Method
List of Equipment
7 clean evaporating dishes
A supply of 1M lead nitrate
A supply of 1M glucose solution
A 10 ml syringe
A stopclock
Microscope
Red onion
Scalpel
Ceramic Tile
A beaker
Slides and Coverslips
Forceps
1. Preparing Lead Nitrate Concentrations
Six concentrations of lead nitrate should be prepared, 0.2M, 0.3M 0.4M 0.5M, 0.6M and 0.7M. To do this, a supply of 1M lead nitrate is needed. Then, using a syringe to measure amounts, the concentrations above should be prepared according to the table below.
20ml of each concentration of lead nitrate should be should be put into a separate, clean, evaporating dish. Clean them if they have already been used once. Only use enough to cover the evaporating dish, so that a piece of membrane can be completely immersed.
3. Preparation of Onion Membrane
Cut the red onion in half on the ceramic tile using the scalpel. Then peel away the dry outer layers of skin, then the dark red layer using forceps if necessary, make sure it is dark red but not too dry. This slightly moist red layer should be cut up into 6 small pieces (around 5mm2 should be sufficient) with the scalpel.
4. Immersion in Distilled Water (5 minutes), and Lead Nitrate (8 minutes)
Immerse each membrane in water for 5 minutes to ensure each cell is turgid.
Put a piece of membrane in the evaporating dishes of lead nitrate for 8 minutes (use the stopclock). The 0M lead nitrate solution will act as the control for the experiment.
5. Immersion in Glucose Solution (2 minutes)
Convey the membrane into the glucose solution for 2 minutes (stopclock).
6. Check for % Plasmolysis by Microscopy
Put a membrane under the microscope. Align the cells with the graticule, and count how many cells on the graticule line have plasmolysed, and how many have not. Convert to a percentage as below, and record in the table.
No. Plasmolysed * 100 = Percent Plasmolysed
Total No. Of cells
Repeat for each of the other membranes, making sure not to mix them up.
7. Immersion In Distilled Water (2 minutes)
Take the membrane and immerse it in an evaporating dish of distilled water for 2 minutes (stopclock).
8. Check for % Plasmolysis by Microscopy
Repeat step 6 for the onion membranes
9. Repeat steps 3 to 8 with fresh solutions until 2 extra sets of results have been completed
Variables/Constants
Variables:
Constants:
- Sugar concentration (the 0.6M is made in a large batch to minimise variation
- Temperature and Pressure (variation should be minimal in a lab during a particular day)
- Time in lead nitrate or glucose solution (careful watching of the stopclock should ensure time remains constant)
- Thickness of onion membrane (the pieces of membrane should have come off the onion at the same time in a large piece before being cut up. Care should ensure that this membrane is only 1 cell thick)
Preliminary Work
√ = Evidence of Significant Plasmolysis
X = No Evidence of Significant Plasmolysis
From these preliminary results, 1M glucose was chosen as the concentration for the glucose solution, it was concentrated enough to force plasmolysis of the very weakly poisoned cells in the 2 minutes, but was not strong enough to force some of the more significantly poisoned cells. 0.6M and 0.2M were too weak, and even cells where it is doubtable that any poisoning had taken place did not undergo plasmolysis.
It also showed that the lowest concentration for poisoning to occur was around 0.4M, so the range in this follow-up experiment has been reduced for increased accuracy.
Table of Results (to be filled in)
The percentage of plasmolysed cells in each case should be recorded in the table above. Graphs can easily be plotted (see sample below), to analyse data.
Risk Assessment
Lead Nitrate is toxic. Care should be taken so that it does not come into contact with skin. Avoid swallowing any.
Scalpels are extremely sharp. Take care when cutting the onion.
Reliability
Reliable results are extremely important. Make sure that a new and uncontaminated glucose solution is used for after each bathing of the membranes, as lead could get into it and will build up as the experiment progresses.
Although it is not critically important, make sure that each piece of membrane is around the same size. Any difference should be negated by the fact it is one cell thick, however.
Ensure that all times are well kept to. Do not run over, especially in the glucose solution and distilled water, otherwise the percentage plasmolysed cells would be altered.
References
1. Toxicology - David Pascoe
2. Biology 1 - Cambridge Advanced Sciences
3. Microsoft Encarta - Microsoft
4. HealthWorld Online -
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