Membrane Permeability

Totty, Reza and Dhila

P05 Savero Jendrizzy



PhysioEx


TM

PhysioEx is a simulation for cell transport mechanism, permeability and

membrane potential. It’s like an online Frozen dress up game but about physiology
and there’s many number and I hate it. If you don’t have anything to do and you
want to try it you can access it at http://www.physioex.com/login.html

Username: physiologyui
Password: musculo1819


A. Simple Diffusion
All molecules possess kinetic energy and are in constant motion. As molecules move about
randomly at high speeds, they collide and bounce of one another, changing direction with each
collision. Smaller molecules tend to move faster than larger molecules because kinetic energy is
directly related to both mass and velocity (KE = ½ mv2).
Diffusion is the movement of molecules from a region of its higher concentration to lower
concentration, and the driving force behind it is the kinetic energy of the molecules themselves.
It occurs because there is a concentration gradient – the molecules will move so that they are
evenly distributed throughout the environment.
Molecules can diffuse passively through the plasma membrane if they are small enough to
pass through the pores (aided by an electrical and/or concentration gradient). A membrane is
called selectively permeable/differentially permeable/semipermeable because they allow some
solute particles to pass but not others.
Simple diffusion is the diffusion of solute particles dissolved in water through a selectively
permeable membrane, while osmosis is the diffusion of water through a differentially permeable
membrane. Both of them involve movement of a substance from a region of its higher
concentration to lower concentration.
The experiment will try out the diffusion of molecules with different molecular weight
(sizes) across several membranes with different MWCO value (Molecular Weight Cut Off); the
largest molecular weight it can let through. The bigger the MWCO value, the bigger the pore is,
allowing larger molecules to pass through.

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 TM

Picture 1.1 PhysioEx activity 1 interface

The instruction is on the left side of the screen, and based on it you have to click and drag to alter
the dialysis membrane and concentration of solute that is instructed, and after it you can record
the data of each experiment.

1) Procedure
1. Prepare as follows:

Picture 1.2 simple diffusion activity interface

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No. Beaker A Beaker B Dialysis membrane
+ -
1. Na Cl 9.00 mM Deionized water 20 MWCO
2. Urea 9.00 mM Deionized water 20 MWCO
3. Na+Cl- 9.00 mM Deionized water 50 MWCO
4. Na+Cl- 18.00 mM Deionized water 50 MWCO
5. Na+Cl- 9.00 mM Deionized water 100 MWCO
6. Urea 9.00 mM Deionized water 100 MWCO
7. Glucose 9.00 mM Deionized water 200 MWCO
8. Albumin 9.00 mM Deionized water 200 MWCO
Table 1.1 simple diffusion activity instructions
2. Click start to run the test.
3. Observe the concentration display to the side of each beaker for any activity.
4. Record the data.

2) Data Collected

No. Beaker A Beaker B Time to reach Average
concentration concentration equilibrium diffusion rate
1. 9.00 mM 0.00 mM - 0.0000
2. 9.00 mM 0.00 mM - 0.0000
3. 4.50 mM 4.50 mM 10 minutes 0.0150
4. 9.00 mM 9.00 mM 11 minutes 0.0273
5. 4.50 mM 4.50 mM 10 minutes 0.0150
6. 4.50 mM 4.50 mM 16 minutes 0.0094
7. 4.50 mM 4.50 mM 36 minutes 0.0042
8. 9.00 mM 0.00 mM - 0.0000
Table 1.2 data collected from simple diffusion activity tuh kan angkanya banyak beud
Note:
• The concentrations noted are the final concentration after diffusion has been
allowed to happen and equilibrium has been reached.
• The diffusion rate is not directly proportional to time as it is also affected by the
initial concentrations.

3) Conclusion
• Molecules cannot pass through the membrane if the membrane pores are too small
(MWCO values too low) – see experiment no. 1 and 2: Both Na+Cl- and Urea are
made of molecultes too big to pass the 20 MWCO pore.
• Concentration difference affects the rate of diffusion. Higher difference means
higher diffusion rate – see experiment no. 3 and 4: Experiment 4 has a higher
diffusion rate as it had an initial concentration difference of 18mM while experiment
3 had only 9mM difference.
• If a molecule can pass through a certain MWCO value, increasing the membrane
MWCO value will not change the rate of diffusion. Higher MWCO value does not

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 mean faster rate – see experiment no. 3 and 5: The difference in MWCO resulted in
the same diffusion rate as the concentration difference were both the same being

B. Facilitated Diffusion
Facilitated diffusion is the process of passive transport of molecules through the
membrane. Substance that are too large or not lipid soluble (e.g.: sugars, amino acids, ions) need
special protein to pass through the membrane called carrier proteins and channel proteins.
These special proteins are usually substance-specific (e.g.: GLUT4 - solute carrier family 2,
facilitated glucose transporter member 4), and only allows diffusion in one way, down the
concentration gradient. The greater the concentration difference between one side of the
membrane and the other, the greater the rate of diffusion.

