University of Bridgeport Cellular Membranes: Effects of Chemical stress there is pre lab questions need to be answered i will attach the questions and the

University of Bridgeport Cellular Membranes: Effects of Chemical stress there is pre lab questions need to be answered i will attach the questions and the other file is to read it to be able to answer the questions. if u have any questions let me know. BIOL-102 General Biology lab_Kita
Pre-lab 5 (cellular membranes and osmosis)
2019 Fall
All questions are 10 points each unless specified.
1. In exercise 1 (seeing the effect of acetone or ethanol), what are you trying to see (what
is happening on cell membranes by treating with those organic solvents)? Briefly (a few
sentences) explain.
2. Briefly describe the definition of osmosis.
3. Briefly describe the definition of diffusion in biology. You have to mention on cell
membrane (or the plasma membrane).
4. In exercise 2, we will use dialysis tubes. Why? Briefly explain (A few sentences).
5. Related to exercise 2: when you place a tied dialysis tube (filled with 1% sucrose) in
water, what do you expect over time?
6. Related to exercise 2: when you place a tied dialysis tube (filled with 1% sucrose) in
25% sucrose solution, what do you expect over time?
7. Briefly describe the definition of plasmolysis.
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BIOL-102 General Biology lab_Kita
Pre-lab 5 (cellular membranes and osmosis)
2019 Fall
8. When you place red blood cells in water, why do cells explode?
9. Plant cells will not explode under turgid condition. Why? Briefly explain.
10. What is the independent valuable in exercise 2 (5 points), and in graphs, which axis do
we put the independent valuable in general (5 points)?
answer (independent valuable): _______________
answer (axis): __________
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CELLULAR MEMBRANES AND OSMOSIS – LAB 7
Exercise 1:Cellular Membranes: Effects of Chemical stress
Membranes separate and organize the myriad of reactions within cells and allow communication
with the surrounding environment. Although they are only a few molecules thick (6-10 nm),
membranes (l) retard diffusion of selected molecules; (2) house receptor molecules that detect
other cells, organelles, and hormones; (3) provide sites for active and passive transport of
selected molecules; (4) orga-nize life processes by providing surfaces to accommodate chemical
reactions; and (5) help maintain the integrity of cells.
Membranes consist of a phospholipid bilayer; attached to or embedded within this bilayer are
thousands of proteins. A phospholipid molecule consists of a phosphate group and two fatty
acids bonded to a three-carbon, glycerol chain. Phospholipids have an unevenly distributed
charge; that is, they have charged (polar) and uncharged (nonpolar) areas. In phospholipids the
phosphate group and glycerol are polar and hydrophilic (“water-loving”), whereas the fatty-acid
chains are nonpolar and hydrophobic (“water-fearing”). Such molecules with two different
affinities are amphipathic, and amphipathic phospholipids have a natural tendency to selfassemble into a double-layered sheet. In this double layer, the hydrophobic tails of lipids form
the core of the membrane, and the hydrophilic phosphate groups line both surfaces. This elegant
assembly is stable, self-repairing, and resists penetration by most hydrophilic molecules.
Membranes also include proteins dispersed throughout the fluid bilayer of lipids. These proteins
are not fixed in position; they move about freely and may be densely packed in some membranes
and sparse in others. Carbohydrate chains (strings of sugar molecules) are often bound to these
proteins and to lipids. These chains serve as distinctive tags that identify particular types of
cells. These elaborate molecular elements form the fluid mosaic model of membrane structure.
Membranes are selectively permeable. The proteins embedded in the phospholipid bilayer can
selectively take up or expel molecules that otherwise could not penetrate the membrane. In
doing so, these proteins function as pores, permitting and often facilitating the passage of
specific ions and polar molecules. In addition to forming pores and sites for active transport,
membrane-bound proteins also function as enzymes and receptors that detect signals from the
environment or from other cells.
Cellular Membrane- Image modified from OpenStax Biology.
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CELLULAR MEMBRANES AND OSMOSIS – LAB 7
Phospholipid- Image credit: OpenStax Biology.
Observe the Effects of Organic Solvents on Cellular Membranes
Organic solvents dissolve a membrane’s lipids. Acetone and methanol are common solvents for
various organic molecules, but acetone has the greater ability to dissolve lipids.
