Osmosis: The spontaneous movement of water through a selectively permeable

membrane from a region of high water potential to a region of low water potential.

Osmolarity: The total solute concentration which is expressed as osmoles per litre
of solution. Osmoles refers to the number of moles of a solution that contribute to
osmotic pressure of solution. For example, 1 mol/L NaCl solution has an osmolarity
of 2osmol/L. This is because NaCl dissociates fully in water into Na+ and Cl- ions.
Hence, the dissociation of NaCl in the solution results in 1 mol/L concentration Na+
and 1mol/L concentration of Cl-, and since Na+ and Cl- both contribute to the
osmotic pressure of solution, the solution as an osmolarity of 2osmol/L.

Tonicity: Tonicity is a measure of the osmotic pressure gradient (as a result of water
potential) of two solutions separated by a semi-permeable membrane. Tonicity is
only affected by solutes that cannot cross the membrane.

Wcell stands for the water potential of the cell. Water potential is the potential
energy of water per unit volume relative to pure water in reference conditions. Wpi
stands for the solute potential and Wp stands for pressure potential. Increasing the
concentrations of dissolved solutes lowers the water potential while pressure
exerted on the cell increases the water potential.

3. Animal cells do not have a cell wall, hence animal cells do not have a pressure
potential (nothing to exert pressure on cell).

4. Red blood cells are permeable to water but relatively impermeable to salts. Intact
erythrocytes transmit very little light when suspended in a liquid as each erythrocyte
acts as an opaque disc. This is a property of both the haemoglobin and also the size
and shape of erythrocytes. When cells hemolyze, the suspension will become
transparent. This characteristic allows one to determine the tonicity of various
solutions to red blood cells. Hence both of these properties of erythrocytes allow it
to be used in preference of other cells.
















5. Table of results for Haemolysis time of erythrocytes when placed in different
solution
Heamolysis time (s)
Solution that
RBC was
placed in
Trial 1 Trial 2 Trial 3 Mean
Heamolysis
time (s)
Distilled
water
2.0 1 1 1.7
Isotonic
saline
Non-
haemolytic
Non-
haemolytic
Non-
haemolytic
Non-
haemolytic
50mM NaCl
solution
5.0 6.0 7.0 6.0
100 mM NaCl
Solution
Non-
haemolytic
Non-
haemolytic
Non-
haemolytic
Non-
haemolytic
150 mM NaCl
solution
Non-
haemolytic
Non-
haemolytic
Non-
haemolytic
Non-
haemolytic

Note: All times given to 2 significant figures
Also, non haemolytic means there was no evidence of haemolysis after 2
minutes.

6. The difference in haemolysis times can be attributed to the differences in the
osmotic pressure gradient between inside the RBC and the solution that RBC were
placed in (distilled water and isotonic saline). The water potential of RBCs is the
same (or approximately the same) as the water potential of isotonic saline and
there should not be any net movement of water due to osmosis and as a result
haemolysis should not occur. This was recorded in the practical as non-haemolytic
after 2 minutes. However the water potential of distilled water is much higher than
of RBCs and hence there would be a net rapid movement of water into the RBC due
to osmosis which will quickly cause haemolysis.

7. The NaCl solution is dissociative as NaCl splits into Na+ and Cl- ions in water.
Hence the Na+ and Cl- ions each contribute to the osmolarity of the solution.
However the Na+ and Cl- ions are not permeant.

As a result the 50 mM solution of NaCl has a osmolarity of 100 mosM which is lower
than the osmolarity of RBCs hence the 50mM NaCl solution is hypotonic. This is
supported as haemolysis occurred with RBCs were placed in this solution.

The 100 mM NaCl solution therefore has osmolarity of 200 mosM, which is again
lower than osmolarity of RBCs hence the 100 mM NaCl solution is hypotonic. Even
though the results in the table say haemolysis did not occur after 2 minutes, it is
likely that the rate of haemolysis was slow in this solution and is likely to have
occurred at a time greater than 2 minutes.

The 150 mM NaCl solution therefore has osmolarity of 300 mosM, which is either
higher or equal to the osmolarity of the RBC (depends on the true osmolarity of the
RBC). Hence the 150 mM NaCl solution is isotonic or hypertonic. This is supported as
haemolysis did not occur within 2 minutes when RBCs were placed in this solution
(or crenation occurred but was not visible).



8. Distilled water is hypotonic compared to the red blood cells in the body while
isotonic saline is (isotonic). Hence there is a high water potential in the distilled
water and a lower water potential in the RBC (due to impermeant solutes in RBC). As
a result, water will flow into the RBCs due to osmosis ( net water flow from a region
of high water potential to a region of low water potential) and the RBCs will swell
and eventually burst (haemolysis).