TM
Picture 2.1 PhysioEx activity 2 interface

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1) Procedures
1. Prepare as follows:

Picture 2.2 facilitated diffusion activity interface

No. Beaker A Beaker B Membrane glucose
carriers
1. Glucose 2.00 mM Deionized water 500
2. Glucose 8.00 mM Deionized water 500
3. Glucose 2.00 mM Deionized water 700
4. Glucose 8.00 mM Deionized water 700
5. Glucose 10.00 mM Deionized water 100
6. Glucose 2.00 mM Na+Cl- 2.00 mM 700
Table 2.1 facilitated diffusion activity instructions
2. Click start to run the test.
3. Observe the concentration display to the side of each beaker for any activity.
4. Record the data.

2) Data Collected
No. Beaker A Beaker B Time to reach Average
concentration concentration equilibrium diffusion rate

1. 1.00 mM 1.00 mM 43 minutes 0.0008

2. 4.00 mM 4.00 mM 58 minutes 0.0023
3. 1.00 mM 1.00 mM 33 minutes 0.0010
4. 4.00 mM 4.00 mM 43 minutes 0.0031

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 5. 6.824 mM 3.176 mM - 0.0028
Glucose 1.00 Glucose 1.00 33 minutes 0.0010
6. mM, Na+Cl- mM, Na+Cl- 1.00 (glucose), 8 mins (glucose),
1.00 mM mM (Na+Cl-) 0.0042 (Na+Cl-)
Table 2.2 data collected from facilitated diffusion activity

3) Conclusion
• Concentration difference affects the rate of diffusion. Higher difference means
higher diffusion rate – see experiment no. 1 and 2: Experiment 2 had a higher
difference hence higher diffusion rate
• There is no correlation between having a different solute in the experiment (NaCl)
and the rate of diffusion – see experiment no. 3 and 6: Even though experiment 6
has NaCl, both experiments produced the same rate of diffusion, as the
concentration difference of Glucose was the same.

C. Osmotic Pressure
A special form of diffusion, called osmosis, is the diffusion of water through a selectively
permeable membrane down its water potential gradient. Because water can pass through the
pores of most membranes, it can move from one side of the membrane to the other relatively
freely. Osmosis takes place whenever there is a difference in water concentration between two
sides of a membrane. More solute dissolved means less water concentration. In other words,
water will move towards the region with higher dissolved solute concentration (water moves
down its own concentration gradient).
Osmotic Pressure is defined as the minimum pressure that must be applied to prevent
the movement of the water molecules. In a closed container, it is also the pressure that is
exerted by the water to the container due to the increasing volume. It is usually measured with
millimeters of mercury (mm Hg). Generally, more permeable membrane results in smaller
osmotic pressure.

Picture 3.1
TM
PhysioEx
activity 3
interface

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 1) Procedures
1. Prepare as follows:


















Picture 3.2 osmotic pressure activity interface

No. Beaker A Beaker B Dialysis membrane
+ -
1. Na Cl 5.00 mM Deionized water 20 MWCO
2. Na+Cl- 10.00 mM Deionized water 20 MWCO
3. Na+Cl- 10.00 mM Deionized water 50 MWCO
4. Glucose 8.00 mM Deionized water 100 MWCO
5. Glucose 8.00 mM Glucose 8.00 mM 100 MWCO
6. Glucose 8.00 mM Deionized water 200 MWCO
7. Albumin 9.00 mM Glucose 10.00 mM 200 MWCO
Table 3.1 facilitated diffusion activity instructions

2. Click start to run the test.
3. Observe the concentration display to the side of each beaker for any activity.
4. Record the data.

