Anatomy & Physiology I Lab – Cell Transport
© Atsma/Rohrer 2014
The plasma membrane is one of life’s most essential features. It keeps the cell’s special
chemistry of life separate from the outside world. It does this by being selectively
permeable or semipermeable, meaning it allows some things to move across but is a
barrier to most particles. It is the membrane’s chemical structure that determines which
materials may move in or out of the cell.
The plasma membrane is basically two layers of phospholipids with cholesterol and
protein molecules wedged in between them. Phospholipids are structurally similar to
triglycerides, with a phosphate group attached to one of the carbons at the glycerol
“head” end, and two fatty acid “tails.”
Why is this molecule so special? The fatty acid “tails” are non-polar, hydrophobic molecules
that attempt to "hide" from water, while the glycerol/phosphate head is polar and
hydrophilic (“water-loving”). Thus, this molecule normally orients itself with the head facing
water and its two fatty acid tails attempting to turn away from water.
This tendency of phospholipids alone might not make for a good membrane for a cell, but
membranes use a double layer of phospholipids, set up as mirror images. This is referred to
as the phospholipid bilayer. As you look at the figure below, you can appreciate how
effective this strategy is. The water inside and outside the cell is facing a layer of
phospholipid heads, while the intertwined fatty acid tails stick together in order to remain
hidden from the water.
~~~~~~~~ Water outside the cell ~~~~~~~~
~~~~~~~~ Water inside the cell ~~~~~~~~
Because most of the distance molecules would have to move across the membrane is nonpolar and hydrophobic, the plasma membrane's phospholipid bilayer is a natural barrier to
most molecules important to living things such as proteins, sugars, and sodium, potassium,
calcium, and chloride ions. Thus, after a cell goes through the trouble of making a sugar or
protein, it stays trapped inside and will not be lost by accidentally dissolving out of the cell
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into the surrounding water. Ions and smaller polar molecules such as glucose can get
across the membrane, but require the aid of proteins for transport.
Many different proteins are found in the membrane. The ones relevant to today’s lab are
the transport proteins. These integral proteins span the membrane and make it possible
for the cell to move things such as ions across the membrane that otherwise could not pass
through a phospholipid bilayer. Channel proteins form a tube for passage of particles that
crosses through the hydrophobic region of the membrane. Carrier proteins bind to
particles on one side of the membrane then change shape in order to transport the particle
to the opposite side of the membrane.
Movement of particles across the membrane occurs by either passive or active processes.
Active transport requires the cell use energy in the form of ATP, and passive transport
does not require the use of ATP, usually relying on simple diffusion of the particle.
There are forms of membrane transport that are dependent more on the size of the
particles than chemical properties such as polarity. For today’s lab you will study passive
processes in semi-permeable membranes: diffusion, osmosis, and filtration.
Define the term, “semipermeable membrane.”
Being selectively permeable or semipermeable means it allows some things to move across
but is a barrier to most particles.
What kinds of particles can move across a phospholipid bilayer? What are excluded?
Non-polar molecules and water molecules can pass through, ions and polar molecules
(proteins, sugars, and sodium, potassium, calcium, and chloride) are excluded.
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Diffusion
Diffusion is the movement of particles from an area where they are present at a high
concentration to an area of lower concentration. Diffusion may occur in mixtures of gases
or liquids. For example, perfume diffuses through the air, and a drop of food coloring will
diffuse throughout a cup of water.
Since the source of energy for this movement is kinetic energy, motion due to heat
energy absorbed by the particles, it is a passive process. Kinetic energy induces random
movement of particles, causing them to bounce away from each other when they collide.
Molecules in an area of high concentration are likely to collide with each other and bounce
outward, while molecules moving through an area of low concentration have nothing to
stop them until they reach the wall of the container.
This net flow of particles down their concentration gradient (from high to low
concentration) is diffusion. Diffusion will continue as long as there is a concentration
gradient. When the concentration of particles is uniform throughout the mixture, the
particles will continue to move, but there will be no net movement toward a particular area;
the system is at equilibrium.
Activity – Diffusion through an Agar Gel
The rate at which particles move during diffusion is proportional to their molecular weight.
In the following experiment you will determine the relationship between molecular weight
and rate of diffusion. You will compare the rate of diffusion of potassium permanganate
(MW = 158) with methylene blue (MW = 320) through an agar gel. The agar gel is similar
to gelatin, consists of 98.5% water, and is freely permeable to these particles despite its
somewhat solid appearance.
