NOTE: begin answering from question 3 and upIMPORTANT NOTE!! Please use your own words to answer these question because my instructor is so picky and if you answer in a professional way he will deduct points as well as assuming it’s cheating. So, use a common language/basic English to answer these question with the biological terms used in the questions. Answer each question as is DO NOT add additional information! Make is short and simple!NOTE: I have provided a PDF for the lab so you can look at it and answer these questions based on the lab.NO copying off of the internet or plagiarism! Question 3. Describe the Brownian motion in your own words:Question 4. Describe, in your own words, how this observation indicates that all visible and invisible molecules are in motion?After approximately one hour, measure the diameter (in millimeters) of the circle the solutions diffused.Potassium permanganate: Methylene blue: Examine your Petri dish. Shade the agar in the diagram to demonstrate the movement of the solutions in the diagram below. Shade to show where the concentration of the potassium permanganate is higher and where it is lower.Top view of Petri dish: Side view of Petri dish: Question 5. Your Petri dish illustrates the most basic characteristic of diffusion. Complete the following to understand this basic principle:The net diffusion of potassium permanganate appears to be from the area of its ___concentration to the area of its concentration. (“lower to higher” – or – “higher to lower”?)Question 6. In addition to concentration, molecular weight of a diffusing molecule is also an important factor that can determine the rate (speed) of diffusion. We happen to know that potassium permanganate’s molecular weight is 150, while that of methylene blue molecule is 374.Use your own observations (above) to explain how molecular weight appears to affect the rate of diffusion here:Question 7. Based on the results, what is the relationship between molecular weight and diffusion rate?Circle one: Inverse DirectState the basis of your conclusion: Question 8. How would you expect the dyes to be distributed in the agar gel after several days? Would you expect to still see evidence of a concentration gradient?Question 9. Calculate the molecular weights for the following molecules so you can predict which will diffuse more rapidly.Note: Molecular weights are calculated by adding up the atomic weights (masses) for all the atoms that make up a given molecule.) Look at the Periodic Table on the wall in the laboratory. It will give you the atomic weights of all the atoms in the two molecules below. Calculate the weight of these two molecules. Show your work here:Water: H2O Sucrose: C12H22O11 Question 10. Which substance, water or sucrose, is more likely to be able to diffuse through a semipermeable membrane, and on what basis did you make your decision?Question 11. Phospholipids are each made of the phosphate-containing “head” which is hydrophilic (easily combine with water), and two long fatty acid chains which are hydrophobic (do not combine with water, fatty acids are oils). Color the circular phosphate heads blue; color the fatty acid chains of the phospholipids red. Then label them hydrophobic and hydrophilic based on your reading above.Figure 1. Selectively Permeable Cell Membrane Question 12. Observe Figure 1. What kinds of molecules are able to pass through the phospholipid bilayer membrane easily? What kinds of molecules are not able to pass through easily? What are the characteristics of each (charge, polar vs. nonpolar), and size?Move easilyDo not move easilyBecause water is a small and flat molecule (despite it being polar), it can either pass through the phospholipid bilayer membrane. It can more quickly pass into and out of cells through tunnel-like proteins called aquaporins that traverse the cell membrane.Question 13. Label the aquaporin protein in Figure 1. What does the aquaporin allow to pass through the membrane? Question 14. Define the terms Solvent and Solute:Question 15. Below is a diagram of three model cells filled with three different solutions. Make a prediction. Review Figure 2 and complete the Table 1 to its right. For column 4, predict whether the “model cell” will increase or decrease in weight after one hour. (Hint: Will the net diffusion of water be into the “model cell” or into the beaker?)Table 1: Prediction for Model Cells Cell Letter% solutes in cell% water in cellIncrease or decrease after 1 hour?ABCFigure 2. Diagram showing the experimental set up for the 3 model cells Write down the colors of the solutions here: 1% 25% 50% At 15-minute intervals, for 1 hour, record the “total weight” of each cell in Table 2 Calculate the net mass change for each model cell in the last row of Table 2.Net mass change = Final mass (at time 60) – initial mass (at time 0)Table 2. Change in Mass of Three Model Cells by Osmosis over TimeMass (g)Time (minutes)Cell ACell BCell C0 15 30 45 60 Net mass change Question 16. How do your results compare with your prediction in Table 1? Graph the Osmosis data Include the data for the three model cells as 3 separate curves (lines) using three different colors on the same graph. Create a key for identifying the colored line according to its corresponding Cell letter and % Solutes.Write the title “Change in Mass of Three Model Cells with Varying Concentration of Solutes via Osmosis over Time” at the top of the graph. Question 17. Do the words hypertonic, hypotonic, and isotonic refer to the solute or solvent concentration?Question 18. Which solution has the highest concentration of water (the “solvent”): a 1% Sucrose solution or a 50% Sucrose solution? Question 19. Look at your osmosis data in Table 1:Did the water move into or out of all three model cells? If not, is this what you expected?Would you describe the environment outside the three model cells as hypertonic or hypotonic? Question 20. A concentration gradient for water must exist between the inside and the outside of a cell’s membrane for osmosis to occur. Observe the graph. Which of the three model cells represents the one with the steepest concentration gradient of water?Question 21. The steepest concentration gradient of water should result in the highest rate of diffusion (osmosis). Examine your Table 3 osmosis data for the 15 to 30-minute interval. Did the greatest changes in weight occur in the model cells with the greatest concentration gradients of water? Question 22: A cell can be placed in various solutions. Based on the information in the above reading and the concentration gradient, fill in the following table:In the second and third columns, write “more,” “less,” and “the same amount of.”In first blank of the fourth column, write in or outside.In the second blank of the fourth column, write in, outside of, inside of, into, or equally in and out of. A cell placed in a hypertonic solution has …__________ solutes than the surrounding environment__________ water than the surrounding environment.The water __________ the cell will move __________ the cell.A cell placed in a hypotonic solution has …__________ solutes than the surrounding environment__________ water than the surrounding environment.The water __________ the cell will move __________ the cell.A cell placed in an isotonic solution has …__________ solutes than the surrounding environment__________ water than the surrounding environment.The water __________ the cell will move __________ the cell.Draw a picture of each cell in its solution to the left of the description help you clarify this.Question 23. Draw a sketch of several red blood cells observed in each of the three environments. Label their plasma