2) Data Collected
Beaker A Beaker B
No.
Concentration Pressure Concentration Pressure
1. 5.00 mM 170 mm Hg 0.00 mM 0 mm Hg
2. 10.00 mM 340 mm Hg 0.00 mM 0 mm Hg
3. 5.00 mM 0 mm Hg 5.00 mM 0 mm Hg
4. 8.00 mM 136 mm Hg 0.00 mM 0 mm Hg
5. 8.00 mM 0 mm Hg 8.00 mM 0 mm Hg
6. 4.00 mM 0 mm Hg 4.00 mM 0 mm Hg
Albumin 9.00 Albumin 0.00
7. mM + Glucose 153 mm Hg mM + Glucose 0 mm Hg
5.00 mM 5.00 mM
Table 3.2 data collected from facilitated diffusion activity astaghfirullah

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 3) Conclusion
• Concentration of solutes affects the osmotic pressure. The higher the
concentration, the higher the osmotic pressure. (see experiment no. 1 and 2)
• Present of special proteins affects the rate of diffusion. The more glucose carriers
or special proteins there are in the membrane, the faster the diffusion rate is. (see
experiment no. 1 and 3, or no. 2 and 4)
• Using different solutes also affects the osmotic pressure

D. Filtration
Filtration is a passive process which water and solutes pass through a membrane from an
area of high hydrostatic pressure to low hydrostatic pressure. For example, fluid and solutes
filter out of the capillaries in the kidneys into the kidney tubules because blood pressure in the
capillaries is greater than the fluid pressure in the tubules, so if blood pressure increases, the
rate of filtration increases.
Filtration is not a selective process. The amount of filtrate – the fluids and solutes that pass
through the membrane, depends almost entirely on the hydrostatic pressure gradient and the
size of membrane pores (Molecular Wight Cutoff). Solutes that are too large to pass through
are retained by the capillaries. Ions and smaller molecules like glucose and urea can pass
through, while large solutes like blood cells and proteins cannot pass through.

TM
Picture 4.1 PhysioEx activity 4 interface

1) Procedures
1. Prepare as follows:

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Picture 4.2 filtration activity interface

No. Top beaker Pressure Dialysis membrane
Na+Cl- 5.00 mg/ml
Urea 5.00 mg/ml
1. Glucose 5.00 mg/ml 50 mm Hg 20 MWCO
Powdered charcoal
5.00 mg/ml
Na+Cl- 5.00 mg/ml
Urea 5.00 mg/ml
2. Glucose 5.00 mg/ml 50 mm Hg 50 MWCO
Powdered charcoal
5.00 mg/ml
Na+Cl- 5.00 mg/ml
Urea 5.00 mg/ml
3. Glucose 5.00 mg/ml 50 mm Hg 200 MWCO
Powdered charcoal
5.00 mg/ml
Na+Cl- 5.00 mg/ml
Urea 5.00 mg/ml
4. Glucose 5.00 mg/ml 100 mm Hg 200 MWCO
Powdered charcoal
5.00 mg/ml
Table 4.1 filtration activity instructions
2. Click start to run the test.
3. Observe the concentration display to the side of each beaker for any activity.
4. Record the data.

2) Data Collected
No. Solute Start Filter Filter rate

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 concentration concentration
Na+Cl- 5.00 mg/ml 5.00 0.00
Urea 5.00 mg/ml 5.00 0.00
1. Glucose 5.00 mg/ml 5.00 0.00 1
Powdered charcoal
5.00 0.00
5.00 mg/ml
Na+Cl- 5.00 mg/ml 5.00 4.81
Urea 5.00 mg/ml 5.00 0.00
2. Glucose 5.00 mg/ml 5.00 0.00 2.5
Powdered charcoal
5.00 0.00
5.00 mg/ml
Na+Cl- 5.00 mg/ml 5.00 4.81
Urea 5.00 mg/ml 5.00 4.74
3. Glucose 5.00 mg/ml 5.00 4.39 10
Powdered charcoal
5.00 0.00
5.00 mg/ml
Na+Cl- 5.00 mg/ml 5.00 4.81
Urea 5.00 mg/ml 5.00 4.74
4. Glucose 5.00 mg/ml 5.00 4.39 20
Powdered charcoal 5.00 0.00
5.00 mg/ml

Table 3.2 data collected from filtration activity

3) Conclusion
• Hydrostatic pressure affects the rate of filtration. The higher the hydrostatic
pressure, the higher the filtration rate. (see experiment no. 3 and 4)
• Size of membrane pores affects the rate of diffusion. The higher the molecular
weight cutoff, the higher the filtration rate. (see experiment no. 1, 2 and 3)

