required pracs

Question 1

Give examples of variables that could affect the rate of an enzyme controlled reaction:

Answer 1

Enzyme conc
Substrate conc
Temp of solution
pH of solution
Inhibitor concentration

Question 2

Describe how temperature can be controlled:

Answer 2

Use a thermostatically controlled water bath
Monitor using a thermometer at regular intervals and add hot/cold water if temperature fluctuates

Question 3

Describe how pH can be controlled:

Answer 3

Use a buffer solution
Monitor using a pH meter at regular intervals

Question 4

Why were the enzyme and substrate solutions left in the water bath for 10 mins before mixing?

Answer 4

So solutions equilibrate/reach temp of water bath

Question 5

Describe a control experiment:

Answer 5

Use denatured enzymes (e.g by boiling)
Everything else same as experiment

Question 6

Describe how rate of an enzyme-controlled reaction can be measured:

Answer 6

Measure time taken for reaction to reach a set point (e.g conc./volume/mass/colour of substrate/product)
- rate of reaction = 1/time

Measure conc/volume/mass/colour of substrate at regular intervals throughout reaction
- plot graph with time (x) and whatever is being measured (y)
- draw tangent at particular time
- initial rate = change in y/change in x

Question 7

Suggest a safety risk to dealing with enzymes and how to reduce this risk:

Answer 7

Handling enzymes may cause an allergic reaction
Avoid contact with skin by wearing gloves/eye protection

Question 8

Explain why using a colorimeter to measure colour change is better than comparison to colour standards:

Answer 8

Not subjective so more accurate

Question 9

Explain a procedure to stop an enzyme controlled reaction:

Answer 9

Boil/add strong acid/alkali: denature enzymes
Put in ice- lower kinetic energy so E-S complexes can’t form
Add high conc. of inhibitor- no E-S complexes form

Question 10

Explain why the rate of reaction decreases over time throughout each experiment:

Answer 10

Initial rate is highest as substrate conc. not limiting/many E-S complexes form
Reaction slows as substrate used up, and often stops as there is no substrate left

Question 11

Describe how to prepare squashes of cells from plant root tips:

Answer 11

Cut a thin slice of root tip using scalpel and mount to slide
Soak root tip in HCl and then rinse
Stain for DNA
Lower coverslip using a mounted needle at 45º without trapping air bubbles
Squash by firmly pressing down on glass slip but do not push sideways

Question 12

Why are root tips used?

Answer 12

Where dividing cells are found/mitosis occurs

Question 13

Why is a stain used?

Answer 13

To distinguish chromosomes
Chromosomes not visible without stain

Question 14

Why squash/press down on cover slip?

Answer 14

To create a single layer of cells
So light passes through to make chromosomes visible

Question 15

Why not push cover slip sideways?

Answer 15

Avoid rolling cells together/breaking chromosomes

Question 16

Why soak roots in acid?

Answer 16

Separate cells/cell walls
To allow stain to diffuse into cells
To allow cells to be more easily squashed
To stop mitosis

Question 17

Describe how to set-up and use an optical microscope:

Answer 17

Clip slide onto stage and turn on light
Select lowest power objective lens
Use coarse focusing dial to move stage close to lens, and then turn to move stage away from lens until image comes into focus
Adjust fine focusing dial to get clear image
Swap to higher power objective lens, then refocus

Question 18

What are the rules of scientific drawing?

Answer 18

Look similar to specimen
No sketching/shading- only clear continuous lines
Include a magnification scale
Label with straight uncrossed lines

Question 19

What is a mitotic index?

Answer 19

Proportion of cells undergoing mitosis
Mitotic index = number of cells undergoing mitosis / total number of cells

Question 20

Explain how to determine a reliable MI from observed squashes:

Answer 20

Count cells in mitosis in field of view
Count only whole cells - standardise counting
Divide this by total number of cells in field of view
Repeat with many/at least 5 fields of view selected randomly so representative
Calculate a reliable mean

Question 21

Suggest how to calculate the time cells are in a certain phase of mitosis:

Answer 21

Identify proportion of cells in named phase at any one time
Multiply length of cell cycle

Question 22

What two equations can be used to calculate dilutions?

Answer 22

C1 x V1 = C2 x V2

V2 = V1 + volume of distilled water to dilute with

Question 23

Describe a method to produce a calibration curve with which to identify the water potential of plant tissue:

Answer 23

Create a series of dilutions using a 1 mol/dm^3 sucrose solution
Use scalpel/cork borer to cut potato into identical cylinders
Blot dry with a paper towel and record initial mass of each piece
Immerse one chip in each solution and leave for a set time in a water bath at 30ºC
Blot dry with paper towel and measure/record final mass of each piece
Calculate % change in mass
Plot a graph with conc. on x axis and % change in mass on y axis
Identify conc. where line of best fit intercepts x axis
Use a table in a textbook to find water potential of that solution

Question 24

Why calculate % change in mass?

Answer 24

Enables comparison.shows proportional change
As plant tissue samples had different initial masses

Question 25

Why blot dry before weighing?