membranes. In the last row write whether water moved into or out of the cells, or whether there was no change.HypertonicIsotonic:Hypotonic Drawing:Description of relative size and shape of cellsWater movement?Important: The red blood cells in the isotonic solution are in a “normal” osmotic condition, the same as when they were in the living animal. If we assume that the cell membrane is semipermeable, do you think water is moving in and out of the cell membrane in an isotonic environment? Describe the equilibrium.Question 24. Notice these plant cells are shaped different than the animal cells (human cheek cells). What give these plant cells the brick-like shape? Question 25. Note the green, jelly-bean shaped chloroplasts. Where are they located within the cell? In the center or around the edge near the cell membrane and cell wall?Question 26. Sketch several Elodea (plant) cells in the space below to the left titled “Cells in tap water.” Label with a word(s) and an arrow pointing to their cell walls, central vacuoles, and chloroplasts. Tap waterHypertonicDrawing:Description of relative size and shape of cellsWater movement?Question 27. Where are the chloroplasts now in relation to the center or the cell wall? Question 28. What had to have happened to the water in the central vacuole for the chloroplasts to be located where they are now after placing hypertonic solution on the slide?Question 29. Observe the large plastic model of a plant cell. Notice that the central vacuole occupies most of the plant cell’s mass. The fluid concentration within this vacuole, called cell sap, contains a high concentration of salt, sugar and protein molecules. How does this help explain the large size of the central vacuole in the Elodea cells when placed in fresh tap water? (Hint: osmosis plays a part here.)Question 30. The plant phenomenon observed above is called plasmolysis. In one word, what has happened to the Elodea cell? Question 31. Notice that a plant cell, unlike an animal cell, has not only a plasma (cell) membrane, but also has a cell wall encasing it. Did the cell wall appear to swell, shrink or remain the same during your observations?Question 32. Placing the wilted elodea cells (or wilted lettuce leaves that you might put into a salad) into tap water causes the plant cells to become turgid once again. Explain, in terms of osmosis, why the elodea leaf (or lettuce leaf) becomes “crisper” after placing it in tap water:Question 33. Explain why the Elodea (plant) cells did not lyse when placed in tapwater as did the animal cells (red blood cells). In other words, what is the reason that there is a difference between animal and cell’s response to being placed in these various solutions? REVIEW Question 34. Compare animal and plant cells in varying solutions:An animal cell placed in a(n)Will· lose water· gain water· maintain same waterCausing the cell to· be unaffected· shrink (crinkle)· or swell (explode)Hypertonic solution Isotonic solution Hypotonic solution A plant cell placed in a(n)Will· lose water· gain water· maintain same waterCausing the cell to· swell (be unaffected, normal, turgid)· become flaccid (drooping plant)· or plasmolyzed (wilting plant)Hypertonic solution Isotonic solution(see your textbook) Hypotonic (tap water) solutionExercise 6
DIFFUSION AND OSMOSIS
Student Learning Outcomes
At the completion of this exercise you should:
(l) Be able to define the terms diffusion and osmosis.
(2) Be able to list and discuss four mechanisms that cells use to move molecules across their
plasma membranes.
(3) Be able to explain what Brownian motion tells us about atoms and molecules.
(4) Be able to explain the relationship between molecular weight and the rate of a molecule’s
diffusion.
(5) Be able to list the characteristics of molecules that can, and those that cannot, move
passively across a cell’s plasma membrane.
(6) Be able to describe how the solute concentration, inside of a cell, affects the rate (speed) of
osmosis.
(7) Be able to define the following terms: concentration gradient, selectively permeable
membrane, hypertonic, hypotonic, isotonic, and homeostasis.
I. Introduction
Virtually all life forms are composed of cells. The cell is called the fundamental unit of life
because within it occur most of the biochemical life processes. One of the phenomena of life
is that the chemical composition of a cell remains fairly constant, in spite of the fact that the
cell continually uses substances from its external environment and at the same time
discharges other substances into its environment: This state of chemical constancy in living
systems is called homeostasis. This homeostasis, in addition to the fact that a cell’s
surroundings are always of relatively different chemical composition from its inside, leads us
to hypothesize that there must be some very selective means of chemical exchange across a
cell’s plasma membrane. Today, we will investigate the processes of movement of some
substances into and out of cells.
Question 1. Most “cells” do not appear to have an obvious “mouth” or other visible structures in
their cell (“plasma”) membranes. Suggest one other way in which materials might be able to pass
through the cell’s membrane:
Question 2. Cell biologists tell us that there are 4 basic mechanisms that cells use to get
molecules across their membranes. You need to learn these 4 strategies. Go to your textbook, or
other reference source, and define the following 4 mechanisms:
1. Osmosis:
2. Facilitated diffusion:
3. Endocytosis and Exocytosis:
4. Active Transport:
Brownian Motion
Robert Brown made an interesting observation in 1827 that led to the principle that “all atoms
and molecules” are in constant motion. Dr. Brown was a botanist and army surgeon who was
looking at particles inside pollen grains when he noticed the “rapid oscillatory motion of
microscopic particles.” He later observed the same movement when looking at substances, like
India Ink. India Ink is made of water and billions of suspended clumps of carbon atoms. Under
high magnification, Brown observed that the clumps of carbon atoms were vibrating wildly in all
directions. He hypothesized that moving water molecules, which cannot be seen, must be
colliding with the clumps of carbon, forcing them to move. Further study has shown that Dr.
Brown was correct and, today, we call this kind of observation “Brownian motion”.
A physical scientist would tell you that particles (atoms and molecules) are moving because
“they have heat energy (Kinetic energy)”. If you ask what kinetic energy is, you will be told that
it is “random molecular motion” which, of course, is a circular argument. The point here is that
we really do not know the ultimate reason why all atoms and molecules on earth are moving,
only that they are and that the more heat energy (“kinetic energy”) atoms or molecules have, the
faster they move.
You are about to make this observation under the compound microscope yourself. The proper
way to carry the compound microscope will be demonstrated. Always use two hands. Make sure
that the cord is not dangling to prevent a tripping hazard. One hand should be holding the base
of the microscope, while the other should hold the arm of the microscope.
Before using the microscope, check for the condition the microscope was left in from the prior
class.
1. Was the microscope placed back in its assigned compartment with the arm facing out
toward you?
2. Was the cord wrapped between the stage and objectives with the plug tucked inside the
cord?
3. Was the cord relatively untangled?
4. Was the ocular lens clean? (If not, clean the dirty lens with the appropriate lens cleaner
and lens paper (not a paper towel).
