Flashcards in Experiments to know (Chapter 4 and 3) Deck (22):
What is visking (dialysis) tubing?
A partially permeable non-living membrane made from cellulose
- it possesses molecular-sized pores which are small enough to prevent the passage of large molecules e.g. starch/sucrose but will allow the passage of small molecules e.g. glucose by diffusion
How could you demonstrate diffusion using visking tubing?
1) Fill a length of visking tubing (roughly 15cm) with a mixture of starch and glucose solutions
2) If the tubing is suspended in a boiling tube of water for a period of time, the presence of starch and glucose outside the tubing can be tested for at intervals to monitor whether diffusion out of the tubing has occurred
3) Results should indicate that only glucose diffuses out of the tubing
How could you demonstrate diffusion using visking tubing more quantitively? (2 ways)
1) Try to estimate the glucose concentration at each time interval by setting up separate tubes, one for each planned time interval, and using a semi-quantitive Benedict's test each time - a colorimeter could also be used
2) OR A set of colour standards could be prepared and a graph could be drawn showing how the rate of diffusion changes with the concentration gradient between the inside and outside of the tubing
What could you demonstrate diffusion with?
1) Starch and glucose
2) Sucrose and sucrase
3) Starch and amylase
How could you investigate the effect of size (SA:vol) on diffusion?
By timing the diffusion of ions through blocks of agar of different sizes:
1) Solid agar is prepared in suitable containers i.e. ice cube trays and made up with very dilute NaOH and universal indicator (so solution is purple)
2) Cubes of the required dimensions e.g. 2x2, 1x1, 0.5x0.5 cm can be cut from the agar, placed in a container and covered with a diffusion solution e.g. dilute HCl with a higher molarity
3) Either the time taken for the acid to completely change the colour of the agar blocks, or the distance travelled into the block by the acid in a given time e.g. 5 mins can be measured - the times can be converted into rates
4) The rate of diffusion (colour change) can be plotted against SA:vol ratio
Why is it better to use acid as a diffusion solution rather than universal indicator?
Because the colour will be affected by the pH of the water in the agar
How could you investigate the effect of environmental factors e.g. chemicals and temperature?
1) Pieces of beetroot can be placed into water at different temps or into diff alcohol concentrations
2) Any damage to the cell membranes results in the red pigment, which is normally contained within the large central vacuole, leaking out of the cells by diffusion
3) Changes in the colour of the surrounding solution can be monitored qualitatively or quantitively
- a colorimeter or a set of colour standards could be used
- could put the tubes in order and make up a colour scale e.g. 0-10 (water=0, darkest=10)
Explain the observations when you put the beetroot in ethanol/high temps?
- The red dye diffuses from a region of high concentration in the vacuoles to a region of low concentration in the solution outside the beetroot
- Diffusion is normally prevented by the partially permeable nature of the membranes (tonoplast), but in this case, the membranes have been damaged by the alcohol/temp
What happens to a cube as you cut it into smaller pieces?
The SA increases, but the volume stays the same
Therefore, the SA:vol ratio increases
Why do you cover water/ethanol/any liquid with e.g. clingfilm/parafilm/bungs when you leave a solution for a period of time?
- To prevent evaporation of the liquid which would affect the concentration/water potential of the solution and therefore the accuracy of the results would decrease
Why do you wash the visking tubing before putting it into distilled water?
To wash off any spillage of solution on the outside of the tubing
Why do you wash the beetroot pieces in distilled water before carrying out the experiment?
To remove excess pigment on the surface of the beetroot cylinders
How does ethanol affect membrane permeability?
- Ethanol disrupts the membrane by interacting with the phospholipid bilayer as the OH group can interact with the phosphate group
- Ethanol increases the fluidity of the membrane
- It may denature the proteins embedded in the membrane
What would you use to observe osmosis (plasmolysis) in plant cells?
Epidermal strips because coloured sap makes observation easier e.g. red cabbage, rhubarb, red onion storage leaves
How could you investigate osmosis in plant cells?
1) Place the strips of epidermis in a range of molarities of sucrose solution (up to 1moldm3) or NaCl solutions of up to 3%
2) Place small pieces of the strips on glass slides, mounted in the relevant solution and observed with a microscope
3) Plasmolysis may take several minutes, if it occurs
Describe the principle of the experiment to determine the water potential of a plant tissue
To find a solution of known water potential which will cause neither a gain nor a loss in water of the plant tissue being examined
How could you determine the water potential of a plant tissue?
1) Samples of the tissue e.g. potato are allowed to come into equilibrium with a range of solutions e.g. sucrose of different water potentials and changes in mass/volume of the tissue are recorded
2) Plotting a graph of the results allows the solution that causes no change in mass/volume to be determined
3) This solution will have the same water potential as the plant tissue
Why is it difficult to measure the reaction rate of the breakdown of starch by amylase?
Because the substrate and product remain as colourless substances in the reaction mixture
How could you measure the reaction rate of the breakdown of starch by amylase?
Measure the rate at which starch disappears from the reaction mixture
1) Take samples from the mixture at known times, and adding each sample to the some iodine in potassium iodide solution
2) As starch forms a blue-back colour with this solution, wising a colorimeter you can measure the intensity of the blue-black colour obtained, and use this as a measure of the amount of starch still remaining
3) If you do this over a period of time, you can plot a curve of 'amount of starch remaining' against 'time'
4) You can then calculate the initial reaction rate in the same way as for the catalase-H2O2 reaction
What is another way to measure the reaction rate of the breakdown of starch by amylase but why is it not ideal?
Mix starch, potassium iodide solution and amylase in a tube, and take regular readings of the colour of the mixture in this one tube in a colorimeter
- This is not ideal because the iodine interferes with the rate of the reaction and slows it down
How could you immobilise enzymes e.g. lactase?
1) The enzyme is mixed with a solution of sodium alginate
2) Little droplets of this mixture are then added to a solution of calcium chloride
3) The sodium alginate and calcium chloride instantly react to form jelly, which turns each droplet into a little bead which contains the enzyme
- The enzyme is held in the bead (immobilised)