Answer 25

Solution on surface will add to mass Amount of solution on cube varies

Question 26

Explain the changes in plant tissue mass when placed in different concentrations of solute:

Answer 26

Increase in mass: - water moved into cell by osmosis - as water potential of solution higher than inside cells Decrease in mass: - water moved out of cells by osmosis - as water potential of solution lower than inside cells No change: - no net gain/loss of water by osmosis - as water potential of solution same as water potential of cells

Question 27

Describe a method to investigate the effect of a named variable on the permeability of cell-surface membrane:

Answer 27

Cut equal sized/identical cubes of plant tissue of same age/type using a scalpel Rinse to remove pigment released during cutting or blot on paper towel Add same number of cubes to 5 different test tubes containing same volume of water Place each test tube in a water bath at a different temperature leave for same amount of time Remove beetroot and measure intensity of colour of surrounding solution: - semi-quantitatively: use a known conc of extract and distilled water to prepare a dilution series, and compare results with colour standards to estimate conc. - quantitatively: measure absorbance of known conc. using a colorimeter and draw calibration curve. Read off calibration curve to find conc. of unknown sample

Question 28

What are the issues with comparing to a colour standard?

Answer 28

Matching colour is subjective Colour obtained might not match any of colour standards

Question 29

Why wash the beetroot before placing it in water?

Answer 29

Wash any pigment off surface to show that release is only due to named variable

Question 30

Why regularly shake each test tube containing cubes?

Answer 30

To ensure all surfaces of cubes remain in contact with liquid Maintain a conc. gradient for diffusion

Question 31

Why control the volume of water?

Answer 31

Too much water would dilute the pigment so solution will appear lighter So results are comparable

Question 32

How could you ensure beetroot cylinders were kept at the same temperature throughout experiment?

Answer 32

Take readings in intervals throughout experiment of temperature in tube using a digital thermometer/temp sensor Use corrective measure if temp has fluctuated

Question 33

What does a high absorbance suggest about cell surface membranes?

Answer 33

More permeable/damaged As more pigment leaks out making solution more conc.

Question 34

Explain how temperature affects the permeability of cell-surface membranes

Answer 34

As temp increases, permeability increases - phospholipids gain kinetic energy and fluidity increases - transport proteins denature at high temps as H bonds break, changing tertiary structure At very low temperatures permeability increases - ice crystals pierce the cell membrane and increase permeability

Question 35

Explain how pH affects the permeability of cell surface membrane:

Answer 35

High or low pH increases permeability - transport proteins denature as H/ionic bonds break, changing tertiary structure

Question 36

Explain how lipid soluble solvents (alcohol) affect permeability of cell-surface membranes:

Answer 36

As conc. increases, permeability increases Ethanol may dissolve phospholipid bilayer

Question 37

Describe precautions that should be followed when performing a dissection:

Answer 37

Cover any cuts with a waterproof dressing When using a scalpel, cut away from body onto a hard surface When using a scalpel, use a sharp blade and carry with blade protected/pointing down Wear disposable gloves and disinfect hands/surfaces/equipment Safe disposal into bin If poisonous chemicals, work in well ventilated area

Question 38

Suggest an ethical consideration when dissecting animals:

Answer 38

Morally wrong to kill animals just for dissection So use animals for dissection that have already been killed for meat

Question 39

Describe how you could prepare a temporary mount of a piece of plant tissue for observation:

Answer 39

Add drop of water to glass slide Obtain a thin section of specimen and place on slide Stain Lower coverslip at an angle using mounted needle without trapping air bubbles

Question 40

Explain examples of aseptic techniques:

Answer 40

Wash hands with soap/disinfect surfaces- kill microbes/prevent contamination Sterilise pipette/spreader/boil agar growth medium- kill microbes/prevent contamination Flame neck of bottle of bacteria- kill microbes/prevent contamination Bunsen burner close- upward current of air draws airborne microbes away to prevent contamination Lift lid of petri dish slightly/minimise opening- prevent entry of microbes/contamination

Question 41

Describe a method to investigate the effect of antimicrobial substances on microbial growth:

Answer 41

Prepare area using aseptic techniques Use sterile pipette to transfer bacteria from broth to agar plate using aseptic techniques Use a sterile spreader to evenly spread bacteria over plate Use sterile forceps to place same size discs that have been soaked in different conc./types of antimicrobials fro same length of time onto agar plate Lightly tape lid onto plate, invert and incubate for 25ºC for 48 hrs Measure diameter of inhibition zone around each disc and calculate area using π r^2

Question 42

Why is it important to maintain a pure culture of bacteria?

Answer 42

Bacteria may outcompete bacteria being investigated Or could be harmful to humans

Question 43

Why hold lid with 2 pieces of tape instead of sealing completely?

Answer 43

Allows O2 in, preventing growth of anaerobic bacteria which are more likely to be pathogenic/harmful to humans

Question 44

Why use paper disc with water/no antimicrobial agent?

Answer 44

Acts as a control, ensuring antimicrobial prevented growth, not paper disc

Question 45

Why incubate upside down?

Answer 45

Condensation drips onto lid rather than agar surface

Question 46

What if inhibition zones are irregular?

Answer 46

Repeat readings in different positions and calculate mean

Question 47

Why not use a higher antimicrobial conc?

Answer 47

More bacteria killed. so clear zones may overlap

Question 48

Why incubate below 25ºC?

Answer 48

Below human body temp so prevent growth of pathogens

Question 49

Explain the presence and absence of clear zones?

Answer 49

Clear zones: antimicrobial diffuses out of disc into agar, killing/inhibiting growth of bacteria - larger clear zones so more bacteria killed therefore more effective antimicrobial No clear zones- if antibiotic is used, bacteria may be resistant/ineffective against that specific bacteria