E. Active Transport
Active transport is the process of moving substances across its membrane against its
concentration gradient using energy (ATP). There are several reasons that a substance might not
be able to pass through a membrane by diffusion: it might be too large to pass, it might not be
lipid soluble, it might have to move against its concentration gradient.
The process is enabled by protein carriers called solute pumps. The solute pumps aid in
the movement of the molecules from lower to higher concentration (against its concentration
gradient) with the help of ATP. Solute pumps are usually solute specific, meaning they only
transport a certain molecule, also called a uniporter. However, there are other types, for
example the carrier protein for Na+ ions and K+ ions in cell membranes. These carrier proteins
are said to participate in coupled transport. For protein carriers that move the solutes in
opposite directions are called antiporters. Whereas protein carriers that transfer two molecules
in the same direction are called symporters.

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 TM

Picture 5.1 PhysioEx activity 5 interface
1) Procedures
















Picture 5.2 active transport activity interface

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 1. Prepare as follows:

No. Solutes Na+-K+ Glucose
ATP
Intracellular Extracellular pumps carriers
Na+Cl- 9.00 K+Cl- 6.00
1. 1 mM 500 -
mM mM
Na Cl 9.00 K+Cl- 6.00
+ -
2. 3 mM 500 -
mM mM
+ -
Na Cl 9.00 Deionized
3. 3 mM 500 -
mM water
Na Cl 9.00 K+Cl- 6.00
+ -
4. 3 mM 800 -
mM mM
K+Cl- 6.00
Na+Cl- 9.00 mM,
5. 3 mM 800 400
mM Glucose
10.00 mM
Table 5.1 active transport activity instructions
2. Click start to run the test.
3. Observe the concentration display to the side of each beaker for any activity.
4. Record the data.


2) Data Collected
No. Solutes Time for
Rate
Intracellular Extracellular transport
+ -
Na Cl 5.61 Na+Cl- 3.39
1. 3 minutes 0.0188
mM mM

K Cl 2.26 mM K+Cl- 3.74 mM
+ -
3 minutes 0.0125
Na+Cl- 0.00 Na+Cl- 9.00
60 minutes 0.0025
2. mM mM
K+Cl- 6.00 mM K+Cl- 0.00 mM 60 minutes 0.0017
Na+Cl- 9.00 Na+Cl- 0.00
3. - 0.0000
mM mM
+ -
Na Cl 0.00 Na+Cl- 9.00
36 minutes 0.0042
4. mM mM
K Cl 6.00 mM K+Cl- 0.00 mM
+ -
36 minutes 0.0028
Na+Cl- 0.00 Na+Cl- 9.00
36 minutes 0.0042
mM mM
5. K+Cl- 6.00 mM K+Cl- 0.00 mM 36 minutes 0.0028
Glucose 4.95 Glucose 5.05
- 0.0028#
mM mM
Table 5.2 data collected from active transport activity penderitaan belum berakhir

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3) Conclusion
• Concentration of ATP affects the active transport. The higher the concentration, the
more ions are transported. (see experiment no. 1 and 2)
• Present of special proteins affects the rate of transport. The more Na+-K+ pumps or
solute pumps there are in the membrane, the faster the transport rate is. (see
experiment no. 2 and 3)
• Adding other solutes does not affect the rate of transport. Adding glucose does not
disrupt the Na+ or K+ transport, because the pumps that are used to transport the
ions are solute specific. (see experiment no. 4 and 5)

F. Resting Membrane Potential

Resting Membrane Potential is the difference in charge between inside and outside the
cell. It depends on the permeability of the membrane to the ions (K+ ions and Na+ ions for
neurons) and their concentrations. In neurons, the Na+ ion concentration inside the cell is low
and the K+ ion concentration is high (Na+ ~5 mM, K+ ~150 mM). Outside the cell the
concentration of Na+ ion is high while K+ ion is low (Na+ ~150 mM, K+ ~5 mM). This condition is
regulated by the ion pumps (Na+-K+ pump. Note that there are K+ ion leak channel, allowing the
diffusing of the ions. Combined with the active transport of the ion pumps, it maintains the
resting membrane potential. The extracellular environment is relatively positive compared to
intracellular environment—due to the presence of large anions that are unable to pass through
the membrane—hence, the negative resting potential value (-70 mV).
The experiment tests the effects of changing extracellular environment, in regards to the

ion concentrations. If the extracellular environments have a high concentration of K+ ions, less K+

