The implications of a hypertonic, hypotonic, and isotonic environment on human red blood cells

Josh Matthew R. Oronce

Biology Department, College of Science, De La Salle University
Email: josh_oronce@dlsu.edu.ph

ABSTRACT

Osmosis is the process wherein water molecules move from areas possessing a higher concentration of
solutes to areas with a lower concentration of solutes. Based on their osmotic pressure, a solution can
either be hypertonic, hypotonic, or isotonic. Red blood cells will swell when placed in hypotonic solution
and shrivel when placed in a hypertonic solution. This is because of osmosis which leads to a net
movement of water towards areas with a higher concentration. This study sought to observe the effects of
a hypertonic, isotonic, and hypotonic solution on a red blood cell. In order to ascertain whether the
hypothesis was true, the researcher prepared three slides. Slide 1 containing a 0.9% NaCl solution which
is isotonic relative to the red blood cell, slide 2 containing a 5% NaCl solution which is hypertonic
relative to the red blood cell, and slide 3 containing distilled water which is hypotonic relative to the red
blood cell. Blood was then collected from the subject and mixed with each slide. The slides where then
studied under a microscope in order to observe the effect of each solution on the red blood cell. The
results of the experiment showed that red blood cells became lysed when placed in hypotonic solution,
shriveled when placed in a hypertonic solution, and had no structural changes when placed in an isotonic
solution thus affirming the hypothesis of the study.

INTRODUCTION

The cell membrane divides the regions containing the intracellular fluids and the extracellular
fluids. Most solutes such as sugars and ions are not lipid soluble and therefore cannot pass through the
lipid bilayer. A concentration gradient of these solutes however can lead to a net movement of water
across the cell membrane. This happens because water molecules form hydration shells around polar
solute molecules and when a membrane separates two solutions with differing charges, the concentration
of free water molecules on the opposite sides of the cell membrane also differ. The side possessing a
higher solute concentration has more water molecules in hydration cells and thus has lesser free water
molecules. This difference leads free water molecules to pass through the membrane and move towards
the area with a higher solute concentration in a process known as osmosis. The concentration of solutes in
a solution dictates the osmotic concentration of a solution. If two solutions have equal osmotic
concentration, the solutions are isotonic. If they are unequal however, the solution possessing a osmotic
concentration is hypertonic and the solution with a lower osmotic concentration is said to be hypotonic
(Mason et al. 2015)

Red blood cells will swell when placed in hypotonic solution and shrivel when placed in a
hypertonic solution. This effect can be attributed to osmosis and the selectively permeable nature of the
cell membrane which allows for the movement of certain solutes but inhibits the movement of others thus
leading to implications not just the on cell’s volume but on its integrity as well (Goodhead and Macmillan
2017). When in a hypotonic solution, the osmotic concentration of the fluid within the red blood cell is
higher than that of the solution causing an influx of water towards the red blood cell. The same concept
applies to when the red blood cell is in a hypertonic solution. The osmotic concentration of the fluid
within the red blood cell is lesser than that of the solution. This leads to an efflux of water from the red
blood cell. (Roiger and Bullock 2019).

The medical field will benefit from the findings of this study considering that the presence of red
blood cells is crucial in sustaining the lives of most organisms. Medical practitioners will gain insights on
how the relative concentration of solutions to red blood cells affects the cells volume and integrity. This

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information is integral when it comes to certain medical procedures such as administering intravenous
fluids to patients and storing blood. Being informed on how red blood cells react to different
concentrations will also help in the development of future treatments. For researchers, this study may be
used as a guide to further their own studies regarding osmosis.

This study sought to observe the effects of a hypertonic, isotonic, and hypotonic solution on a red blood
cell. This was done in order to ascertain if a hypertonic solution leads to the red blood cell shriveling and
if a hypotonic solution leads to the red blood cell swelling. The effects of a isotonic solution on the red
blood cell was used as a baseline. The effects that these solutions had on the red blood cells were used to
either prove the study’s hypothesis.

METHODOLOGY

Three slides were prepared. Slide 1 has a These solutions were prepared because they provided an
0.9% NaCl solution, slide 2 has a 5% NaCl, isotonic, hypertonic, and hypotonic environment for the
and slide 3 has distilled water. red blood cells in which their reactions could be
observed
The researchers washed their hands and This prevented the slides from being contaminated with
prepared the sterile lancet other substances. The sterile lancet on the other hand
was used to puncture the subject’s finger in order to get a
small blood sample
The subject was asked to massage either his The process of massaging the subject’s finger from the
middle finger or ring finnger of his non- base to the tip was done in order to improve blood
dominant hand from the base to the tip circulation which made it easier to obtain more blood.
The subject’s fingertip was cleaned with an This was done to remove and bacteria that could enter
alcohol swab and then dried. the subject’s body when the needle from the lancet
penetrates the skin.
The opening of the lancet was positioned The fingertip was chosen because it has a higher
against the subject’s fingertip and the release concentration of capillaries this area was also easier to
level was pressed. penetrate with the lancet which made it easier to obtain
blood.
Blood was gently squeezed from the By applying pressure on the subject’s finger, more blood
subject’s finger. The blood was then was extracted from the capillaries.
transfered onto the different slides using a
toothpick.
The blood and the solution were mixed using These procedures finalized the preparation of the wet
a toothpick and the cover slip was applied. mounts to be observed in the study. Blood was mixed
with solution in order to disperse the red blood cells
which made them easier to find under a microscope.
The appearance of the red blood cells in The slides were viewed under the high power objective
each of the solutions was observed under a to clearly see the difference in the shapes and sizes of
microsope using the high power objective. the red blood cells. This was done to prove the
researcher’s hypothesis

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 RESULTS & DISCUSSION

The red blood cells mixed with the 0.9% NaCl solution did not undergo any observable structural
changes. They appeared to be oval when viewed under a microscope (Fig. 1).

