“Diffusion Through Living Membranes”

Introduction

a.) Background Information–

A membrane is a thin layer of cells that act as a boundary. It separates the interior of ALL cells from the outside environment. The plasma membrane allows nutrients to enter the cell and keeps out unwanted substances. This process is known as selective permeability. There are two types of transport of molecules in and out of cells: passive and active processes. Passive processes transport is a basic way that transport through the plasma membrane can occur. In passive processes, pressure differences drive the movement. Diffusion is the movement of molecules from an area of high concentration to an area of low concentration. Molecules generally diffuse passively though the plasma membrane. Simple diffusion is the unassisted transport across a plasma membrane of a very small particle. Just as in simple diffusion, facilitated diffusion moves down a concentrated gradient. However, it is used by large molecules that can pass through the plasma membrane independently. Diffusion of a solvent through a selectively permeable membrane from a dilute solution to a concentrated one is called osmosis. Simple diffusion, facilitated diffusion, and osmosis are the three types of passive transport across the plasma membrane. Molecules have kinetic energy and are in constant motion. Because of this they move around randomly at high speeds, causing them to be unevenly distributed. When this happens a concentration gradient is present, resulting in areas of high and low concentration. During osmosis, water moves along a concentration gradient. Because the concentration of water is related to that of solutes, if the solutes are able to diffuse across the membrane so can the water. If there are particles present in a solution that prevent the solutes from crossing the membrane, then only the water can move by osmosis. When this happens, changes in the volume occur on either side of the membrane, which is why there is tonicity. Tonicity is a measure of the ability of a solution to cause a change in a cell’s shape by promoting osmotic flows of water. There are three types of tonicity: hypertonic, hypotonic, and isotonic. A hypertonic solution is a solution that has a higher concentration of nonpenetrating solutes than the reference cell. The concentration of solutes is greater outside the cell than inside it, causing it to shrink. Crenation usually results from a hypertonic solution. Crenation happens when red blood cells undergo shrinkage and take on a scalloped surface. Hypotonic solutions have less nonpenetrating solutes than the reference cell. These solutions have greater concentration inside of the cell than outside, causing them to swell and burst. Hemolysis occurs in hypotonic solutions. Hemolysis is the destruction of red blood cells. An isotonic solution has a concentration of nonpenetrating solutes equal to that found in the reference cell. In active processes a cell uses the bond energy of ATP to move substances across the plasma membrane. ATP is required because the molecule has to go against the concentration gradient.

In this experiment, the tonicity of varying solutions, all of which containing red blood cells was determined. The independent variables are the two NaCl solutions and the distilled water. The responding are the red blood cells in slides 2-4. The controlled variable are the red blood cells in slide 1.

b.)  Purpose–

The purpose of the experiment is to determine tonicity. The lab report is to record the data and results of the experiment.

c.)   Hypothesis–

Slide #1 will be isotonic.

Slide #2 will be hypertonic.

Slide #3 will be hypertonic.

Slide #4 will be hypotonic.

Procedure

a.)   Equipment Used–

0.9% NaCl  Filter paper-if needed  4 clean slides

10% NaCl  Wooden applicator stick  Cover-slips

 dH2O   1 disposable mat    Microscope

Sheep’s blood Disposable gloves   Goggles 

Wax pencil

b.)  Collection of Data–

Using the disposable mat, place the four slides, cover-slips and sheep’s blood on top. Then, number the slides 1-4. Place one drop of blood on slide 1.

With the wooden applicator stick, dip one end into the blood and place a single drop on the next 3 slides.

Dispose of the wooden applicator stick in the biohazard waste container.

On slide 2, add 1 drop of .9% NaCl with the drop of blood. Add a cover-slip on top and use filter paper to absorb excess liquid.

Observe the cells under scan, low power, then high power. Draw observations.

Repeat steps, adding 10% Nacl to slide 3 and dH2O to slide 4, in replace of .9% NaCl.

Place ALL biohazard waste in the appropriately marked bins and dispose of gloves. The slides with the solution should be placed in the appropriate container.

Results

a.)   Table–

Slide #

Contents

Tonicity

Results

Appearance

1

Sheep’s Blood

Isotonic

Normal

Cell

Circle shaped

2

Blood &

.9% NaCl

Isotonic

Normal

Cell

Circle shaped

3

Blood &

10% NaCl

Hypertonic

Crenation

Shriveled Circle

4

Blood &

dH2O

Hypotonic

Hemolysis

Burst

b.)  Description of Data–

The sheep’s blood under a microscope was light red in color with smaller red dots. When set at a higher power the cells looked like small clear dots. On both setting the cells were almost perfect circular shapes. The solution was isotonic because it was a normal cell.

Adding the .9% NaCl caused the overall color to change from red to white on low power. At a closer look the cells appeared to be gray and farther spaced apart than in slide #1. The shape remained the same. The solution was isotonic because it was a normal cell.

Adding 10% NaCl to the sheep’s blood made the color look brown. The shape of the cells were shriveled looking circles. The solution was hypertonic because crenation occurred.

Sheep’s blood and dH2O together did not show any cell, because it blew up. The solution was hypotonic because hemolysis occurred.

Discussion–

a.)   Support of Hypothesis–

Slide #1 will be isotonic: supported

Slide #2 will be hypertonic: not supported Slide #2: isotonic

Slide #3 will be hypertonic: supported

Slide #4 will be hypotonic: supported

b.)  Explanation–

Slide one contained the controlled variable; the red blood cells. This was the control because it remained unchanged throughout the experiment. The purpose of the control variable is to have an example of what the red blood cell looks like by itself. The NaCl and dH2O were the independent variables because they were changed throughout the experiment. The independent variable represents the reason for the outcome of the experiment. The dependent (responding) variable in this experiment were the red blood cells in slides 2-4. These were the ones being tested and measured, therefore they were dependent on the independent variables to yield an outcome. Slide one was isotonic because it was a “normal cell”. It had the same osmotic pressure across a selectively permeable membrane. Osmosis did not occur in this solution because it had an equal solute/water concentration. Slide two was isotonic as well, because even after adding the .9% NaCl the concentration of nonpenetrating solutes was still equal to that of the reference cell (slide 1). Osmosis occurred, however the cells neither gained or lost any water. Slide three was hypertonic because it underwent crenation causing the shape of the cell to become shriveled. Osmosis occurred because there were more nonpenetrating solute particles than the interior of the cell. Slide four was hypotonic because hemolysis occurred and resulted in the cell bursting or blowing up. Osmosis occurred because there were fewer nonpenetrating solute particles than the interior of the cell.

c.)   Clinical Application-

Diffusion is used in the radiology field in diffusion-weighted imaging (DW-MRI). DWI-MRI is a form of MRI based on measuring the motion of water molecules within a voxel of tissue. DW-MRI is useful in differentiating epidermoid carcinoma and malignant lymphoma and distinguishing radiotherapy-induced tissue changes from persistent and recurrent cancer.

References–

Human Anatomy & Physiology Laboratory Manual: 13th Edition; Exercise 5: The Cell: Transport Mechanisms and Cell Permeability pages 51-58.

Human Anatomy & Physiology, 11 Edition