Paper 1 Practicals

Question 1

describe a method to investigate the effect of temperature on the rate of enzyme (trypsin) activity

Answer 1

1. make a control sample of just milk solution
2. add milk solution to 5 test tubes labelled with 20C, 30C, 40C, 50C & 60C
3. in separate test tubes at 2cm^3 trypsin solution & 2cm^3 pH7 buffer solution
4. put test tubes containing milk solution & trypsin with buffer into a water bath at 20C for 5 mins
5. repeat for all temperatures
6. add trypsin & buffer solution into the test tube of milk solution, place back into water bath & time for 4 minutes using a stopclock
7. after 4 mins, put contents of test tube into a cuvette into a colorimeter & measure absorbance immediately
   (alternatively time how long it takes for solution to go colourless)
8. plot graph of temperature vs absorbance
9. the smaller the absorbance, the greater the rate of enzyme activity

Question 2

describe a method to use an optical microscope to identify the stages of mitosis in root tip squashes & to calculate mitotic index

Answer 2

1. heat 1 mol of HCl in 60C hot water bath
2. cut root sample using a scalpel, place in HCl & leave in hot water bath for 5 minutes
3. wash roots in cold distilled water & cut the tips of the roots with a scalpel
4. place into acetic orcein stain & leave in water bath for 5 mins
5. place stained root tip on slide
6. place cover slip on top of root tip & squash them down
7. put the prepared slide under the microscope & set objective lens to lowest magnification
8. use the coarse & fine adjustment knob to focus the image
9. on highest magnification objective lens (X40), count the number of cells in mitosis & the total number of cells
10. make a biological drawing
11. mitotic index = # cells with visible chromosomes/total # cells

Question 3

describe a method to identify the water potential of potato tuber cells by a dilution series

Answer 3

1. make a series of dilutions with distilled water of 1M sucrose (or salt) solution at 0.0M, 0.2M, 0.4M, 0.6M, 0.8M & 1.0M sucrose
2. measure 10cm^3 of each dilution into separate test tubes
3. use a ruler & scalpel to cut out 6 potato chips of equal dimensions. blot with paper towel to remove excess water (do not squeeze)
4. weigh each potato chip using a mass balance
5. place 1 potato chip in each test tube & leave for 20 mins
6. remove each potato chip, blot dry with paper towel & reweigh them
7. calculate the % change in mass for each potato chip
8. plot graph of conc vs % change in mass

Question 4

describe a method to investigate the effect of temperature on the permeability of cell surface membranes (beetroot leakage)

Answer 4

1. cut beetroot into 10 equal dimension cubes
2. label 10 test tubes with relevant temperature (20-80) & put 5cm^3 distilled water in each
3. put each test tube in a water bath at the given temperature
4. put 5 beetroot cubes into the water bath for 1 minute then put in labelled test tube for 20mins
5. after 20mins, pour coloured water into a cuvette & into a colorimeter
6. set colorimeter to blue filter, calibrate with distilled water, then measure absorbance
7. plot graph of temperature vs absorbance
8. higher absorbance = higher pigment conc. = higher membrane permeability

Question 5

what must be done before using a colorimeter?

Answer 5

calibrate colorimeter with distilled water
= zero it

Question 6

describe precautions when dissecting e.g. a mammalian heart & lungs

Answer 6

disinfect surfaces
wear gloves
observe outside of heart & lungs before dissection
use scalpel & make sharp, precise cuts

Question 7

a. describe a method to investigate the effect of antimicrobial substances on microbial growth

Answer 7

1. use aseptic technique (describe throughout)
2. use a sterile pipette & flame neck of bottle to remove some E.coli broth
3. lift lid of petri dish facing the Bunsen flame & place a few drops of E.coli on the agar plate
4. flame a glass spreader, let it cool, then spread bacteria evenly on surface of agar
5. use sterile forceps to place filter paper discs soaked in antimicrobial solution onto the centre of the agar plate
6. secure the lid of the petri dish with 2 pieces of tape & put upside down in incubator @ 25C for 48 hrs
7. after incubation, measure the diamater & calculate the area of the zone of inhibition for each antibiotic using A=pir^2

Question 8

describe & explain examples of aseptic technique

Answer 8

wash hands with soap & disinfect surfaces
use sterile pipette
flame the neck of the bottle, glass spreader & forceps in a (roaring) Bunsen flame
lift the lid of the petri dish towards the Bunsen safety flame, which creates convection current/upwards air movement so microogranisms moved away
discard apparatus in a beaker of disinfectant

Question 9

b. describe a method to investigate the effects of antimicrobial substances on the growth of bacterial populations in a broth culture

Answer 9

1. use aseptic technique (describe throughout)
2. use a sterile pipette & flame neck of bottle to remove some M. luteus broth & place 1cm^3 in the first bottle of disinfectant
3. remove 1cm^2 from first bottle & place in 2nd bottle & repeat for all bottles, removing 1cm^3 from final bottle & discarding it
4. for each dilution take 0.1cm^3 with a sterile pipette
5. lift lid of petri dish towards Bunsen flame & place dilution solution on agar plate
6. flame a glass spreader, let it cool, & spread dilution evenly over the surface of the agar
7. secure the lid of the petri dish with 2 pieces of tape & put upside down in incubator @ 25C for 48 hrs
8. after incubation, count individual colonies of bacteria, X by dilution factor X 10 to calculate an estimation for # of bacterial cells in original sample per mL