Required Practical 4

• The investigation into the effect of a named variable on the permeability of cell surface membranes
• Cutting plant tissue

  1. Cut equal sized cubes of plant tissue of same type using a scalpel

  2. Rinse to remove pigment released during cutting or blot on paper towel
• Preparing test tubes

  Add same number of cubes to 5 different test tubes containing same volume of water such as 5cm^3
• Incubating test tubes

  1. Place each test tube in a water bath at a different temperature e.g, going up in 10 degrees

  2. Leave for same amount of time e.g 20 mins
• Measuring colour intensity

  Remove the beetroot and measure the intensity of colour of surrounding solutions
• Semi-quantitative measurement

  • Use a known conc. of extract and distilled water to prepare a dilution series (colour standards)
  • Compare results with colour standards to estimate conc.
• Quantitative measurement

  • Measure absorbance of light of known concentrations using a colorimeter
  • Draw a calibration curve -> plot a graph of absorbance (y) against conc. of extract (x) and draw a line of best fit
  • Absorbance value for sample read off calibration curve to find associated extract conc.
• The issues with comparing to a colour standard is that matching to colour standards is subjective and the colour obtained may not match any of the colour standards
• The beetroot should be washed before placing it in water in order to wash off any pigment on the surface, to show that release is only due to a named variable
• Regularly shaking each test tube with the cubes is essential to ensure all surface of cubes remain in contact with liquid and also to maintain a concentration gradient for diffusion
• Controlling the volume of water is essential because too much water would dilute the pigment so solution will appear lighter and more light passes through in colorimeter than expected so results are comparable
• You could ensure that the beetroot cylinders were kept at the same temperature throughout the experiment by taking readings in intervals throughout experiment of temperature in tube using a digitial thermometer or a temperature sensor. If temperature has fluctuated then use a corrective measure.
• High absorbance suggests that the cell membrane is more permeable and damaged. This is because more pigment leaks out making surrounding solutions more concentrated.
• Temperature affects the permeability of the cell surface membrane as when it increases, permeability increases this is because the phospholipids gain kinetic energy and fluidity increases. Then transport proteins denature at high temperatures as H bonds break, changing the tertiary structure. At low temperatures, the permeability increases as ice crystals can form which pierce the cell membrane and increase permeability.
• pH can affect permeability of cell surface membranes as at high or low pH incraeses permeability. This is because transport proteins denature as H or ionic bonds break, changing the terary structure.
• Lipid soluble solvents such as alcohol can affect permeability of cell surface membranes as concentration increases, permeability increases. Ethanol may also dissolve phospholipid blayer forming a gap.