This workbook accompanies the SimBio Virtual Labs® Diffusion laboratory

26 Jun This workbook accompanies the SimBio Virtual Labs® Diffusion laboratory

Question
SimBio Virtual Labs®

OsmoBeaker®: Diffusion
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SimBio Virtual Labs®

Diffusion
Introduction
The best chocolate shop in Amsterdam is partway down a side street in the downtown area. (It’s
called Puccini, if you ever visit.) They make the chocolate right in the store and have a beautiful
display counter that’s pretty much irresistible once you step inside. But how do they persuade people
to come look? Well, the kitchen in back where the chocolate is cooking has quite an enticing smell,
which is piped out to the street. When chocolate is being made, your nose knows all the way at the
corner of the block, and it’s the rare person who could resist sniffing along down the side street to
determine the source of the aroma.
Whenever you smell something, it means that molecules of whatever it is you smell have traveled to
and entered your nose. (WARNING: you might not want to think about this too much!) The outflow
pipe of the Puccini chocolate shop was designed so it releases molecules of chocolate to the outside
air. However, on a still day with no breezes, how can those molecules make it all the way to your
nose when you’re at the end of the block?
The movement of molecules is important for olfaction (sensing via smell), and to most other biological
systems as well. For example, oxygen molecules must move from your lungs to your blood vessels,
and from your blood vessels to the inside of cells on the other side of the blood vessel walls. Within
a single cell, molecules such as ATP (the main energy source for cellular processes) are made in
one place (mitochondria for ATP) but used in many others. All of these processes, from chocolate
attractant to oxygen transfer to ATP delivery, occur through a physical process called diffusion.
On one level, diffusion is intuitive. If some type of molecule is concentrated in a certain area
(e.g., chocolate molecules exiting an outflow pipe), those molecules will spread out over time. But
why, exactly, does diffusion of molecules happen? It turns out that many people (and even many
biology textbooks!) are confused about how diffusion works. This lab explores how diffusion works
using a series of simulations of different biological systems.

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SimBio Virtual Labs® | Diffusion

Exercise 1: Diffusion Across a Cell Membrane
The cell membrane (also known as the “plasma membrane”) is a very thin structure that surrounds
the cell’s cytoplasm and keeps its organelles in place. It is the outermost part of animal cells, and is
situated just inside the cell wall in plant cells. The cell membrane is composed of a phospholipid
bilayer (lipid = fat; bilayer = two layers) with embedded proteins. Certain proteins that cross the
bilayer are important for actively transporting specific ions and small molecules into and out of the
cell. Because the cell membrane is semi-permeable, some very small molecules can move passively
into and out of the cell.
One of the cool features of cell membranes is that they function more as flowing liquids than as
solids. This is due to the structure of the phospholipids in the membrane. The phospholipids have two
distinct ends: a hydrophilic head that is attracted to water and two hydrophobic tails that are repelled
by water. The phospholipids react to the watery environments inside and outside of the cell by
orienting their hydrophilic heads towards the water on the interior and exterior of the cell and their
tails away from the water. This reaction of phospholipids to water generates the liquid-like bilayer
structure of cell membranes, an important feature both for cellular transport and for cell recognition.

Figure 1. Schematic diagram of a cell membrane

Exercise 1 explores how molecules passively diffuse across cell membranes. The experimental set-up
involves an artificial membrane that has been stretched across the middle of a beaker, dividing the
left and right sides. The beaker contains two types of molecules; peptides (small proteins) and water.

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SimBio Virtual Labs® | Diffusion

[ 1 ]

If you haven’t already, start SimUText® by double-clicking the program icon on your computer or
by selecting it from the Start menu.

[ 2 ]

When the program opens, enter your Log In information and select the Diffusion lab from your My
Assignments window.

[ 3 ]

Select Artificial Membrane from the Select an Exercise menu at the top of the screen.

[ 4 ]

The lab will open showing the Beaker view, with a membrane stretched down the middle of a
beaker separating the left and right sides. The left side will be filled with orange peptide molecules.
The right will be filled with blue water molecules.

