Cell Membranes and Temperature

Located within the vacuole of beetroot cells is a red pigment call Betalains. Typically these pigments are contained within the cell vacuole by the tonoplast (vacuole membrane), however When beetroot is heated these red pigments escape the vacuole. This experiment aims to explore the effect of temperature on the permeability of the cell membranes (i.e. Tonoplast). ‘The cell surface membrane is the plasma membrane that surrounds cells and forms the boundary between the cell cytoplasm and the environment…..It controls the movement of substances in and out of the cell.’ (Glen Toole and Susan Toole, 2008, p.552). The tonoplast is identical to the cell surface membrane except it surrounds only the vacuole, compared to the cell surface membrane which surrounds the entire cell. A plasma membrane is made up of a phospholipid bilayer. Each layer which consists of a hydrophobic fatty acid tail and a hydrophilic phosphate head. Each layer

Apparatus

• Beetroot
• Size 4 Cork Borer
• White Tile
• Scalpel/Knife
• Ruler
• Water Baths (0°C, 10°C, 20°C, 30°C, 40°C, 50°C, 60°C, 70°C)
• Plastic Beaker (~250cm3)
• 2 Boiling Tube Racks

• 8 Boiling Tubes
• Crushed Ice
• 8 Thermometers (one per water bath)
• Stopwatch
• Distilled Water
• Pipette
• Cuvette
• Colorimeter

Method

(N.B. – All personal protective equipment must be worn and care must be taken when using the cork borer or Scalpel/knife. All laboratory rules must be followed)

Eight cylinders where cut from the Beetroot, using a cork borer, and then cut to 1cm using a knife or scalpel. Eight water baths were set up at temperatures of 0°C, 10°C, 20°C, 30°C, 40°C, 50°C, 60°C and 70°C. The 50 60 and 70 water baths were maintained electrically whereas the rest were manual. Eight boiling tubes were taken and labelled according to its corresponding water bath and filled with 5cm3 and left in its matching water bath for 5 minutes to allow acclimatisation of the distilled water. After five minutes one of the eight 1cm long cylinders of beetroot was placed in each of the boiling tubes and left for the 30 minutes in the water baths. After 30 minutes the beetroot pieces were removed from the boiling tubes and the tubes were shaken to disperse the dye. Then using a pipette approx. 2cm3 of the solution from each of the test tubes was taken and placed in a cuvette, with care taken to keep solutions of different temperatures separate. The cuvette was then placed in a colorimeter, ensuring the smooth sides were facing the light source, and the absorbance of the solution was measured. Before each measurement the colorimeter was calibrated to an absorbency of zero using a cuvette of distilled water. This was done for the cuvettes of each of the temperatures.

Results

|Temp. |Absorbance |
|(Initial)/°C | |
|0 |0.12 |
|10 |0.15 |
|20 |0.23 |
|30 |0.21 |
|40 |0.26 |
|50 |0.39 |
|60 |1.70 |
|70 |1.96 |

The general trend followed by the results is, that as the temperature increase, the permeability of the cell membranes also increases. This is indicated by the fact that in the results when temperature increases the absorbance of the solution in the colorimeter also increases, suggesting a deeper colouration of the solution and therefore greater concentration of the pigment Betalains within the solution. This indicates that at a higher temperature, a greater amount of Betalains was allowed to seep out of the cell vacuole and into the surrounding environment.

Theory

As temperature increased, the permeability of the cell membranes also increased as shown by the general trend of the results. This is due to the fact that as the temperature was raised, the kinetic energy of the phospholipids within the cell membranes also increases, causing greater random movement of the phospholipid bilayer, this results in large holes within the bilayer causing the partially permeable membrane to become more permeable. The Betalains then diffused through the tonoplast and out of the vacuole, which is a region of high concentration, and into the distilled water, which is a region of low concentration, down a concentration gradient. The higher the temperature that the beetroot cylinder is exposed to, the greater the damage to the cell membranes and the greater the permeability of the cell.

Conclusion

The results we gained were not completely reliable since there were
many factors that weren’t taken into account and many errors which may have occurred. Possible Errors
• Cylinders of Beetroot may not have been cut accurately, as they were cut using a ruler which is only accurate to 1 mm. resulting in a greater or smaller surface area, which would have had an effect on the rate of diffusion of the Betalains once the cell membranes had become permeable. • There were some technical difficulties when using the colorimeter. For example the colorimeter may not have been calibrated correctly, the colorimeter may not have been on the correct setting, and the cuvettes may have been placed in the colorimeter in the wrong position, i.e. the smooth side not facing the light source. Also this test suffered from some unknown internal problem of the colorimeter and since this was not solved some liability of the anomalous results lies in this region amongst others.