1. Examine the treatments listed in table 1.
2. Hypothesize which treatments will cause the most and least damage. Note your rankings
alongside the tube numbers in the column marked Tube Number.
3. You will receive prepped, beet sections from your TA.
4. Place one of the seven beet sections into each of seven dry test tubes. Do not crush, stab,
or otherwise damage the cylinders when moving them to the test tubes.
5. Label the tubes 1-7 and write the organic-solvent treat-ment on each tube as listed in
table 1.
6. Add 10.0 mL of the appropriate solvent (see table 2) to each of the seven tubes.
7. Keep all beets at room temperature for 20 min and shake them occasionally. Then
remove and discard the beet sections and measure the extent of membrane damage
according to the amount of betacyanin that diffused into the water.
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CELLULAR MEMBRANES AND OSMOSIS – LAB 7
8. Quantify the relative color of each solution between 0 (colorless) and l0 (darkest red). If
color standards are available in the lab, use them to determine relative values for the
colors of your samples. Record the results for your work group in table 1. Also, provide
your results to the instructor to calculate the class averages.
9. Graph Concentration of Organic Solvent versus Relative Color for the class averages
according to a demonstration graph provided by your instructor.
Table 1: The Color Intensity of Betacyanin from Damaged Cells Treated with Organic
Solvents
Tube Number
Treatment
Work Group
1
2
3
4
5
6
7
Color Intensity
Class Average
1% Acetone
25% Acetone
50% Acetone
1% Methanol
25% Methanol
50% Methanol
Isotonic Saline
Exercise 2: Osmosis: Movement across a concentration gradient
Osmosis is diffusion of water across a differentially permeable membrane. Osmosis follows the
same laws as diffusion but always refers to water, the principal solvent in cells. A solution is a
homogenous, liquid mixture of two or more kinds of molecules. A solvent is a fluid that solves
substances, and a solute is a substance dissolved in a solution.
We can simulate osmosis by using dialysis bags to model cells under different conditions and
measuring the direction and rate of osmosis. Each of the four-dialysis bags in the following
experiment is a model of a cell. Bag A simulates a cell with a solute concentration that is
hypotonic native to its environment. Hypotonic describes a solution with a lower concentration
of solutes, especially those solutes that do not pass across the surrounding membrane. Water
moves across semipermeable membranes out tonic solutions. Conversely, the solutionsurrounding bag A is hypertonic relative to the cell. Hypertonic refers to a solution with a high
concentration of solutes.
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CELLULAR MEMBRANES AND OSMOSIS – LAB 7
Osmosis Across a Concentration Gradient
1. Obtain eight pieces of string and four pieces of soaked dialysis tubing 15 cm long. Seal
one end of each tube by folding and tying it tightly.
2. Open the other end of the tube by rolling it between your thumb and finger.
3. Fill the bags as follows: Bag A 10 mL of 1% Sucrose; Bag B 10mL of 1% Sucrose; Bag
C 10mL of 10% Sucrose; Bag D 10mL of 25% Sucrose. To label each bag, insert a small
piece of paper with the appropriate letter (A, B, C, or D written on it in pencil).
4. For each bag, loosely fold the open end and press on the sides to push the fluid up
slightly and remove most of the air bubbles. Tie the folded ends securely, rinse the bags,
and check for leaks.
5. Gently blot excess water from the outside of the bags and weigh each bag to the nearest
0.1 g.
6. Record these initial weights in the table in the first column.
7. Place bags B, C, and D in three individual beakers or one large bowl filled with 1%
sucrose. Record the time.
8. Place bag A in a beaker and fill the beaker with 150 mL of 25% sucrose. Record the
time.
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CELLULAR MEMBRANES AND OSMOSIS – LAB 7
9. Remove the bags from the beakers at intervals for the next hour (or at intervals indicated
by your instructor), gently blot them dry, and weigh them to the nearest 0.1 g. Handle the
bags delicately to avoid leaks, and quickly return the bags to their respective containers.
10. During the 15-min intervals, use your knowledge of osmosis to make hypotheses about
the direction of water flow in each system (i.e., into or out of bag), and the extent of
water flow in each system (i.e., in which system will osmosis be most rapid?).