[ 5 ]

Run the simulation by clicking the GO button in the Control Panel bar at the bottom of the screen.
You can speed up the action by increasing the temperature using the Temperature slider in the
bottom panel. Watch some of the peptide molecules. As you watch the molecules, try to figure out
whether individual molecules tend to move in any particular direction, or whether they are just
moving randomly.

[ 6 ]

If you watched long enough, hopefully you concluded that the individual molecules did not follow
any pattern; rather, they moved randomly—that is, at any given instant, if you were to pick a
molecule to watch, it might be moving up, down, left, or right, with equal probability. There are
a total of 40 peptide molecules on the left and 80 water molecules on the right. Think about and
then write answers to the following two questions in the spaces provided.
[ 6.1 ]

[ 6.2 ]

[ 7 ]

If you could add channels to the membrane that make it permeable to peptides and
water, what do you think would happen to the peptide and water molecules in the
cell?

If, after adding those channels, you were to wait long enough for the system to
equilibrate (i.e., reach a stable state in which the number and type of molecule in
each compartment no longer changes much over time), about how many peptide
molecules would you expect to find on the left side of the cell and how many on the
right? How about the water molecules? Explain your reasoning.

In the Control Panel, find and click the RESET button (circular arrow) to re-initiate the simulation.
Then find the two checkboxes under the Membrane Permeability heading beneath the Beaker.
Checking WATER adds channels that make the membrane permeable to water and checking
PEPTIDE adds channels that make the membrane permeable to peptides.

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3

SimBio Virtual Labs® | Diffusion

[ 8 ]

Add channels to make the membrane permeable to both water and peptides by clicking on both
checkboxes. The channels that appear in the membrane will conveniently match the color of the
molecules that can pass through them.

[ 9 ]

Click the GO button and watch until the system equilibrates (i.e., stops changing much over
time). Monitor the Current Count status numbers in the panels under the Cell View to help you
determine when the counts stabilize, which may take a minute or two. Then click the STOP button.
[ 9.1 ]

After the system equilibrates, do the individual molecules follow the same or different
patterns of movement than when the system was not at equilibrium?

[ 9.2 ]

Were your predictions in Question [ 6.2 ] correct?

Hopefully you saw that the random movement of individual molecules resulted in molecules moving
from areas of higher concentration to lower concentration until the concentration differences
disappeared. A common misconception is that this happens because the molecules “want” or “tend”
to move where there is space to spread out. But as you just saw, the behavior of each molecule was
the same regardless of whether the membrane was sealed or was perforated. The only “rule” the
molecules were following was to move. As long as there were more peptide molecules on the left
side of the membrane, the probability that peptide molecules would move from left to right was
greater, even though the movement was purely random. Over time, the number of peptide molecules
in each compartment evened out, and thus the chance a peptide molecule would cross from right to
left or from left to right was the same.
[ 10 ]

Click the TEST YOUR UNDERSTANDING button at the bottom right corner of the screen. You can
click as many answers as you like to make sure that you understand why only one is the correct
response.
[ 10.3 ] Molecules tend to shift from regions where they are in higher concentration to
regions where they are in lower concentration because (write out the correct onscreen
response in the space below):

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4

SimBio Virtual Labs® | Diffusion

Exercise 2: Moving by Diffusing

Figure 2. The structure of a neuron. The cell body receives signals from dendrites that have been stimulated. Axons then carry impulses
from the cell body to muscles. Like all cells, neurons have a nucleus that contains genes. Neurons also have cytoplasm, mitochondria, and
other organelles.

Exercise 2 focuses on neurons (nerve cells), some of which are the longest cells in the animal kingdom.
The cell bodies of the neurons that control your foot, for instance, sit at the base of your spine. Each
of these neurons has an extension called an axon that travels the length of your leg from your spine
to the muscles and nerves in your foot. Here is yet another case where your body needs to transport
a particular type of molecule. The cell body produces various molecules including peptides, which
are proteins used at the tip of the axon. Those peptides must somehow travel from your spine, down
the axon, and out to the axon tip in your foot, and they must do this quickly. Could peptides travel
the one-meter from cell body to axon tip simply by diffusion?
In the following experiments, you will explore how quickly diffusion can transport molecules different
distances by examining nerve cells with axons of varying lengths. In each case, a peptide molecule
must travel from the cell body where it is produced to the end of the axon where it is used. You will
use your data to determine whether diffusion time is directly proportional to distance. If diffusion
time is directly proportional to diffusion distance, this means it will take twice as long for molecules
to diffuse twice as far, three times to diffuse three times as far, and so on. All three graphs below are
examples of two variables (X and Y) being directly proportional.