9. Experimental variation could have arisen from the difference in volume of the
RBC drops that were placed in the cuvette. This may have affected the haemolysis
time if the volume of blood (hence the number of RBC)s in solution was different in
repeats of the experiment. This may have been improved by predeciding on a fixed
volume of blood to be added to the cuvette for each trial and this volume be
measured (e.g on a measuring cylinder) before being added to solution in cuvette.

Inconsistent determination of the end point could result in variation in results
potentially due to the method (e.g the time taken in covering the cuvette with
parafilm and placing cuvette in front of newspaper). This was more likely when using
solutions where the haemolytic time was very low ( 1-2s for distilled water). As a
result, it may have resulted in inconsistent end point determination that may have
resulted in variance. Increasing the number of repeats of the experiment and then
taking an average haemolytic time is likely to reduce this variance.

11. Table of results for the Haemolysis time of erythrocytes when placed in different
solution in Experiment 2.


Heamolysis time (s)
Solution that
RBC was
placed in
Trial 1 Trial 2 Trial 3 Mean
Heamolysis
time (s)
284 mM
sucrose
solution
Non-
haemolytic
Non-
haemolytic
Non-
haemolytic
Non-
haemolytic
1M urea
solution
4.0 4.0 4.0 4.0
284 mM
sucrose and
1 M urea
solution
Non-
haemolytic
Non-
haemolytic
Non-
haemolytic
Non-
haemolytic


When the RBCs were placed in the solution of 284 mM sucrose, there was no net
movement of solute as sucrose is impermeant. Also the solution of 284 mM sucrose
was isotonic compared to the RBCs as the total osmolarity inside the RBCs and
outside in the solution was approximately equal


When the RBCs were placed in the solution of 1M urea, there was a net movement
of urea into the cell down its concentration gradient by facilitated diffusion until it
has equilibrated across the membrane. Also the solution of 1 M urea was hypotonic
compared to the RBCs. This is because the total osmolarity inside the RBCs (1.284
osM) was higher than the total osmolarity outside in the solution (approximately 1
osM).

When the RBCs were placed in the solution of 284 mM sucrose 1M urea, there was a
net movement of urea into the cell down its concentration gradient by facilitated
diffusion until it has equilibrated across the membrane as urea is only permeant.
Also the solution of 284 mM sucrose 1M urea was isotonic compared to the RBCs.
This is because the total osmolarity inside the RBCs (1.280 +/- 20 osM) was
approximately equal to the total osmolarity outside in the solution (1.284 osM).


12. Initially, before the urea had equilibrated across the RBC membrane, there would
be a net movement of water from the RBC out to the solution and the RBC may have
become crenated. This is because the water potential of the RBC was initially higher
than the initial water potential of the solution , as the solution initially had a higher
total solute concentration. If this experiment was done under a light microscope , it
could be possible to determine that crenation of the RBC did initially occur.

13.
i) If the RBC are placed in a solution of 142nM sucrose and 77 nM NaCl, the
osmolarity of the solution that the RBCs are placed in is 296 mosM (as NaCL
dissociates in solution). Since the osmolarity of RBCS is 280 +/- 20 mosM, the
solution of 142nM sucrose and 77 nM NaCl is approximately isotonic to the red
blood cells. There will be approximately no net water movement and no solute
movement, as all the solutes are impermeant to the membrane. Hence the RBC
should stay the same.

ii) If the RBCs are placed in a solution of 284mM, where urea is a permeant solute,
urea will move across the cell membrane from the outer solution into the RBCs by
facilitated diffusion. After the urea concentration has equilibrated across the
membrane, the osmolarity of the RBCs will be 280 mosM + 284 mosM which equals
564 mosM (+/- 20 mosM) while the osmolarity of the solution will be 284 mosM. As
a result the solution is hypotonic to the RBCs and as a result water will move into the
cell by osmosis and hence the RBC will lyse.

iii) If RBCs are placed in a 1M urea, 154 mM NaCl solution, where urea will move
across the cell membrane from outer solution into the RBCs by facilitated diffusion,
until it has equilibrated across the membrane. After, the osmolarity of the RBC is
280mosM + 1osM which is 1.280 M ( +/- 20 mosM) while the osmolarity of the
solution is 1 osM + 308 mosM (as NaCl dissociates in solution and not permeable_
which equals 1.308 mosM. Hence the solution is hypertonic to the RBC. As a result, a
net flow of water will move out of the RBCs into the solution due to osmosis. As a
result, the RBCs will have crenated.