5. Was the light turned off?
6. Check that the Condenser is at its highest point, directly below the stage. There is a
knob connected to the Condenser that allows it to be moved up and down.
7. Was the mechanical stage centered so that the stage clips don’t hang over the edge of the
stage?
8. Was the scanning objective lens (4x) (not another objective) placed over the stage? If not,
rotate the nose piece until the scanning objective is facing the stage.
9. Was the stage lowered to the lowest setting possible position? If not, use the coarse focus
knob to do so, not the fine focus knob.
10. Were there any slides remaining on the stage? If so, remove the slide and notify your
instructor. It is important that the slide is placed in the correct box, or it may get lost.
11. If any of these conditions were problematic, please let your instructor know.
Procedure:
1. Plug in the cord and turn up the light intensity.to its maximum value and adjust the iris
diaphragm to its most closed setting. As you proceed you can increase the light passing
through the specimen by gradually opening the iris diaphragm.
2. Adjust the distance between the oculars: Without placing the prepared slide on the stage yet,
look through the oculars. You are likely going to see two circles of white light. Do not try to
focus your eyes on any one thing, as nothing is in focus yet. Slowly, move the two oculars
together and/or further apart until the two circles of white light become one circle of light,
the Field of View.
3. Lower the stage using the coarse focus knob, and make sure the (shortest) scanning objective
is facing the stage, so that there is no chance of the slide scratching any objective lenses.
4. Prepare your wet mount slide. Wash and dry a glass slide. Place a small drop of India ink
on a clean slide. Place a cover slip on the slide.
5. Coverslip: Carefully cover the preparation with a clean plastic coverslip as follows:
Place one edge of the coverslip near to the drop. The stain and water with which you mixed
the cells will flow along the junction of the edge of the coverslip and the slide. Carefully
lower the coverslip over the specimen keeping the edge of the coverslip in contact with the
slide. In this way, the water will flow slowly and uniformly about the specimen and force out
air bubbles from beneath the coverslip. (A few air bubbles are not a serious problem for your
first slide.)
coverslip
stain & water drop
with specimen
glass slide
6. Excess fluid: If liquid spills out or may spill out from under the coverslip, gently blot the
excess with a towel, so that it will not later drip onto the stage of the microscope.
7. Before looking through the eyepiece (ocular), open the stage clip, and place the slide on the
stage of the microscope beneath the objective, with the coverslip visible on the upper side.
The stage clip should be holding the slide in place, not pressing the slide under it. Using the
left and right/up and down stage knobs), center the object below the objective without
looking through the oculars. Nothing is in focus yet.
8. Coarse adjustment with scanning lens: With the scanning lens in place, move the stage up
to its highest point without looking through the oculars. Nothing is in focus yet.
Looking through the ocular with your right eye only (squint or cover your left eye), bring the
specimen into focus by turning the coarse focus adjustment knob slowly until the specimen is
generally in focus. Then turning the fine adjustment knob will bring the specimen into
sharper focus.
9. Focus your left eye: Viewing the specimen with both eyes through both oculars, turn the left
ocular diopter until the specimen is clear in both eyes.
10. Iris diaphragm: The light coming through the microscope may be either too bright or too
dim. If the amount of light is not satisfactory, it can be adjusted by carefully regulating the
size of the opening of the iris diaphragm by moving the lever beneath the stage. The iris
diaphragm is part of the condenser which concentrates the light coming from the light source.
11. Adjusting on low and high power objectives – use fine adjustment knobs only: If you wish
to view the specimen using higher magnification, center the specimen in the field, and
carefully rotate the revolving nosepiece to bring the next higher power objective into place
beneath the body tube. The specimen will no longer be in focus. In order to sharpen the
image of the specimen, adjust the focus using only the fine focus adjustment knob.
(Again, the light may have to be adjusted with the iris diaphragm.) Each time you move to
the next higher power objective, be sure you center the specimen beforehand.
12. Examine the drop first under scanning, then low power, then under high power. Be sure
you can see the individual particles of India ink.
13. Focus your attention on one particle (under high power) for several seconds. Look for a
slight but vigorous movement of this particle, independent of the other particles. (Note: You
may need to wait a few minutes. Then, if you see a mass “flowing” movement of all the
particles in one direction like a small river, this is not Brownian motion. Wait for the flowing
to subside, then carefully observe one particle.)
. (Very Import)
This video explains the Brownian Motion Principle
This video shows the actual Brownian Motion you will use to answer the questions.
Question 3. Describe the Brownian motion in your own words:
Question 4. Describe, in your own words, how this observation indicates that all visible and
invisible molecules are in motion?
12. Remove the slide: Once everyone in your group has viewed the Brownian Motion and you
need to remove the slide, be sure to rotate the nose piece to the scanning objective. Then
using the coarse adjustment knob, lower the stage to its lowest position. Then open the stage
clip and remove the slide.
Clean up.
1. Rinse any wet mount slides and place in the container marked “Used slides.” Wash and
dry the coverslips and place them in their original container.
2. Throw away any Kimwipes or other paper.
3. You will need your microscope for other parts of this lab, so leave it available. However,
make sure it is not near where there is sugar solution. Keep it safe.
II. DIFFUSION
Diffusion is the movement of particles from an area of high concentration to an area of low
concentration. This results from the continuous random motion that is characteristic of all
molecules in liquid or the gas states. A few observations about diffusion will help us to
understand how molecules can move from one location to another, perhaps even across cell
membranes.
A. Diffusion through a Colloid
The contents of a cell (the cytoplasm) may be described as a colloid rather than a liquid or solid.
Large protein molecules are present in a cell’s cytoplasm that allow it to be in a transitional state
of matter called a colloid (somewhere between a liquid and a solid). A special kind of colloid,
agar gel, is available in the laboratory and will be used to demonstrate how molecules diffuse
from one place to another once they are inside a cell.
Agar is a carbohydrate extracted from algae in powder form. A gel is prepared by mixing the
powder with water, then heating followed by cooling–similar to the preparation of a gelatin
(animal protein) dessert. The result is a gelatin-like substance composed of intertwined
molecules with water trapped among them. Two compounds (molecules), potassium
permanganate and methylene blue, have been selected to illustrate diffusion through a colloid.
Unlike most components of living cells, these compounds are brightly colored, allowing us to
watch their diffusion.
