Required practical 4 Flashcards
Beetroot
What is this practical investigating
. Effect of alcohol concentration, or temperature on the permeability of the cell surface membrane by looking at leakage of pigment from beetroot through plasma membranes
What is the purple pigment in beetroot called, and why is it used
Betalin found in vacuoles of beetroot cells
. It can’t move across undamaged plasma membranes
How do you make the colour standards of the beetroot
. Using beetroot extract first, add water to prepare a series of 6 boiling tubes containing 5cm3 of different concentrations of extract.
The concentrations should be from 0% extract and 100% water, to 100% beetroot extract and 0% water.
So do 0%, 20%, 40%, 60%, 80% and 100%
These are the colour standards, label them 0,2,4,6,8,10
What do you use the colouremeter for and how do you set it up
. Use it to get a quantitative number for the colour of our solution, so it is not all based on how it looks for the person
Set it up by using water to calibrate it to zero absorbance , then measure the absorbance of each of the colour standards you have prepared
How do you carry out the investigation
. Set water bath to 30 degrees
. Get a second set of boiling tubes and add 2cm3 of 100% alcohol to a test tube and put a bung in the tube
. Label tube with the alcohol concentration, then repeat this with alcohol concentrations of 80%, 60%, 40%, 20%
. Put these in water bath until temperature reaches 30 degrees
. Blot 10 discs of beetroot with paper towel
. Use forceps to put two discs in each alcohol tube, and replace bungs soon after
. Leave tubes in water bath for 5 minutes, and shake gently once per minute then remove
. Immediately pour each solution into a clean boiling tube (labelled correctly) and remove beetroot, you do this immediatly to limit time variation of discs in contact with alcohol in water bath
. Compare each of the solutions with the colour standards made earlier by using colouremeter which which measures light absorbance of each one
Which ever ones have very similar absorbance levels to the standards made earlier; so you know the amount of beetroot pigment that has leaked out
So the darker the solution, the more damaged the cell membrane
Why do you blot beetroot discs with paper towel before putting in alcohol
To remove excess water, because it could change the concentration of the alcohol solution
. It also removes excess pigment on beetroot which may make the solution appear darker even if no beetroot has left the cell membrane
What is the relationship between alcohol concentration and the cell surface membrane
The more alcohol added, the more broken the membrane so it is more permeable and lets more beetroot pigment leak through, making it a darker solution.
Name 2 factors that affect permeability of cell membranes
. Temperature
. Concentration of solvents eg ethanol
How would you investigate the effect of temperature on cell membrane permeability
. With 10 identical beetroot discs, place 2 in each boiling tube, so there are 5 boiling tubes
. Each boiling tube has an equal volume of distilled water
. Place each one in a water bath of different temperatures ranging from
30 degrees to 80 degrees and leave for 20 minutes
. Then filter each solution and put in colouremeter to measure the colour difference quantitatively
Hazards of ethanol
. It is an irritant so may sting and irritate skin so wear gloves and goggles
. It is also flammible so keep away from any naked flames
. Keep sharp scalpel away from fingers
. Don’t burn yourself in hot liquid
Why does increasing temperature increase the amount of betalains from beetroot concentration in the solution
. At higher temperatures, the phospholipid bilayer becomes more fluid so more pigment can leave the membrane into the solution
. Also, the proteins in the membrane may denature at higher temperatures
. The tonoplast around the vacuole may weaken also and let more pigment into the cell
So as a result there will be more absorbance in the colourometer
