AQA A-LEVEL BIOLOGY: Required Practicals

REQUIRED PRACTICAL #1

What is the method for testing the effect of temperature on enzymes?

(1) Make two control samples:

● Take two flat bottomed tubes.

● Add 5cm3 of milk suspension to each tube.

● Add 5cm3 of distilled water to one tube- this control will indicate the absence of enzyme activity.

● Add 5cm3 of hydrochloric acid to the other- this control indicates the colour of a completely hydrolysed sample.

(2) Take three test tubes and measure 5cm3 milk into each. Place in water bath at 10°C for 5 minutes to equilibrate.

(3) Add 5cm3 trypsin to each test tube simultaneously and start the timer immediately.

(4) Record how long it takes for the milk samples to completely hydrolyse and become colourless.

(5) Repeat steps 2-3 at temperatures of 20°C, 30°C, 40°C and 50°C.

(6) Find the mean time for the milk to be hydrolysed at each temperature and use this to work out the rate of reaction.

Rate = (mean time)^-1

REQUIRED PRACTICAL #1

What are the risks in the experiment about testing the effect of temperature on enzyme activity?

(-) Broken Glass [may cut individuals so move glass slowly and keep it away from edge of desk. In emergencies, apply pressure to cuts and seek medical attention].

(-) Hydrochloric Acid [may cause harm/irritation to eyes or cuts. Hence, wear eye protection and gloves and avoid skin contact. Wash it off skin immediately and flood eye/cuts with cold water].

(-) Hot Liquids [may scald. Use tongs to remove boiling tubes. Run any burns under cold water and seek medical attention].

(-) Enzymes [may trigger allergies. Avoid contact with skin and eyes. Seek medical attention if necessary].

REQUIRED PRACTICAL #1

How would you represent the graph of the effect of temperature on enzyme activity and what can you conclude?

Graph: rate of reaction against temperature

Conclusion:

Milk contains a protein called casein which, when broken down, causes the milk to turn colourless. Trypsin is a protease enzyme which hydrolyses the casein protein.

As the temperature increases from 10°C, kinetic energy increases so more enzyme-substrate complexes form. This means that the rate of reaction increases up to the optimum temperature

At temperatures beyond the optimum, bonds in the enzyme tertiary structure break, which changes the shape of the active site. This means that the substrate and enzyme are no longer complementary

REQUIRED PRACTICAL #2

What is the method for calculating the mitotic index of a sample?

(1) Heat 1 mol dm-3 HCl at 60°C in a water bath.

(2) Cut a small sample of the root tip using a scalpel.

(3) Transfer root tip to HCl and incubate for 5 minutes.

(4) Remove from HCl and wash sample in cold distilled water and remove the very tip using a scalpel.

(5) Place tip on a microscope slide and add a few drops of stain (e.g. toluidine blue O). This makes the chromosomes visible and will therefore show which cells are undergoing mitosis.

(6) Lower the cover slip down carefully onto the slide. Make sure there are no air bubbles in the slide which may distort the image, and that the coverslip doesn't slide sideways which could damage the chromosomes.

(7) Place under a microscope and set the objective lens on the lowest magnification.

(8) Use the coarse adjustment knob to move the lens down to just above the slide.

(9) Use the fine adjustment knob to carefully re-adjust the focus until the image is clear (you can use a higher magnification if needed).

(10) To calculate mitotic index, cells undergoing mitosis must be counted (cells with chromosomes visible), as well as the total number of cells.

Formula for mitotic index:

number of cells with visible chromosomes / total number of cells in sample.

REQUIRED PRACTICAL #2

What are the risks associated with calculating a mitotic index?

(-) Hydrochloric Acid [may cause harm/irritation to eyes or cuts. Hence, wear eye protection and gloves and avoid skin contact. Wash it off skin immediately and flood eye/cuts with cold water].

(-) Touldine blue O Stain [may cause harm/irritation to eyes or cuts. Hence, wear eye protection and gloves and avoid skin contact. Wash it off skin immediately and flood eye/cuts with cold water].

(-) Scalpel [may cause cuts as it is sharp. Use forceps to hold a sample whilst cutting, NOT fingers. Elevate cuts and apply pressure and seek medical attention].

(-) Broken Glass [may cut individuals so move glass slowly and keep it away from edge of desk. In emergencies, apply pressure to cuts and seek medical attention].

REQUIRED PRACTICAL #3

What is a callibration curve?

Calibration curves are graphs used to determine an unknown concentration of a sample by comparing the unknown to a set of standard samples with known concentrations - they are also known as standard curves. A dilution series can be used to create a set of samples with known concentrations.

REQUIRED PRACTICAL #3

How can you use a callibration curve to find the water potential of a plant tissue?

(1) Make a series of dilutions of 1M sucrose solution. These should be at 0.0, 0.2, 0.4, 0.6, 0.8 and 1.0M sucrose. Dilute using distilled water.

(2) Measure 5cm3 of each dilution into separate test tubes.

(3) Use a cork borer to cut out six potato chips and cut down the sections into identically sized chips. Dry each chip using a paper towel to remove excess water but do not squeeze.