Procedure:
1. Working with your team, obtain a disposable Petri dish containing agar gel.
2. Using a No. 5 cork borer (a “punch”), make two holes in the agar approximately 5
centimeters apart (See diagram below). Remove the plugs of agar with a toothpick and place
in the trash.
3. Bring your agar dish, the dropper bottles of potassium permanganate solution and the
dropper bottle of methylene blue back to your lab bench.
4. Place two drops of 1% potassium permanganate solution in one of the holes. In the other
hole, place two drops of 1% methylene blue solution. Start a timer for 60 minutes. Return
the bottles back to the side lab benches for others to use.
5. After approximately one hour, measure the diameter (in millimeters) of the circle the
solutions diffused.
Potassium permanganate:
6.
Methylene blue:
Examine your Petri dish. Shade the agar in the diagram to demonstrate the movement of the
solutions in the diagram below. Shade to show where the concentration of the potassium
permanganate is higher and where it is lower.
Top view of Petri dish:
Side view of Petri dish:
Question 5. Your Petri dish illustrates the most basic characteristic of diffusion. Complete the
following to understand this basic principle:
The net diffusion of potassium permanganate appears to be from the area of its
___concentration to the area of its
concentration.
(“lower to higher” – or – “higher to lower”?)
Question 6. In addition to concentration, molecular weight of a diffusing molecule is also an
important factor that can determine the rate (speed) of diffusion. We happen to know that
potassium permanganate’s molecular weight is 150, while that of methylene blue molecule is
374.
Use your own observations (above) to explain how molecular weight appears to affect the rate of
diffusion here:
Question 7. Based on the results, what is the relationship between molecular weight and
diffusion rate?
Circle one:
Inverse
Direct
State the basis of your conclusion:
Question 8. How would you expect the dyes to be distributed in the agar gel after several days?
Would you expect to still see evidence of a concentration gradient?
Question 9. Calculate the molecular weights for the following molecules so you can predict
which will diffuse more rapidly.
Note: Molecular weights are calculated by adding up the atomic weights (masses) for all the
atoms that make up a given molecule.) Look at the Periodic Table on the wall in the
laboratory. It will give you the atomic weights of all the atoms in the two molecules below.
Calculate the weight of these two molecules. Show your work here:
Water: H2O
Sucrose: C12H22O11
Question 10. Which substance, water or sucrose, is more likely to be able to diffuse through a
semipermeable membrane, and on what basis did you make your decision?
Clean up.
1. Throw away your Petri dish, agar included, into the trash can.
2. Wipe down all countertops and student benches with yellow cleaning solution.
III. LIVING CELL MEMBRANES
The membranes that surround all cells allow some molecules to diffuse across while inhibiting
others. In other words, the membranes of cells are said to be “selectively permeable.” The
membranes of all cells are made out of 2 layers of phospholipid molecules with various kinds of
channels passing through them. (See Figure 1). Phospholipids are each made of the phosphatecontaining “head” which is hydrophilic, and two long fatty acid chains which are hydrophobic.
Question 11. Phospholipids are each made of the phosphate-containing “head” which is
hydrophilic (easily combine with water), and two long fatty acid chains which are hydrophobic
(do not combine with water, fatty acids are oils). Color the circular phosphate heads blue;
color the fatty acid chains of the phospholipids red. Then label them hydrophobic and
hydrophilic based on your reading above.
Very small
Polar
molecules
like H2O
Outside of a Cell’s membrane
Ions like Na+
and Cl-
Very large, Polar
Molecules like
Glucose &
Amino Acids
Nonpolar
molecules
like Lipids,
CO2 &
Oxygen
Inside of Cell’s membrane
Figure 1. Selectively Permeable Cell Membrane
Question 12. Observe Figure 1. What kinds of molecules are able to pass through the
phospholipid bilayer membrane easily? What kinds of molecules are not able to pass through
easily? What are the characteristics of each (charge, polar vs. nonpolar), and size?
Move easily
Do not move easily
Because water is a small and flat molecule (despite it being polar), it can either pass through the
phospholipid bilayer membrane. It can more quickly pass into and out of cells through tunnellike proteins called aquaporins that traverse the cell membrane.
Question 13. Label the aquaporin protein in Figure 1. What does the aquaporin allow to pass
through the membrane?
IV. Osmosis and the rate (speed) of diffusion along a concentration gradient
Often, scientists find it helpful to construct a simplified model of an object or phenomenon in
order to understand it more clearly. This is exactly what you will do now. You will make
several different model cells using a plastic membrane material (dialysis tubing) that mimics the
osmosis characteristics of a living cell’s membrane.
The speed or rate at which a molecule diffuses from one area to another depends on the
concentration gradient between the two areas. For example, if the concentration of perfume
molecules is higher in one room compared to an adjacent room connected by an open door, we
would say the concentration gradient between the two rooms is very steep and the rate of
diffusion would be very rapid. Conversely, if the concentration of perfume molecules was equal
in the two rooms, then the rate of diffusion would be zero and the net movement of the perfume
molecules would stop. (Movement would still occur, as Brownian Motion still occurs, but the
diffusion would be at equal rates between the rooms.)
Osmosis follows the same laws as diffusion but always refers to water, the principle solvent in
cells. A solvent is a fluid that dissolves substances, while the term solute is used to describe
substances dissolved in a solvent to make a solution. This means that water will move down its
concentration gradient, too, from an area of high concentration of water to an area of low
concentration of water.
Because water is the universal solvent in all cells its diffusion into or out of cells is critical in
living organisms. Too much water entering or leaving a cell will cause cell death and often the
death of the entire organism. You will now set up several different model cells and measure the
direction and rate (speed) of osmosis.
Question 14. Define the terms Solvent and Solute:
Question 15. Below is a diagram of three model cells filled with three different solutions. Make
a prediction. Review Figure 2 and complete the Table 1 to its right. For column 4, predict
whether the “model cell” will increase or decrease in weight after one hour. (Hint: Will the net
diffusion of water be into the “model cell” or into the beaker?)
Cell A
10 ml of
1%
Sucrose
(solutes)
Inside cell
Cell B
10 ml of
25%
Sucrose
(solutes)
Inside cell
Cell C
10 ml of
50%
Sucrose
(solutes)
Inside cell
Table 1: Prediction for Model Cells
%
%
Increase or
Cell
solutes water
decrease
Letter
in cell in cell after 1 hour?