Figure 3. Three graphs showing directly proportional relationships between X and Y. For instance, in the middle graph, X is always half of Y
for any point (X, Y) on the line.

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SimBio Virtual Labs® | Diffusion

[ 1 ]

Select Axon 15 from the Select an Exercise menu at the top of the screen.

[ 2 ]

You should see a short axon, of length 15 µm (µm = micrometer = 1 millionth of a meter), stretching
to the right of a nerve cell body. The cell body is producing orange peptide molecules. Your Axon
Viewer (the box on top of the axon) shows the peptide molecules on the left and lots of blue water
molecules throughout the axon.

[ 3 ]

Run the simulation by clicking the GO button and watch to see how the molecules move around.
Then click the STOP button.

[ 4 ]

Click the RESET button. Next you will time how long it takes a peptide molecule to move from
the left to the right side of the cell. Click GO to run the simulation. This time, pay attention to the
Stopwatch at the top of your Axon Viewer. The Stopwatch automatically stops as soon as one of
the peptide molecules reaches the right side of the axon. Once the Stopwatch displays a number,
you can click STOP.
[ 4.1 ]

How long did it take? [NOTE: for the following steps, don’t worry about the time units,
just record the numbers.]

[ 4.2 ]

Repeat step 4 four additional times to obtain a total of 5 estimates of the time it takes
peptides to diffuse across the axon. Record each of the 5 estimates (including the one
in the space above) in the 15µm row of the table below and calculate and record their
average. [NOTE: there is a CALCULATOR tool in the bottom panel on your screen.]
PEPTIDE DIFFUSION DATA TABLE
AXON SIZE

DIFFUSION TIME

Trial:

1

2

3

4

5

AVG*

0 µm

–

–

–

–

–

0

15 µm
30 µm
60 µm

[ 4.3 ]

[ 4.4 ]

In the table above, circle the shortest and the longest diffusion times you recorded for
the 15 µm axon. If molecules always traveled the same predictable paths instead of
moving randomly, would you expect these numbers to be the same or different?

Based on the two numbers you circled, why do you think you obtained more than one
estimate (i.e., why did you replicate the experiment)?

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SimBio Virtual Labs® | Diffusion

[ 5 ]

Select Axon 30 from the Select an Exercise menu. The nerve cell in this scenario has an axon
that is twice as long (30 µm) as the previous one but is otherwise identical. As before, peptide
molecules are produced on the left side of the cell.
[ 5.1 ]

[ 6 ]

Repeat step 4 to determine how long it takes the peptide to move across this cell by diffusion.
[ 6.1 ]

[ 7 ]

Record your results in the 30 µm row of the PEPTIDE DIFFUSION DATA TABLE in Step 4.

Select Axon 60 from the Select an Exercise menu. The axon of this nerve cell is 60 µm, four times
as long as the first one (and twice as long as the second one).
[ 7.1 ]

[ 8 ]

If the relationship between diffusion time and distance is directly proportional (e.g.,
refer back to Figure 3), based on the average time it took peptides to diffuse 15µm,
how long do you predict it should take peptides to diffuse across this 30 µm axon?
You can use the onscreen calculator, but show the values you used in your calculation
in the space below (e.g., ( ) x ( ) = __ ) :

If the relationship between diffusion time and distance is directly proportional,
based on the average time it took peptides to diffuse 15 µm, how long
do you predict it should take peptides to diffuse across this 60 µm axon?
Again, show the values used in your calculation.

Once again, repeat step 4 to determine how long it takes the peptide to move across this cell by
diffusion.
[ 8.1 ]

[ 8.2 ]

Do the molecules always move from left to right or do they sometimes travel back to
the left again?

[ 8.3 ]

Record your results in the 60 µm row of the PEPTIDE DIFFUSION DATA TABLE in Step 4.
As you are running the simulation, pay attention to how individual peptide molecules
move.