11. For each 15-min interval, record the total weight of each bag and its contents in the table.
Then calculate and record in the table the change in weight since the previous weighing.
12. Using excel end create a graph with Total Weight (g) versus Time (min). Total Weight
changed in response to differences in the independent variable, so Total Weight is the
dependent variable. The dependent variable is always graphed on the vertical axis. Time
is the variable that you established and actively controlled and, therefore, is the
independent variable. The independent variable is always graphed on the horizontal axis.
13. Graphs must have a title, correctly labeled axes a label showing measurement units, and
values along each axis.
14. Plot the data for total weight at each time interval from table 2.
15. Include the data for all four bags as four separate curves on the same graph.
Table 2: Weight of Dialysis Bags after Specified Time Intervals
0 Min
15 Min
30 Min
45 Min
60 Min
Initial Total
Change Total
Change Total
Change Total
Change
Weight Weight in
Weight in
Weight in
Weight in
Weight
Weight
Weight
Weight
Bag A
Bag B
Bag C
Bag D
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CELLULAR MEMBRANES AND OSMOSIS – LAB 7
Exercise 3: Using Different Concentrations of Sodium Chloride to Observe
Plasmolysis
In plants, cell walls provide structural support that protects cells from osmotic shock.
Taking such physical property of plant cells as an advantage, we can relatively easily observe
shrinking cytosol under hypertonic conditions. In this experiment, you will place plant cells into
several different “environment”; i.e., different concentrations of solute. Your objective here is to
observe turgid (swollen) and plasmolyzed (shrunken) cells to understand how different osmotic
pressures actually affect cells.
As plant cells have cell walls, turgid cells (cells under hypotonic conditions) would be
indistinguishable from normal cells. However, cells dipped in salt (here, we will use sodium
chloride) solutions higher than physiological conditions, you would be able to observe the cell
membrane pulled back away from the cell wall.
This exercise also contains two additional elements – 1) proper handling of small volume
of liquid with micropipettes 2) Quantification of biological results.
Plasmolyzed Cell
Procedures:
1. Prepare a series of sodium chloride (NaCl) solutions in small (~1.5ml) microcentrifuge tubes.
Use micropipettes and tips. Before preparing solutions, carefully calculate the volume of
distilled water and stock NaCl solution to be mixed.
2. Prepare thin outer epidermal cells from a piece of red onion by peeling off. You may require a
forceps. Alternatively, we can get very thin inner epidermal cells from a piece of onions. If you
end up using inner epidermal cells, we may need to stain cells with 0.1% Neutral Red.
2. Place a small piece of the epidermal cells into each tube. Incubate on the bench for 10
minutes to allow the change of osmotic pressure.
4. Take out a piece and place onto each slide glass. Add a few drop of solution from THE
SAME tube that the sample was submerged. Then place a coverslip to observe cells under a
microscope to see how cytosols look like.
5. Count appropriate number of cells (~20 cells or more) in each condition and record number of
plasmolyzed cells.
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CELLULAR MEMBRANES AND OSMOSIS – LAB 7
6. You can calculate the percentage of plasmolyzed cells as:
Number of plasmolyzed cells
Number of total cells counted
×100%
7. (Option but I highly encourage you to try) Data analysis: For example, plotting, in excel, the
percentage of plasmolyzed cells (y-axis) to a variety of the salt concentrations (x-axis) may give
you a quantitative data that can tell you isotonic point.
Table 3: Number of Plasmolyzed Cells
Salt concentration
Number of
plasmolyzed cells
Number of counted
cells
Percentage of
plasmolyzed cells
0% (water)
0.85%
1%
5%
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CELLULAR MEMBRANES AND OSMOSIS – LAB 7
Apply the Knowledge
1. Did water move across the membrane in all bags containing solutions of sugar?
2. In which bags did osmosis occur?
3. A concentration gradient for water must be present in cells for osmosis to occur. Which
bag represented the steepest concentration gradient relative to its surrounding
environment?
4. Based on your results, are lipids soluble in both acetone and methanol?
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CELLULAR MEMBRANES AND OSMOSIS – LAB 7
5. In which solvent are lipids most soluble?
6. Based on your results, which damages membranes more: 50% methanol or 25% acetone?
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