A
B
C
A
B
C
DI water
(0% solutes)
Figure 2. Diagram showing the experimental set up for the 3 model cells
Observe the video on using triple beam balance.
jny01UtU3MgKcuykkYCOXTYDNgip&index=4&t=0s
Procedure steps 1-17
• Observe the video on Model Cells while reading
procedure and take data in Table 2.
re=youtu.be
The (t) = time. The % is the bag Sugar concentration.
Procedure:
1. Obtain a three pieces of paper towel on which to place your plastic tubing. This tubing is
like plastic wrap or plastic bags. While unseen by the naked eye, the plastic has molecular
holes of a specific size that allow some molecules to diffuse across the plastic while blocking
other, larger, molecules. Therefore, the plastic tubing is selectively permeable. This tubing
will simulate the structure of the membrane
2. Obtain a short stack of paper towels, wet the center towels in the stack with deionized water,
so you will have dry towels on the bottom and top and wet towels in the middle. Use this
“bed” to keep you plastic tubing moist. Allowing the tubing to dry out will cause it to lose its
permeability properties
3. Obtain a beaker, and fill it 2/3 full with deionized water
4. Obtain 3 pieces of water-soaked plastic tubing 20 cm long and six pieces of thread. Twist one
end of the first plastic tubing, and fold down. Then tie the folded end tightly with a double
knot.
5. Open the other end of the tube by rolling with your fingers. Don’t let the tubing get dry, or it
may crack.
6. Write down the colors of the solutions here:
1%
25%
50%
7. Fill the 3 plastic tubes with the contents shown below in Figure 2.
8. Twist the other end of the plastic tubing, then fold down and tie a string around that end. The
baggie you have created should not be entirely full of water, but instead have a bit of space
remaining at the top. Tie the folded ends securely. The goal is to create a water-tight bag.
9. Check for leaks. First, press any extra liquid out of the ends created by filling the bags.
Then, by gently but firmly rolling the bag on the paper towel, checking for leaks. Then trim
the excess thread. Keep the model cells on the clean paper towel between measurements of
cells. Have your instructor check.
10. Check to see if your Triple Beam balance is calibrated properly This means the line on the
end of the center beam lines up at the zero line when all of the weights (poises) are slid to the
left at zero. If the lines do not meet at the zero line, ask your instructor to assist you.
11. Next, blot excess water off the model cells, and place it on the center of the balance pan.
First, move the heaviest weight to the right to the first notch which causes the pointer to drop,
then, move it back one notch, causing the pointer to rise. If the pointer goes below the zero,
the weight is too much, so you will need to try a lighter weight. If it stays above the zero, you
will need to add more weight.
12. Add weights as necessary, moving along one beam and then the next lighter beam until the
pointer goes to zero. The weight of the specimen is the sum of the values for all of the weigh
position, read directly from the graduated beams.
13. Weigh each model cell to nearest 0.01 grams using the balance on your lab bench. If you are
not sure how to read this measurement on the Triple Beam balance, please ask your instructor
to assist you. Record the “cell” weights in Table 2 in the column titled “initial weight.”
14. Place cells A, B, and C simultaneously in a beaker filled with 100% distilled water
(solvent) (See Figure 2 again). Note the time here: _________
15. Remove all 3 model cells from their beaker every 15 minutes for the next hour. Place the
model cells in your paper towel bed. Blot dry and weigh each model cell, being careful to dry
the string and the area where you have folded the tubing. Weigh the cells again to the nearest
0.1 gram. Handle the cells very carefully to avoid causing leaks. After you have weighed and
recorded the weight, return the “cells” back to the beaker for the next 15-minute period
16. At 15-minute intervals, for 1 hour, record the “total weight” of each cell in Table 2.
17. Calculate the net mass change for each model cell in the last row of Table 2.
Net mass change = Final mass (at time 60) – initial mass (at time 0)
Table 2. Change in Mass of Three Model Cells by Osmosis over Time
Time (minutes)
0
15
30
45
60
Net mass change
Cell A
Mass (g)
Cell B
Cell C
Question 16. How do your results compare with your prediction in Table 1?
Graph the Osmosis data
1. Using the graph paper below, label the graph with the vertical (“Y”) axis as “Mass (grams)”
and the horizontal (“X”) axis as “Time (minutes).”
2. Plot the data from Table 2 for “Mass” at each of the 15-minute intervals.
3. Include the data for the three model cells as 3 separate curves (lines) using three different
colors on the same graph. Create a key for identifying the colored line according to its
corresponding Cell letter and % Solutes.
4. Write the title “Change in Mass of Three Model Cells with Varying Concentration of Solutes
via Osmosis over Time” at the top of the graph.
Clean up
1. Drain the sugar water from the plastic tubing down the sinks, and then rinse off the tubing.
Flush the sink with plenty of water. Throw the plastic tubing in the trash.
2. Spray a clean paper towel with the yellow cleaning solution, and then wipe your Triple Beam
balance pan.
3. Wipe off all surfaces, including your lab bench, the side lab benches (under the sugar
solution dispersal containers). (Sugar attracts ants!)
4. Throw all paper towels away.
Hypotonic, Hypertonic, and Isotonic Environments.
The three terms above are used to describe solute concentration environments a cell may find
itself in. In two of these environments water will enter or leave the cell and the cell will change
shape and perhaps die. You must learn what these terms mean so you be able to answer questions
that use them.
These three terms always describe the solute concentration on one side of cell’s membrane
relative to the solute concentration on the other side of the cell membrane.
The term hypotonic means “less solute concentration here” relative to the other side of a cell’s
membrane. If you are told the outside environment a cell finds itself in is hypotonic, then you
automatically know that the inside environment is the exact opposite or hypertonic.
As you recall, the cell membrane will not allow large or ionic substances to move across it, but
water may pass. Your job will always be to determine whether the cell will swell up with water
or shrink due to water loss. To do this you must decide which side of the membrane has a
higher water concentration. Once this is done you can predict which way water will move
(diffuse) since molecules (such as water) always diffuse from an area of higher concentration
to an area of their lower concentration.
Look at the osmosis data you collected for Model cell A. We had you put a 1% solute (sucrose)
concentration inside the cell and surrounded it with deionized water (a solution that had 0%
solute concentration). The proper terminology would be that the inside of the model cell was
hypertonic (greater solute concentration) compared to the outside environment that would be
described as hypotonic (meaning a lower solute concentration).