Fig 1. Red blood cells in slide 1 containing a 0.9%

NaCl Solution.

A 0.9% NaCl solution is isotonic relative to red

blood cells because the osmotic concentrations of the
of the intracellular fluid is the same as that of the
0.9% NaCl solution. When red blood cells are in this
environment, the intracellular and extracellular fluids
are in a state of osmotic equilibrium and there is no
net movement in water (Reece et al.2014). Since there
was no net influx or efflux of water, the red blood
cells’ structure remained unaffected and did not
experience swelling nor shriveling. This is in line
with the statement of Purves et al. (1998) that the
integrity of a red blood cell is dependent on the
environment in which it is suspended in, it must be
isotonic with the cells so that it does not burst or
shrivel. This supports the hypothesis that red blood
cells will swell when placed in hypotonic solution and
shrivel when placed in a hypertonic solution since the solution in slide 1 is neither hypertonic nor
hypotonic the cell experienced no change in volume. The effects of a hypertonic and hypotonic solution
on red blood cells were further observed on slides 2 and 3 which contained a 5% NaCl solution and
distilled water respectively.

The red blood cells mixed with the 5% NaCl solution appeared to have become shriveled (Fig. 2).

Fig. 2. Red blood cells in slide 2 containing a 5%

NaCl Solution.

The 5% NaCl solution has a higher osmotic

concentration as compared to the intracellular fluid of
the red blood cells. This makes the solution
hypertonic relative to red blood cells. When subjected
to a hypertonic environment, the red blood cells
shriveled. This is because of osmosis, the process
wherein water moves from areas with a lower
concentration of solutes to an area with a higher
concentration of solutes. This led to a net efflux of
water from the red blood cells causing it to shrivel
(Lodish et al. 2005) The results correspond with the
expected outcome thus supporting the hypothesis that
red blood cells will swell when placed in hypotonic
solution and shrivel when placed in a hypertonic
solution. In order to observe red blood cells in a
hypotonic solution, the red blood cells were then
viewed under slide 3 which contains distilled water.

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 The red blood cells mixed with distilled water appeared to be lysed (Fig. 3).

Fig. 3. Red blood cells in slide 3 containing distilled water.

Distilled water possesses a lower osmotic

concentration as compared to the intracellular fluid of
the red blood cells. This made the solution hypotonic
relative to red blood cells. When subjected to a
hypotonic environment, the cells swelled and
eventually became lysed. Similar to the previous
slides, this result was attributed to osmosis. As water
passed through the cell membrane into the
intracellular fluid since it had a higher osmotic
concentration, the cell swelled. This led to the
cytoplasm exerting pressure on the cell membrane.
The cell membrane was not able to withstand the
internal pressure which caused it to burst (Mason et
al. 2015). The results correspond with the expected
outcome thus supporting the hypothesis that red blood
cells will swell when placed in hypotonic solution and
shrivel when placed in a hypertonic solution.

CONCLUSION & RECOMMENDATIONS

The results of the experiment wherein red blood cells were placed in three different slides
containing a 0.9% NaCl solution, 5% NaCl solution, and distilled water showed that red blood cells
became lysed when placed in hypotonic solution, shriveled when placed in a hypertonic solution, and had
no structural changes when placed in an isotonic solution. This affirmed the hypothesis of the study. The
outcome of the experiment was attributed to osmosis, the process in which water moves from areas with a
lower concentration of solutes to an area with a higher concentration of solutes. This led to a net influx of
water towards the red blood cell in a hypotonic environment causing the red blood cells to become lysed
and a net efflux of water from the red blood cell in a hypertonic environment causing the red blood cells
to shrivel. In an isotonic environment, there was no net movement in water thus leading to no change in
volume or structure.

The researcher recommends studies to be done on the effects of osmosis on different cells in
order to observe how cells from different tissues would react when subjected to environments with
different concentration gradients. Plant cells for example possess a cell wall which could lead to the cells
reacting differently.

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Roiger D, Bullock J.2019. Anatomy, physiology, & disease foundations for the health professions 2nd
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K. Goodhead, Lauren & M. MacMillan, Frances. 2017. Measuring osmosis and hemolysis of red blood
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Mason K, Johnson G, Losos J, Singer S. 2015. Understanding Biology. New York (NY): McGraw-Hill
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Purves K, Orians G, Heller H, Sadava D. 1998. Life the science of biology. New York (NY): W. H.
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