Using the graph paper on the next page, plot your predicted diffusion time values for
the 30 µm and 60 µm axons using r‘s. (These are the values you calculated in Questions
5.1 and 7.1 above, that were based on the assumption that diffusion time and distance
are directly proportional.) Include the point (0,0) since it should take no time to travel
no distance. Draw the trend line for predicted diffusion times by connecting the r‘s
and label the trend line “predicted.”

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SimBio Virtual Labs® | Diffusion

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SimBio Virtual Labs® | Diffusion

[ 8.4 ]

Next, plot the average diffusion time for each axon length you recorded using =’s. You
can plot the point (0,0) on the graph along with your 3 data points. Draw the trend
line for your data by connecting the =’s and label the line “observed”.

[ 8.5 ]

Do your observed data support the prediction that diffusion time and distance are
directly proportional? Explain why or why not, and if not, considering how long it
took peptides to diffuse 15 µm, did it take more time or less time than expected for
peptides to diffuse 30 and 60 µm?

[ 8.6 ]

Based on your graphed data, which of the below choices do you think would be true?
(circle one letter)
a.

b.

It would probably take a little more than twice as long for peptides to diffuse the
length of a 120 µm axon than the length of a 60 µm axon.

c.

[ 8.7 ]

It would probably take about twice as long for peptides to diffuse the length of a
120 µm axon than the length of a 60 µm axon.

It would probably take a lot more than twice as long for peptides to diffuse the
length of a 120 µm axon than the length of a 60 µm axon.

How do your data support your choice in Question [ 8.6 ] above? [Hint: was diffusion
time consistently or increasingly longer for consistently increasing distances?]

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9

SimBio Virtual Labs® | Diffusion

[ 9 ]

Let’s say you were to run and then stop the simulation as soon as the first peptide reached the 30
µm tickmark, as depicted in A and B, below. One peptide has traveled halfway across the axon in B,
so it might seem that it should take the same amount of time to diffuse the remaining distance.
However, you know from your experiments that it takes more than twice as long for peptides to
diffuse 60 µm than 30 µm. Play with the simulation to convince yourself why this is the case, and
then answer the questions below.
A. Initial Conditions:

B. Conditions when first peptide has reached the 30 µm tickmark:

[ 9.1 ]

[ 9.2 ]

Why is the relationship between diffusion distance and time NOT linear? [Hint: In
figure B, how does the concentration of peptides at 30 µm compare to the initial
concentration at 0 µm in figure A? The correct answer has to do with concentration
and how molecules move when they diffuse.]

The cell bodies of neurons controlling your feet are actually located in your spine. The
cell body is where they make peptides such as the ones in this simulation. Their axons
can be a meter long, traveling out to the tips of your toes (about 1,000 times as long as
the 60 µm axon in the simulation). Do you think these neurons use diffusion to move
peptides from the cell body to the axon tip? Explain your reasoning.

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10

SimBio Virtual Labs® | Diffusion

To review, molecules in a solution are continuously moving and bouncing off other molecules. The
direction that a molecule moves is random and constantly changing. If a large number of molecules
are concentrated in one part of a solution, many of them will be moving in each direction at any
given time. If a neighboring area has very few solute molecules, only a few will be moving in
each direction at any given time. Through purely random motion, more solutes will, on average, be
moving out of the highly concentrated area in the direction of the low concentration area than the
other way around. This random motion will tend to equalize the concentrations in both areas, and
that effect is called diffusion.
The axon you’ve been experimenting with shows this pattern—the peptides start on the left side
and move every which way. Some move to the left and some to the right. Since initially there are
no peptides on the right, this process can only lead to more peptides on the right and fewer on the
left, until eventually the concentration of molecules is similar throughout. [Note: since the process
is random, even after a long time the concentration of peptides on either side may not be exactly
the same. Laws of probability predict that the numbers on either side will even out, but just like it
is possible to toss a coin 10 times and get 3 heads and 7 tails rather than the expected result of 5 of
each, it’s possible that you saw different numbers of molecules on the two ends of the axon when you
stopped the model and counted.]
After the molecules in an initially heterogeneous solution (i.e with local concentration differences)
have diffused for a long time and the solution becomes homogeneous, the solution is said to be in
its equilibrium state (i.e., no longer changing). The molecules will continue to move as before, but
without concentration differences, the movements should not cause consistent net changes on either
side. You should have seen this in the axons in your experiments; once the peptides reached the
right-most edge, they did not stop there. Sometimes they stayed close to that edge and sometimes
they moved back to the left. But on average, after running the model for a long time, there should
have been just as many peptide molecules on the left as on the right.
[ 10 ]