To predict which way water will move, you must translate the solute concentrations to water
(solvent) concentrations for each side of the cell. Here, the inside of the cell is hypertonic to the
outside, hence, the water concentration is lower inside the cell when compared to the outside.
So now we can predict that water will move into Model cell A and the cell will get larger. Check
your Osmosis data in Table 1 and you should see that, indeed, Model cell A weighed more after
1 hour because it gained water. (If not, you had a leak somewhere!)
Question 17. Do the words hypertonic, hypotonic, and isotonic refer to the solute or solvent
concentration?
Question 18. Which solution has the highest concentration of water (the “solvent”): a 1%
Sucrose solution or a 50% Sucrose solution?
Question 19. Look at your osmosis data in Table 1:
a. Did the water move into or out of all three model cells? If not, is this what you expected?
b. Would you describe the environment outside the three model cells as hypertonic or
hypotonic?
Question 20. A concentration gradient for water must exist between the inside and the outside
of a cell’s membrane for osmosis to occur. Observe the graph. Which of the three model cells
represents the one with the steepest concentration gradient of water?
Question 21. The steepest concentration gradient of water should result in the highest rate of
diffusion (osmosis). Examine your Table 3 osmosis data for the 15 to 30-minute interval. Did the
greatest changes in weight occur in the model cells with the greatest concentration gradients of
water?
V. OBSERVATIONS OF LIVING CELLS
In any liquid that contains various dissolved solutes, all of the different solutes added together
constitute the total osmolar concentration (osmolarity) of the solution. It is of critical importance
that two solutions which have the same osmolar (solute) concentration also have the same water
(solvent) concentration. This is due to the fact that, even though solute molecules may be of
different weights, most appear to take up about the same amount of space. The result is that two
solutions with different solutes and identical osmolarities also have identical water molecule
concentrations (water potentials).
We know that the liquid inside a cell contains many different dissolved substances; they make up
the cell’s total osmolar concentration. If a cell is placed in a solution with a higher osmolarity
(lower water potential) than inside the cell, we say that the cell is in a hypertonic solution. When
a cell is placed in a hypotonic solution, a lower osmolarity (higher water potential) exists outside
the cell than inside. A solution that has identical osmolar concentration (water potential) to the
cell’s fluid is called isotonic. In this part of the exercise, we will observe the effects of various
saline solutions on living animal and plant cells.
Question 22: A cell can be placed in various solutions. Based on the information in the above
reading and the concentration gradient, fill in the following table:
In the second and third columns, write “more,” “less,” and “the same amount of.”
In first blank of the fourth column, write in or outside.
In the second blank of the fourth column, write in, outside of, inside of, into, or equally in
and out of.
A cell placed
in a hypertonic
solution has …
__________ solutes
than the surrounding
environment
__________ water
than the surrounding
environment.
The water __________ the cell
A cell placed
in a hypotonic
solution has …
__________ solutes
than the surrounding
environment
__________ water
than the surrounding
environment.
The water __________ the cell
A cell placed
in an isotonic
solution has …
__________ solutes
than the surrounding
environment
__________ water
than the surrounding
environment.
The water __________ the cell
will move __________ the cell.
will move __________ the cell.
will move __________ the cell.
Draw a picture of each cell in its solution to the left of the description help you clarify this.
A. Osmosis in Red Blood Cells (Erythrocytes)
Watch this video
• Red blood cells are one type of cell found in blood. There are also platelets
and several types of white blood cells found in blood plasma. Blood transports
oxygen, carbon dioxide, glucose, ions, hormones and proteins throughout the
body.
The first part of the video the blood cells are in an Isotonic solution.
The second part is the Hypertonic solution.
The last part is in a Hypotonic solution.
Procedure:
1. If you already set up your microscope for the Brownian Motion procedure, then skip to
Procedure Step 6.
2. If you did not set up your microscope for the Brownian Motion procedure, then when you
obtain your microscope from the cabinet, confirm that the scanning objective is facing the
stage, the stage is placed at its lowest position, and the cord is wrapped on top of the stage. If
not, please notify your instructor.
3. Each team should set up three microscopes, then prepare the following three slides, and
observe all three slides side by side. If three microscopes aren’t available, you can set up one
or two microscopes. Set them up in locations that are not close to the sugary model cells,
their beaker, or the triple beam balance. Give yourself room so that you will be able to sit
side by side another person using another microscope.
4. Plug in the cord and turn up the light intensity to its middle value and adjust the iris
diaphragm to its most closed setting. As you proceed you can increase the light passing
through the specimen by gradually opening the iris diaphragm.
5. Adjust the distance between the oculars: Without placing a slide on the stage yet, look
through the oculars. You are likely going to see two circles of white light. Do not try to
focus your eyes on any one thing, as nothing is in focus yet. Slowly, move the two oculars
together and/or further apart until the two circles of white light become one circle of light,
the Field of View.
6. Lower the stage using the coarse focus knob, and make sure the (shortest) scanning objective
is facing the stage, so that there is no chance of the slide scratching any objective lenses.
7. Obtain a dry paper towel and then write “Hypertonic,” “Isotonic,” and “Hypotonic” on it.
8. Prepare your wet mount slide. Place a small drop of hypertonic saline solution on a clean
slide. On the same slide, near the saline drop, touch the dropper from the blood dropper
bottle to the slide. A slightly smaller drop of blood is desirable.
9. Using a clean toothpick, mix the blood and saline together. Apply a cover slip. If liquid
spills out from under the coverslip, gently blot the excess with a towel, so that it will not later
drip onto the stage of the microscope.
10. Before looking through the eyepiece (ocular), open the stage clip, and place the slide on the
stage of the microscope beneath the scanning objective, with the coverslip visible on the
upper side. The stage clip should be holding the slide in place, not pressing the slide under it.
Using the left and right/up and down stage knobs) center the object below the objective
without looking through the oculars. Nothing is in focus yet. Alternatively, since the object
is small, and you may not know what blood cells look like yet, move the slide so that the
edge of the coverslip is under the scanning objective.
11. Coarse adjustment with scanning lens: With the scanning lens in place, move the stage up
to its highest point without looking through the oculars. Nothing is in focus yet.
Looking through the ocular with your right eye only (squint or cover your left eye), bring the
specimen into focus by turning the coarse focus adjustment knob slowly until the specimen is
generally in focus. Then turning the fine adjustment knob will bring the specimen (the
coverslip edge) into sharper focus.