(4) Weigh each before the start of the experiment.

(5) Place a potato chip in each test tube (one per sucrose concentration) and leave for 20 minutes.

(6) Remove each potato chip, dry gently using paper towel, and weigh them in turn.

(7) Calculate the percentage change in mass for each sucrose solution.

REQUIRED PRACTICAL #3

What are the risks associated with producing a dilation series to test the water potential of a plant tissue?

(-) Scalpel [may cause cuts as it is sharp. Use forceps to hold a sample whilst cutting, NOT fingers. Elevate cuts and apply pressure and seek medical attention].

(-) Broken Glass [may cut individuals so move glass slowly and keep it away from edge of desk. In emergencies, apply pressure to cuts and seek medical attention].

REQUIRED PRACTICAL #3

What is the graph that you should draw to find the water potential plant tissue + what are the conclucions made from this?

Plot a graph of change in mass against concentration of sucrose solution.

The point at which the line of best fit crosses the x axis (zero change in mass) indicates the point at which the solution is isotonic. This is when the water potential of sucrose solution is the same as the water potential of the potato tissue, so there is no net movement of water in or out of the potato

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Potato chips in lower concentrations of glucose solution will increase in mass, whilst those in the higher concentrations of glucose solution will decrease in mass.

In the dilute glucose solutions, the solution has a higher water potential than the potato, so water passively moves via osmosis to the area of lower water potential (the potato). This causes the potato to increase in mass.

In concentrated glucose solutions, water will move out of the potato, thus the potato will decrease in mass.

REQUIRED PRACTICAL #4

What is the method for testing how variables affect the permeability of the cell-surface membrane?

The permeability of a membrane can be measured by using beetroot cells, which contain a purple pigment called betalain. When the cell-surface membrane has a higher permeability, more pigment leaks out of cells. The permeability can therefore be measured by the amount of pigment leaked from beetroot cells into an aqueous solution using a colorimeter.

1) Cut beetroot into 6-10 identical cubes using a scalpel.

2) Wipe/rinse to clean off any pigment released as a result.

3) If investigating temperature: place each of the cubes of beetroot in an equal volume of distilled water (5-15ml).

4) Place each test tube in a water bath at a range of temperatures (30-80°C).

5) If investigating concentration of solvents: create a dilution series of ethanol using distilled water. Ethanol concentrations should range from 0-100% ethanol.

6) Leave the samples for 20 minutes - the pigment will leak out of the beetroot.

7) Set the colorimeter to a blue filter and zero using a cuvette with distilled water

8) Filter each sample into a cuvette using filter paper.

9) Measure the absorbance for each solution. A higher absorbance indicates higher pigment concentration, and hence a more permeable membrane.

REQUIRED PRACTICAL #4

What are the risks associated with testing how variables affect the permeability of the cell-surface membrane?

(-) Scalpel [may cause cuts as it is sharp. Use forceps to hold a sample whilst cutting, NOT fingers. Elevate cuts and apply pressure and seek medical attention].

(-) Broken Glass [may cut individuals so move glass slowly and keep it away from edge of desk. In emergencies, apply pressure to cuts and seek medical attention].

(-) Ethanol [may cause harm/irritation to eyes or cuts. Hence, wear eye protection and gloves and avoid skin contact. Wash it off skin immediately and flood eye/cuts with cold water].

REQUIRED PRACTICAL #4

What is the graph and conclusions made from testing how variables affect the permeability of cell-surface membranes?

TEMPERATURE

As the temperature increases, the permeability of the cell-surface membrane also increases. This is because the proteins in the membrane denature as the heat damages the bonds in their tertiary structure. This creates gaps in the membrane, so it is easier for molecules to pass through it.

At low temperatures, phospholipids have little energy and are packed closely together to make the membrane rigid. This causes a decrease in permeability and restricts molecules from crossing the membrane

At very low temperatures, ice crystals can form which pierce the cell membrane and increase the permeability.

ETHANOL

Ethanol causes the cell-surface membrane to rupture, releasing the betalain pigment from the cell. Higher concentrations of ethanol will cause more disruption to the membrane and more gaps will form. Thus, as concentration of ethanol increases, the permeability of the cell-surface membrane also increases.

REQUIRED PRACTICAL #5

What is the risk assessment for a dissection?

Biohazard [can contaminate and spread unwanted pathogens. Use disinfectant and wash hands are touching it].

Disinfectant [it is flammable so keep away from naked flames. Seek out assistance if anything occurs].

Sharp Tools [may cut individuals so move glass slowly and keep it away from edge of desk. In emergencies, apply pressure to cuts and seek medical attention].

REQUIRED PRACTICAL #5

How do you draw a diagram from a dissection?

Use a sharp HB pencil

Make sure drawing is large enough for detail and labels (at least half a page advisable)

No shading

Use single and continuous lines (no sketching)

Label lines should be drawn with a ruler

Do not cross label lines

Lines should not have any arrows

Label all the structures

Do not use colour

Include a magnification/scale