Make sure you are still on the Axon 60 exercise screen. Then click the TEST YOUR UNDERSTANDING
button in the bottom right corner of the screen.
[ 10.1 ] Which choice is the correct answer?

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11

SimBio Virtual Labs® | Diffusion

Exercise 3: Diffusion Challenge
Green plants are critical to life on earth, and their existence depends on diffusion. Plants convert
energy from the sun into useful chemical energy via photosynthesis, a process that requires a way to
take in carbon dioxide and to eliminate oxygen. The exchange of oxygen and carbon dioxide (as well
as the loss of water vapor in transpiration) occurs through special pores in plant leaves called stomata
(singular = stoma).
Stomata are interesting plant structures. They open when
light strikes leaves in the morning and close during the
night. Energy from sunlight changes ion concentrations
in the cells on the surface of leaves, which in turn causes
water to diffuse into the “guard cells” on either side of
the stoma. (This special type of diffusion is referred to as
“osmosis” and is the subject of another SimBio Virtual
Lab.) The inner wall of each guard cell is thick and
elastic. The influx of water creates pressure within the
guard cells, which causes their thin outer walls to bulge
out and forces their inner walls into a crescent shape.
This opens the stoma. When guard cells lose water, their
elastic inner walls regain their original shape and the
stoma closes.
In addition to opening and closing stomata, diffusion is
also used by plants to move gas molecules into and out
of the leaves through the stomata. Unlike the scenario
you explored with neurons, the distance that gases must
diffuse, even in large plants, is not great. Cells in plant
leaves are loosely packed, providing an interconnected
system of air spaces through which gases can diffuse.
Importantly, plants both produce and use gases as they
photosynthesize and respire, thus the concentration of
each gas within the leaf can be quite different than it is
in the air surrounding the plant.

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12

SimBio Virtual Labs® | Diffusion

The final set of experiments investigates whether a molecule of a gas starting out in a region where
the substance is concentrated will diffuse faster than a molecule of the gas starting in a region where
it is less concentrated.
[ 1 ]

Choose Diffusion Challenge from the Select an Exercise menu at the top of the screen.

[ 2 ]

Examine the cross section view of leaf tissue on the right side of the screen. If you could look
through a leaf from the side under a microscope, these are some of the structures you would
see. Notice how the loose packing of cells creates air spaces. If you were to look at the leaf from
the bottom under a microscope, you’d see the stomata, depicted below the cross section view.
Remember that water vapor and CO2 molecules enter and exit leaves through these openings.

[ 3 ]

The left side of the screen shows cross-sections of slices from five different leaves, each one
representing one experimental leaf section, with stomata closed. CO2 molecules are represented
by large balls, and water vapor molecules are represented by smaller (blue) balls. The water vapor
and CO2 molecules in the simulation will behave identically. You should notice that two of the CO2
molecules in each leaf section have different labels; one labeled “A”at the top of the leaf starts out
in a region of low CO2 concentration, and one labeled B” at the bottom starts out in a region of
high CO2 concentration (surrounded by other CO2 molecules.
[ 3.1 ]

[ 4 ]

Run the simulation by clicking the GO button and watch the molecules move around, paying
attention to how the different types of molecules respond to collisions. The stopwatches at the top
and bottom of each leaf section will record when the marked CO2 molecule from the opposite side
has traveled all the way from one side of the leaf to the other and collided against the opposite
wall. When this occurs, the stopwatch will mark the time and display an “A” or “B” indicator.
[ 4.1 ]

Considering the initial configurations of CO2 molecules in the leaf sections, do you
predict that either the “A” or the “B” CO2 molecu…