Materials
1. Petri dish with agar
2. Cork borer
3. Wax pencil
4. Ruler marked in millimeters
5. 0.1 M Potassium permanganate in a
dropper bottle
6. 0.1 M Methylene blue in a dropper
bottle
Procedure
1. Use the cork borer to gently punch two holes approximately 3 centimeters apart in the
agar plate. You will be able to easily remove the agar if you seal the top end of the cork
borer with you finger before pulling away from the agar. Press down on the agar
around the holes so that it will seal to the petri plate.
2. Mark the bottom of the petri dish with the wax pencil so you will know which dye goes
into which hole.
3. Partially fill one of the holes with potassium permanganate solution and the other with
methylene blue. Work carefully so that you do not spill or overfill the holes.
4. Record the time.
5. Cover the dish and set it aside where it will not be disturbed.
6. Use a ruler to measure the distance (in millimeters) that each dye diffused after
approximately 15, 30, and 45 minutes. Record your results in the table below.
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Time
Distance traveled by
methylene blue
Distance traveled by
potassium
permanganate
15 minutes
30 minutes
45 minutes
1. Which substance diffused faster?
Potassium permanganate
2. Was the substance that diffused faster the larger or smaller one?
Smaller
3. What can you conclude about the relationship between molecular weight and the speed
of diffusion based upon these results?
The smaller the molecule, the faster the rate of diffusion.
OSMOSIS
The diffusion of a solvent1, such as water, through a semi-permeable membrane is called
osmosis. In the human body, water is usually the solvent.
Osmosis occurs when a semi-permeable membrane separates two solutions with different
solute concentrations. Since water molecules attach to solutes through hydrogen bonding,
a solution with more solute has less water molecules available for diffusion. Water will
diffuse down its own concentration gradient moving from the solution with less solute (and
more free water molecules) to the solution with high solute concentration (and fewer free
water molecules). The movement of water across the membrane during osmosis produces
Osmotic Pressure. Osmotic pressure plays an important role in the regulation of fluid and
electrolyte balance in the human body.
Activity – Osmosis
In this experiment you will be able to observe osmosis by using Karo syrup (mostly
disaccharides) to create a concentration gradient across the membrane. Disaccharides are
too large to move through the cellophane tubing membrane.
Materials:
1. Glass tubing
2. 500 ml beaker
3. ringstand
4. burette clamp
5.
6.
7.
8.
thread
20 cm cellophane tubing, pre-soaked in water
Karo Syrup + Congo red dye in a squeeze bottle
Small ruler
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A solution is a homogeneous mixture of two or more substances; generally a liquid and some other solid or gaseous
substance. In a solution the solvent and solute always remain mixed. The most abundant substance in the mixture (usually a
liquid) is the solvent. The other substances are the solutes. In most biological systems, water is the solvent and salts, sugars,
and other small polar molecules that dissolve in the water are the solutes. Suspensions are heterogeneous mixtures in which
the particles tend to separate. Flour and water form a suspension.
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Procedure:
1. Tie a knot in the end of a piece of cellophane tubing so that it forms a bag about 5
inches long. Keep the cellophane wet as you work with it.
2. Fill the bag approximately two-thirds full with the Karo Syrup + Congo red dye.
3. Insert approximately 1 inch of the glass tube into the bag, and tie the bag tightly to the
glass tube with thread. Make sure the bottom of the glass tube is submerged in liquid.
4. Rinse the bag under tap water to remove any Karo syrup from the bag’s outer surface.
5. Use the clamp and ringstand to suspend the bag in the beaker filled with tap water. The
Karo syrup + Congo red solution should be visible in the glass tube.
6. Use the ruler to measure your reference point for the level of the red solution in the
tube (how far above or below the thread the liquid starts).
7. Visually check for signs that the bag is leaking (the reason for the red dye). If it is
leaking, tighten the knot, tightly retie the string, or start over if that doesn’t work.
8. Measure the change in the level of the solution in the tube after 10, 20, and 30 minutes.
Record your results in the table below.
Steps 1-2:
Steps 3-5
Time
Distance traveled by the solution
10 minutes
Your data will vary, but the solution
20 minutes
should have moved steadily up the
30 minutes
tube to near the top.