12. Focus your left eye: Viewing the specimen with both eyes through both oculars, turn the left
ocular diopter until the specimen (the coverslip edge) is clear in both eyes. Now you can
move your attention to the very tiny red blood cells. It is best to look in a location where
there are fewer of them (less red on the slide macroscopically – that is, when you look at the
slide without the aid of the microscope). When they are cramped together, it is difficult to
determine if the shape is due to them being cramped together or not. Focus so that you can
see individual cells.
13. Iris diaphragm: The light coming through the microscope may be either too bright or too
dim. If the amount of light is not satisfactory, it can be adjusted by carefully regulating the
size of the opening of the iris diaphragm by moving the lever beneath the stage. The iris
diaphragm is part of the condenser which concentrates the light coming from the light source.
14. Adjusting on low and high power objectives – use fine adjustment knobs only: If you wish
to view the specimen using higher magnification, center the specimen in the field, and
carefully rotate the revolving nosepiece to bring the next higher power objective into place
beneath the body tube. The specimen will no longer be in focus. In order to sharpen the
image of the specimen, adjust the focus using only the fine focus adjustment knob.
(Again, the light may have to be adjusted with the iris diaphragm.) Each time you move to
the next higher power objective, be sure you center the specimen beforehand.
15. Draw the red blood cells (placed in hypertonic solution) under the high power objective.
16. When you are done with the hypertonic solution, remove the slide and place it on the paper
towel where you wrote “Hypertonic solution.” Then repeat steps 8 and 9, this time using
isotonic and hypotonic solutions on each of your slides, using a clean toothpick each time.
Question 23. Draw a sketch of several red blood cells observed in each of the three
environments. Label their plasma membranes. In the last row write whether water moved into
or out of the cells, or whether there was no change.
Hypertonic
Isotonic:
Hypotonic
Drawing:
Description
of relative
size and
shape of cells
Water
movement?
Important: The red blood cells in the isotonic solution are in a “normal” osmotic condition, the
same as when they were in the living animal. If we assume that the cell membrane is
semipermeable, do you think water is moving in and out of the cell membrane in an isotonic
environment? Describe the equilibrium.
B. Osmosis in Plant Cells
Procedure:
1. If you already set up your microscope for the Brownian Motion and/or red blood cell
procedures, then skip to Procedure Step 5 below.
2. If you did not set up your microscope for the Brownian Motion and/or red blood cell
procedures, then when you obtain your microscope from the cabinet, confirm that the
scanning objective is facing the stage, the stage is placed at its lowest position, and the cord
is wrapped on top of the stage. If not, please notify your instructor.
3. Plug in the cord and turn up the light intensity to its middle value and adjust the iris
diaphragm to its most closed setting. As you proceed you can increase the light passing
through the specimen by gradually opening the iris diaphragm.
4. Adjust the distance between the oculars: Without placing a slide on the stage yet, look
through the oculars. You are likely going to see two circles of white light. Do not try to
focus your eyes on any one thing, as nothing is in focus yet. Slowly, move the two oculars
together and/or further apart until the two circles of white light become one circle of light,
the Field of View.
5. Lower the stage using the coarse focus knob, and make sure the (shortest) scanning objective
is facing the stage, so that there is no chance of the slide scratching any objective lenses.
6. Prepare your wet mount slide. Remove a bright green, young leaf from an Elodea plant, and
place it in a drop of tap water on a clean slide. Try to obtain a leaf from the tender growing
tip of the sprig and avoid taking leaves which have been drying out of the water. Apply a
cover slip.
7. Place your slide on the stage by opening the stage clips. Do not place the slide under the
stage clips or they will bend.
8. Coarse adjustment with scanning lens: With the scanning lens in place, move the stage up
to its highest point without looking through the oculars. Nothing is in focus yet.
Looking through the ocular with your right eye only (squint or cover your left eye), bring the
specimen into focus by turning the coarse focus adjustment knob slowly until the specimen is
generally in focus. Then turning the fine adjustment knob will bring the specimen into
sharper focus.
9.
Focus your left eye: Viewing the specimen with both eyes through both oculars, turn the
left ocular diopter until the specimen) is clear in both eyes. It is best to look along an edge of
the leaf, rather in the center where the vein is. There are fewer layers of cells along the edge
of the leaf.
10. Iris diaphragm: The light coming through the microscope may be either too bright or too
dim. If the amount of light is not satisfactory, it can be adjusted by carefully regulating the
size of the opening of the iris diaphragm by moving the lever beneath the stage. The iris
diaphragm is part of the condenser which concentrates the light coming from the light
source.
11. Adjusting on low and high power objectives – use fine adjustment knobs only: If you wish
to view the specimen using higher magnification, center the specimen in the field, and
carefully rotate the revolving nosepiece to bring the next higher power objective into place
beneath the body tube. The specimen will no longer be in focus. In order to sharpen the
image of the specimen, adjust the focus using only the fine focus adjustment knob.
(Again, the light may have to be adjusted with the iris diaphragm.) Each time you move to
the next higher power objective, be sure you center the specimen beforehand.
12. Focusing on one cell at a time, note the shape of Elodea cells.
Exercise 6: Diffusion and Osmosis (SIM 8/2020)
Tap Water
Question 24. Notice these plant cells are shaped different than the animal cells (human cheek
cells). What give these plant cells the brick-like shape?
Question 25. Note the green, jelly-bean shaped chloroplasts. Where are they located within the
cell? In the center or around the edge near the cell membrane and cell wall?
13. Note the evidence of a central vacuole within each cell. (You can’t actually see the vacuole
clearly, since it is a large, transparent membrane filled with fluid in the middle of the cell).
Occasionally, you will see a cell with its nucleus visible (a colorless oval next to the cell
wall). Use your fine adjustment to perceive the depth dimension of the cells.
14. After a few minutes under the microscope light, you may notice the chloroplasts moving
around the inner edge of the cell membrane. This is known as “cytoplasmic streaming.”
• Complete Questions 26 – 34, plus the tables.
Tap Water
Hypertonic
Solution.
Question 26. Sketch several Elodea (plant) cells in the space below to the left titled “Cells in
tap water.” Label with a word(s) and an arrow pointing to their cell walls, central vacuoles, and
chloroplasts.
Tap water
Hypertonic
Drawing:
Description of relative
size and shape of cells
Water movement?
