cup. Pour 100 mL of the assigned sucrose solution into the cup. Cover the cup with plastic wrap. Why do you think this step is necessary? If you have been assigned a second concentration of sucrose to test, repeat steps 1–6 using that solution. Stop. Allow the potato sections to remain in the sucrose solution overnight. Then, proceed to the next step. Continue with the following only after the potato sections have been in the sucrose solution overnight. Perform these steps for each solution you are testing. Record the temperature as given by your teacher 35. Ambient temperature = ________ °Celsius Remove the potato sections from the sucrose solution. Blot them on paper towels to remove excess solution. If you cannot immediately use a balance to mass your cores, cover them as before to prevent evaporation. Use a balance to determine the final mass. Determine the change in mass of the potato sections. Record this data in the Biology Unit 3 Activity Workbook – EXCEL spreadsheet on the class website, enter the data in Laboratory 3 – F, Individual Data for Potato Cores section.. Obtain and record the class averages in the Class Data for Potato Cores section on the Biology Unit 3 Activity Workbook – EXCEL spreadsheet on the class website, enter the data in Laboratory 3 – F. Be sure to save your Excel program as instructed Unit Two Activity Cell Structure Page 24 36. On your completed graph, find the point where the line of your data crosses the 0 line (x‐axis) of the grid. This is the equilibrium point; at this point there is no net gain or loss of water from the potato cells. Read the corresponding value of sucrose molarity for this point. This is the molar concentration of sucrose that produces equilibrium. Below, record this concentration of sucrose as your experimentally determined value for C. Convert ambient temperature from °C to K. a. C = _________ mole/liter b. T = _________ K 37. Using the formula ΨS = –iCRT, calculate the solute potential at equilibrium. Show your calculations in the space below. ψS = _______ bars 38. Using the formula Ψ = Ψp + ΨS, give the following: a. The water potential of the solution at equilibrium _______________ b. The water potential of the potato cells at equilibrium ____________ Unit Two Activity Cell Structure Page 25 39. A chef chops vegetables into a bowl of water. Would you expect the vegetable slices to gain or lose water? Explain your answer in terms of water potential. 40. Imagine that you are an agri‐science consultant to a large corporate farm that raises 7,000 acres of wheat on desert land adjoining the Mediterranean Sea. Just before the wheat matures, all the wells used for irrigation water run dry. The farm manager wants to irrigate the fields with water drawn from the Mediterranean. From previous tests, you know that the average solute potential of root tissue taken from the wheat fields is –11.13 bars. You test the seawater and determine its solute potential to be –24.26 bars. What will you advise the farm manager and why? 41. A marine clam is mistakenly added to a freshwater aquarium. What will happen to the clam and why? Unit Two Activity Cell Structure Page 26 42. A 1 cm diameter, 6 cm long core is removed from a carrot (see diagram). The resulting hole is filled with corn syrup (highly concentrated sugar), and a glass tube is inserted into the hole to make a watertight seal. The carrot is suspended in a cup of pure water. Beginning with the carrot cells adjoining the hole, and going out to the carrot epidermal cells exposed to the water, describe what will happen to the carrot cells. Also, what will happen to the level of liquid in the glass tube and why? Unit Two Activity Cell Structure Page 27 Testing Your Knowledge: • • • • • • • • • • 43. Write the name of the cell part in the box next to its description/function. Cell membrane • Nuclear envelope Centrioles • Nucleolus Chloroplast • Nucleus Chromatin • Peroxisome Cytoplasm • Ribosomes, bound Endoplasmic reticulum, rough • Ribosomes, free Endoplasmic reticulum, smooth • Vacuole Golgi apparatus • Vesicle, secretory Lysosome • Vesicle, transport Mitochondria Unit Two Activity Cell Structure Page 28 44. Indicate if each of the following is true of chromosomes or chromatin or both by placing an A, B, or AB in the line preceding each description. A. Chromosomes B. Chromatin a. ______ Consist of DNA & proteins e. ______ Dispersed b. ______Condensed f. c. ______ Tightly coiled g. ______ Uncoiled ______Decondensed d. ______Visible when stained Unit Two Activity Cell Structure Page 29 45. Determine if each of the following is true of free or bound ribosomes or both by placing a FR, BR, or B on the line in front of the description. FR. Free Ribosomes BR. Bound Ribosomes B. Both a. ______ Produce proteins for use within the cell b. ______ Produce proteins for export c. ______ Attached to rough ER d. ______ Suspended in the cytosol e. ______ Consist of 2 subunits f. ______ Composed of rRNA and proteins 46. Determine if the each of the following is true of microtubules, microfilaments, or intermediate filaments by placing a MT, MF, or IF on the line in front of the description. MT = Microtubules MF = Microfilaments IF = Intermediate filaments a. ______ Straight, hollow tubes b. ______ Made of tubulin c. ______ Involved in cell transport d. ______ Provides tracts for organelle movement e. ______ Made of actin f. ______ Produces cytoplasmic streaming g. ______ Intermediate in diameter h. ______ More permanent Unit Two Activity Cell Structure Page 30 47. Match the cell part with the correct letter from the diagram. ______ Cell membrane ______ Centrioles ______ Chromatin ______ Cytoplasm ______ Golgi ______ Lysosome ______ Mitochondria ______ Nuclear envelope ______ Nucleolus ______ Ribosomes ______ Rough ER ______ Smooth ER 48. Is the cell pictured an animal or plant cell? 49. How do you know? Unit Two Activity Cell Structure Page 31 50. Determine if each of the following characteristics or examples are true of plasmodesmata, tight junctions, desmosomes, or gap junctions. Use the key below to indicate by placing one, two, or all three letters in the correct blanks. P. Plasmodesmata T. Tight Junctions D. Desmosomes G. Gap Junctions a. ______ Found in plant cells b. ______ Found in animal cells c. ______ Form channels between cells d. ______ Allow free passage of water and small solutes between cells e. ______ Found in embryos, cardiac muscle tissue, and endocrine glands f. ______ Hold cells tightly together g. ______ Block intercellular transport of materials h. ______ Found in epithelial layers that separate two kinds of solutions i. ______ Found in the lining of the digestive tract and the blood‐brain barrier j. ______ Rivet cells together k. ______ Well developed in cells subjected to considerable mechanical force l. ______ Found in skin cells m. ______ Permit intercellular transport n. ______ Glycoprotein (intermediate) filaments penetrate and bind plasma membranes of two adjacent cells Unit Two Activity Cell Structure Page 32 51. Match the structure with the correct letter from the diagram. Unit Two Activity Cell Structure Page 33 ...
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