Note: If the experiment is set up correctly, the results usually happen quickly. If you have no
movement in the fluid level after 10-15 minutes, you may have a faulty setup (e.g. may not
have tied the string tightly enough) and should fix it or start over.
1. Will the solute concentration inside the bag ever become equal to the solute
concentration in the beaker? No
2. Will osmosis stop in this setup if you wait for a longer period of time? No
Explain your answer:
Since the solute is trapped inside, there will always be more
solute (and less free water) inside the bag. Water will continue to move in.
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3. What would have happened if you had filled the bag with glucose (which is small enough
to diffuse through the bag) solution instead of Karo syrup?
You may have observed some osmosis initially, but equilibrium would eventually
occur between the bag contents and the water outside. Osmosis would stop then.
FILTRATION
Bulk flow occurs when a solution or suspension is forced to move by some external force
such as hydrostatic (water) pressure, blood pressure, or air pressure. Bulk flow moves
much larger amounts of material than diffusion. Filtration occurs if the mixture is forced
through a membrane that separates the particles according to their size. The material
(solvent and particles) that passes through the filter is called the filtrate. Since the
energy for filtration is supplied by fluid or air pressure, it is a passive process. For
example, filtration is physiologically important for urine formation in the kidneys.
Activity – Filtration
In this procedure you will filter a mixture and look at the effects of hydrostatic pressure on
filtration rate.
Materials
1. beaker
2. funnel
3. filter paper
4. small test tube
5. Ringstand with ring clamp
6. dropper bottle of Lugol’s Iodine (IKI )
7. copper sulfate, starch & charcoal
suspension
Procedure
1. Fold the filter paper into a “triangle” by folding the circle of filter paper in half and in half
again. Pull three of the four layers in one direction to open the filter paper into the
shape of a cone. Place the cone into the funnel. Use a few drops of water on each side
to help it stick to the funnel.
2. Place the funnel in the ring clamp and place the empty beaker under the funnel.
Position the filter several inches above the rim of the beaker.
3. VIGOROUSLY SHAKE THE copper sulfate, starch & charcoal suspension, then quickly fill the
filter paper cone. Do not fill above the filter paper.
4. Observe the filtrate leaving the funnel. As soon as it is possible to count individual
drops, do so for 15 seconds. Record that number: ______.
5. When the filter is about half full, again count the number of drops dripping from the
funnel stem in 15 seconds. Record that number: _______.
6. Determine which substances are in the filtrate and which have been retained by the
filter paper. Record your results in the following table.

Copper sulfate is blue. Visually check for its presence.

The charcoal consists of fine black particles. Visually check for its presence.

Test for starch using Lugol’s Iodine (IKI). Pour about 1 ml of the filtrate into a small
test tube and add 2 drops of Lugol’s Iodine. The appearance of a dark blue-black
color indicates the presence of starch.
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Substance
Retained in Filter Paper
Copper Sulfate
Filtrate
X
Charcoal
X
Starch
X
1. Refer to your data in steps 4 and 5. Did the number of drops increase or decrease in
your second reading? ___Decrease___. Propose a reason for your answer.
With half the fluid gone, there was less pressure driving molecules across the
filter paper (it’s also true that some of the pores were blocked).
2. From your table, which items were small enough to pass through the filter paper?
Copper sulfate (and water of course).
Which were too large to pass through?
Starch and charcoal.
3. Suggest a reason why it is normal to find electrolytes but not proteins in urine.
Electrolytes are ions such as sodium and chloride, and are small enough to pass
through the filtration membrane in the kidneys; proteins are generally too large.
4. Consider the last two experiments you did. If the glass tube in the osmosis experiment
were both thicker in diameter and much longer, could the force of gravity on the liquid in
the tube eventually cause enough pressure to oppose osmotic pressure? _Yes__
Using the terminology and concepts from today’s lab, explain what would be happening in
the situation described in this question.
Initially, osmotic pressure would be strong enough to force
fluid uphill against gravity. However once the weight of fluid in the tube
exerted hydrostatic pressure equal to osmotic pressure, movement would stop.
5. Is there a situation in the human body like the one described in the previous question?
(Hint: the answer is yes  and has something to do with why you need a lymphatic
system.) Explain:
Yes. Hydrostatic pressure forces fluid out of our capillaries and into our tissues,
and osmotic pressure pulls some of it back. The lymphatic system picks up the rest.
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