15. Remove the slide. (Don’t forget to rotate down to the scanning objective and moving the
stage down before you remove the slide.)
16. Now allow 1-2 drops of hypertonic saline solution to flow under one edge of the cover slip,
and draw the solution under the coverslip by touching the fluid at the opposite edge with a
piece of paper towel. Then view the cells under the same power you did with the cells in tap
water.
Question 27. Where are the chloroplasts now in relation to the center or the cell wall?
Question 28. What had to have happened to the water in the central vacuole for the chloroplasts
to be located where they are now after placing hypertonic solution on the slide?
Question 29. Observe the large plastic model of a plant cell. Notice that the central vacuole
occupies most of the plant cell’s mass. The fluid concentration within this vacuole, called cell
sap, contains a high concentration of salt, sugar and protein molecules. How does this help
explain the large size of the central vacuole in the Elodea cells when placed in fresh tap water?
(Hint: osmosis plays a part here.)
Question 30. The plant phenomenon observed above is called plasmolysis. In one word, what
has happened to the Elodea cell?
Question 31. Notice that a plant cell, unlike an animal cell, has not only a plasma (cell)
membrane, but also has a cell wall encasing it. Did the cell wall appear to swell, shrink or remain
the same during your observations?
Question 32. Placing the wilted elodea cells (or wilted lettuce leaves that you might put into a
salad) into tap water causes the plant cells to become turgid once again. Explain, in terms of
osmosis, why the elodea leaf (or lettuce leaf) becomes “crisper” after placing it in tap water:
Question 33. Explain why the Elodea (plant) cells did not lyse when placed in tapwater as did
the animal cells (red blood cells). In other words, what is the reason that there is a difference
between animal and cell’s response to being placed in these various solutions?
VI. REVIEW
Question 34. Compare animal and plant cells in varying solutions:
An animal cell placed in a(n)
Will
• lose water
• gain water
• maintain same water
Hypertonic solution
Isotonic solution
Hypotonic solution
Causing the cell to
• be unaffected
• shrink (crinkle)
• or swell (explode)
A plant cell placed in a(n)
Will
• lose water
• gain water
• maintain same water
Causing the cell to
• swell (be unaffected, normal, turgid)
• become flaccid (drooping plant)
• or plasmolyzed (wilting plant)
Hypertonic
solution
Isotonic solution
(see your textbook)
Hypotonic (tap
water) solution
Graphing your Model Cell Data
•
•
•
If you know how to use Excel or other graphing program, you may use
that.
If you would like to graph the data by hand in the lab manual, you may
do so.
If you do not know how to use Excel or other graphing program, would
not like to graph by hand, or you would like to learn another method,
please click on the following link, and then follow the directions on the
next slide.
https://www.onlinecharttool.com/graph
Exercise 6: Diffusion and Osmosis (SIM 8/2020)
Graphing your Data on “Online Chart Tool”
1) Clickonthefollowinglink,andthenfollowthedirectionsonthenext slide.
https://www.onlinecharttool.com/graph
2) Selecttheappropriatecharttype
•
Most commonly this will be a line graph, a bar graph, or a scatter plot.
•
•
•
Line graphs should be used when observing a continuous change over
time, that is, when the experimental or independent variable is time.
A bar graph is used when the data are categories and you are comparing
groups.
A scatterplot is like a trend line, but in that there will be an
experimental variable
with groups being compared over time.
•
Are we comparing groups over time in this exercise? Yes, then we will
use a scatterplot graph.
Exercise 6: Diffusion and Osmosis (SIM 8/2020)
Graphing your Data on “Online Chart Tool” (cont.)
3) Under Scatter, you will see many options.
1. a) Direction: Horizonal
2. b) Legend: Right
3. c) The remainder can remain the same, though in the future, you may
choose to personalize your graphs with colors, etc.
4. d) Click Next.
4) Next add your data (you may need to scroll up):
a) Create a graph title using this format: The Effect of (the Experimental
Variable) on (the Dependent Variable) via Osmosis over Time
1. The Experimental Variable here is the Varying Concentrations of
Solutes
2. The Dependent Variable here is the Change in Mass of Three Model
Cells
3. Time is the Independent Variable here.
Exercise 6: Diffusion and Osmosis (SIM 8/2020)
Graphing your Data on “Online Chart Tool” (cont.)
4) Next Add your data (you may need to scroll up) (cont.):
b) Type in the X-Axis Title
1. This should be Time, plus units
c) Type in the Y-Axis Title
2. This should be the Dependent Variable, plus any units. Since you already
stated “of Model Cells” in your overall title, you can write Change in Mass (g),
only.
d) You will have 5 items and 3 groups: The 5 items are the data points in your
table in Figure 2. The 3 groups are the cells.
e) Label the Groups as Cell A, Cell B, and Cell C, and type the % sucrose in
parentheses behind each.
Exercise 6: Diffusion and Osmosis (SIM 8/2020)
Graphing your Data on “Online Chart Tool” (cont.)
5) Input your data (cont.)
b) ThePointrepresentsthefirstdatapointonthegraph 1. X = time. Point 1’s X
equals the initial time at 0 minutes.
2. Y = mass. Point 1’s Y equals the original mass at 0 minutes. 3. Point 2’s X
equals the second time at 20 minutes.
c) d)
4. Point 2’s Y equals the change in mass at 20 minutes, etc.
You may wish to change colors to personalize your graph. Click Next.
6) Choose labels and fonts.
a) ClickNoforShowLabels.
b) YoumaychangecolorsorfontsbutleaveeverythingelseasisforData labels.
Remember, it should be easy to read.
c) Click Next.
Exercise 6: Diffusion and Osmosis (SIM 8/2020)
Graphing your Data on “Online Chart Tool” (cont.)
8) Download or Save Your Graph
9)
a) Depending on your instructions from your instructor.
b) You may need to download and submit to your instructor. In this case, save
as a jpg.
c) If you Save your Chart and Data, then you must make an account.
d) You can copy your graph, by right clicking on the graph, and copy image,
then paste. (This process depends on the type of computer you use.)
Add trendlines.
1. a) Normally trendlines should be included between each of the points
of the same color.
2. b) However, this program is limited, so you can hand draw them in if
you print out the graph.
3. c) You can also just leave it without trendlines and visualize the trend
over time when answering the questions in your lab manual.
Exercise 6: Diffusion and Osmosis (SIM 8/2020)
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