• The?permeability?of?cell membranes?is affected by different?factors?or conditions, such as:
  • Temperature
  • Solvent concentration

   

• You can investigate how these different factors affect membrane structure and permeability using?beetroot
• • Beetroot cells contain a?dark purple-red pigment
  • The?higher?the permeability of the beetroot cell membrane, the?more?of this pigment leaks?out of the cell

   

Investigating the effect of temperature on membrane permeability

• In this practical, you need to show knowledge and understanding of how to use?appropriate apparatus?to?record a range of quantitative measurements, including mass, time, volume, temperature, and length. For example:
  • Mass?should be recorded using a?digital balance
  • Time?should be recorded using a?digital stopwatch
  • Volume?should be recorded using a?measuring cylinder
  • Temperature?should be recorded using a?digital thermometer?(although water baths have one built-in)
  • Length?should be recorded using a?ruler

   

• In addition, you need to show knowledge and understanding of how to use another piece of apparatus, known as a?colorimeter, that is also appropriate for use in this practical
  • A colorimeter is a machine that passes light through a?coloured liquid sample?and measures how much of that light is absorbed (and therefore gives an indication of how much of the colour is present in the solution)
  • A colour filter is used in the light path to ensure that the correct wavelength of light is used to measure the optical density of the specific pigment in the solution (e.g. the beetroot pigment called betalain)
  • The colorimeter must be?zeroed?before each colorimeter tube (called a?cuvette) is inserted. This can be done using?distilled water?in a cuvette

   

Apparatus

• Scalpel
• Cork borer (optional)
• Cutting board
• Ruler
• Digital balance
• Test tubes
• Measuring cylinder
• Water baths
• Digital stopwatch
• Colorimeter

Method

• Using a scalpel, cut five?equal-sized?cubes of beetroot
  • The pieces must have the same dimensions so that they all have?equal surface areas and volumes, as these factors could affect the rate at which the pigment leaks out
  • A cork borer can also be used, as long as the cores are cut to the?same length
  • You should also use a digital balance to check that all pieces have the?same mass

   

• Rinse?the beetroot pieces
  • To?remove?any pigment released during cutting

   

• Add the beetroot pieces to five different test tubes, each containing the?same volume of water?(e.g. 5cm3)
• Put each test tube in a water bath at a different temperature (e.g. 10℃, 20℃, 30℃, 40℃, 50℃) for the?same length of time
  • The time should be?long enough to allow the pigment to diffuse into the water?(e.g. around 30 minutes)

   

• Remove the beetroot pieces, leaving just the coloured liquid in the five test tubes
• Use a?colorimeter?to measure how much?light?is?absorbed?as it passes through each of the five samples of coloured liquid
  • The?higher?the?absorbance, the?more?pigment?must have been released, due to a?greater?membrane?permeability

   

Results

• The general pattern you would expect to see is that?as temperature increases, membrane permeability also increases
  • As temperature increases, the phospholipids within the cell membrane?move?more because they have more?energy
  • Increased movement means the phospholipids are?not?as?tightly packed?together, increasing the permeability of the membrane
  • At high temperatures, the phospholipid bilayer may even start to melt and?breakdown, further increasing the permeability of the membrane
  • In addition, the volume of water inside the cells?expands, putting pressure on the membrane, causing channel and carrier proteins to?deform?so they can no longer control what enters and leaves the cell. These factors also increase the permeability of the membrane
  • Temperature also affects the?conformation?(3D shape)?of proteins?as at high temperatures the?intermolecular forces?between amino acids are?broken?which affects the protein’s?specificity and function

   

• If experimenting with?temperatures below 0oC, membrane permeability may also be increased (once the cells have thawed again)
  • Increased permeability can be caused by channel or carrier proteins?deforming?at these low temperatures
  • Ice crystals?that form can also?pierce?the cell membrane, making it highly permeable

   

Example results showing the effect of temperature on membrane permeability

Limitations

• Cuvettes?are the small cuboid containers that?hold?the liquid to be measured in a colorimeter
• Cuvettes may differ in?thickness?(very slightly). A thicker (or?scratched) cuvette will absorb slightly?more light?than a thinner unscratched cuvette
  • Solution: use the?same cuvette?for every reading, or?repeat?the investigation many times and find a?mean

   

• The beetroot pieces may not be identical in?size?and shape, meaning some test tubes could contain slightly more beetroot tissue than others
  • Solution: cut the discs as?accurately?as possible using a?scalpel and ruler, and?repeat?each investigation several times to find a mean

   

• Some?parts?of beetroot tissue have?more pigment?in their cells than others
  • Solution: conduct several?repeats, using?different parts?of the beetroot and find a?mean