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 ion will diffuse out of the leak channels, resulting in a less negative potential difference (-40
mV). In extracellular environments with low Na+ ions, less Na+ ion will diffuse into the cell,
making the potential difference more negative (-72 mV). However, the effect on Na+ ions is not
as significant as the change in K+ ion concentration. This means that the carrier proteins for the
Na+ ions are mostly closed.
TM
Picture 6.1 PhysioEx activity 6 interface


1) Procedures
1. Prepare as follows:

Picture 6.2 resting membrane potential activity interface


Membrane potential from microelectrode position
No. Extracellular Fluid Cell body Axon
Intracellular Extracellular Intracellular Extracellular
Control ECF
(5 mM K+, 150 mM
1.
Na+)

High K+ ECF
(25 mM K+, 130
2.
mM Na+)

Low Na+ ECF
3. (5 mM K+, 30 mM
Na+, 120 mM

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 TmA+)

Table 6.1 resting membrane potential activity instructions
2. Click start to run the test.
3. Observe the concentration display to the side of each beaker for any activity.
4. Record the data.


2) Data Collected
Voltage from microelectrode position
No. Extracellular Fluid Cell body Axon
Intracellular Extracellular Intracellular Extracellular
Control ECF
(5 mM K+, 150 mM
1. -70 mV 0 mV -70 mV 0 mV
Na+)

High K+ ECF
(25 mM K+, 130
2. -40 mV 0 mV -40 mV 0 mV
mM Na+)

Low Na+ ECF
(5 mM K+, 30 mM
3. Na+, 120 mM -72 mV 0 mV -72 mV 0 mV
+
TmA )

Table 6.2 data collected from resting membrane potential activity

3) Conclusion
• The concentration of Na+ and K+ outside of the cell affects the resting membrane
potential.
• The number of leak channels and open carrier proteins that are available in the cell
membrane affects the resting membrane potential. Manipulating the concentration
of K+ outside of the cell will change the resting membrane more significantly than by
manipulating the concentration of Na+, because K+ have more leak channels and the
carrier proteins for Na+ are mostly closed. (See experiment no. 2 and 3)

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Investigating Diffusion


Through Living Membranes



1) TL;DR Procedure
v Two deshelled eggs (same size) are soaked in glucose 30% sucrose solution and distilled
water with equal volume for each egg
v The mass of the eggs are measured every 20 minutes for 1 hour and calculate the
percent change.

2) Data Collected
v The egg mass in 30% sucrose solution is decreasing and the colour is whiter as it
shrinks
v The egg mass in distilled water is increasing and become more transparent as it gained
more volume (just like a balloon)

3) Conclusion
v The process of osmosis happened in the egg
v The egg in 30% sucrose solution loses its water content because the egg osmotic
pressure is higher than the solution osmotic pressure. This is because the solute
concentration of the solution is higher than the solute concentration of the egg
(hypertonic solution).
v The egg in distilled water gained more water content because the egg osmotic
pressure is lower than the solution osmotic pressure. This is because the solute
concentration of the solution is lower than the solute concentration of the egg
(hypotonic solution)

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The Influence of Different



Conditions on Erythrocytes

1. Procedure
v Place a very small drop of Physiologic saline on a slide.
v Using the medicine dropper, add a small drop of blood to the saline on the slide. Tilt the
slide to mix, cover with a cover slip, and immediately examine the preparation under
the high-power lens. Observe, write down what happened.
v Prepare another wet mount of blood, but this time use a 5% sodium chloride solution as
a suspending medium. Carefully observe the red blood cells under high-power lens.
Write down what happened.
v Add a drop of distilled water to the edge of the coverslip. Fold a piece of filter paper in
half and place its folded edge at the opposite edge of the coverslip. It’ll absorb the
saline solution and draw the distiller water across the cells.
v Watch the red blood cells as they float across the field. Describe the change in their
appearance.
TL;DR : basically you just need to put your drop of blood in NaCl 5%, Physc. Saline, and in water. Put
the mixture in the slide, cover it with a cover slip, obsereve.

2. Outcome and Conclusion
a. 0.9 % Salt solution (Physiologic Saline)
Nothing happen to the erythrocyte. This because the condition is isotonic.
b. 5% NaCl Solution.
The cells are crenated (shrink) because the water inside the cells diffused out due to the
Hypertonic condition.
c. Distilled water
The cells are swollen because the water from the environment difused in to the cells due
to the Hypotonic condition. The cells can be lysis (busrt) if the diffusion continues.

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