A Level Biology

• Describe how pigments from a leaf of a plant can be isolated with paper chromatography

  1. Crush leaves with solvent to extract pigments

  2. Draw a pencil line on filter/chromatography paper, 1cm above bottom

  3. Add a drop of extract to line(point of origin)

  4. Stand paper in boiling tube of(organic)solvent below point of origin

  5. Add lid and leave to run(solvent moves up,carrying dissolved pigments)

  6. Remove before solvent reaches top and mark solvent front with pencil
• Explain why the origin should be drawn in pencil rather than ink.(2)

  ● Ink is soluble in solvent

  ● So ink would mix with pigments/line would move
• Explain why the point of origin should be above the level of the solvent.(2)

  ● Pigments are soluble in solvent

  ● So would run off paper/spots dissolve into solvent
• Explain why a pigment may not move up the chromatography paper in one solvent.(1)

  ● May be soluble in one solvent but insoluble in another
• Describe how pigments can be identified

  ● Rf value=distance moved by spot/distance moved by solvent front

  ● Compare Rf value to published value
• Explain why the solvent front should be marked quickly once chromatography paper is removed.(1)

  ● Once solvent evaporates, solvent front not visible
• Explain why the centre of each pigment spot should be measured.(1)

  ● Standardises readings as pigment is spread out

  ● So allows comparisons to be made
• Explain why the obtained Rf values were similar, but not identical, to the published values.(1)

  ● Different solvent/paper/runningconditions may affect Rf value
• Explain why Rf values are used and not the distances moved by pigment spots.(2)

  ● Solvent/pigment moves different distances

  ● Rf value is constant for same pigment/can be compared
• Describe the role of the enzyme dehydrogenase in photosynthesis

  ● Catalyses the reduction of NADP in the light-dependent reaction

  ○ NADP accepts (gains) electrons from photoionisation of chlorophyll / photolysis of water
• Describe how rate of dehydrogenase activity in extracts of chloroplasts can be measured

  1. Extract chloroplasts from a leaf sample using the method in '2.1.3 Methods of studying cells'

  2. Set up test tubes as follows:

  A. Control 1- set volume of DCPIP (redox indicator dye, electron acceptor), water and chloroplasts in isolation medium, covered in foil to block light

  B. Control 2-set volume of DCPIP, water and isolation medium without chloroplasts

  C. Standard-set volume of water and chloroplasts in isolation medium, without DCPIP D. Experiment- set volume of DCPIP, water and chloroplasts in isolation medium

  3. Shine light on test tubes and time how long to it takes for DCPIP to turn from blue (oxidised) to colourless (reduced) in tube D (tube A and B should show no change)

  ○ Compare to a colour standard(tube C)to identify endpoint

  4. Rate of dehydrogenase activity (s-1) = 1/time taken
• Give examples of variables that could be controlled.(3)

  ● Source of chloroplasts

  ● Volume of chloroplast suspension ● Volume/concentrationof DCPIP
• Explain the purpose of control 1 (tube A).(2)

  ● Shows light is required for DCPIP to decolourise

  ● Shows that chloroplasts alone do not cause DCPIP to decolourise
• Explain why DCPIP in control 1 stays blue. (2)

  ● No light so no photoionisation of chlorophyll

  ● So no electrons released to reduce DCPIP
• Explain the purpose of control2 (tube B).(2)

  ● Shows chloroplasts are required for DCPIP to decolourise

  ● Shows that light alone does not cause DCPIP to decolourise
• Explain why DCPIP changes from blue to colourless. (2)

  ● DCPIP is a redox indicator/DCPIP gets reduced by electrons

  ● From photoionisation of chlorophyll
• Suggest a limitation with the method and how the experiment could be modified to overcome this.(4)

  ● Endpoint(colour change)is subjective

  ● Use a colorimeter

  ● Measure light absorbance of sample at set time intervals

  ● Zero colorimeter using the colour standard
• Why is respiration important?

  ● Respiration produces ATP (to release energy) ● For active transport, protein synthesis etc.
• strucutre of the mitochondria

  outer mem

  cristae - inner mem fold

  matrix:

  - small 70S ribosomes

  - circular DNA
• Summarise the stages of aerobic & anaerobic respiration

  Aerobic respiration Anaerobic respiration

  1. Glycolysis- cytoplasm (anaerobic) 2. Link reaction- mitochondrial matrix

  3. Krebs cycle- mitochondrial matrix

  4. Oxidative phosphorylation- inner mitochondrial membrane

  1. Glycolysis- cytoplasm 2. NAD regeneration- cytoplasm
• Describe the process of glycolysis

  1.. Glucose phosphorylated to glucose phosphate

  ○ Using inorganic phosphates from 2ATP 2. Hydrolysed to 2 x triose phosphate

  3. Oxidised to 2 pyruvate

  ○ 2 NAD reduced

  ○ 4 ATP regenerated(net gain of 2)
• Explain what happens after glycolysis if respiration is anaerobic

  1. Pyruvate converted to lactate (animals & some bacteria) or ethanol (plants & yeast) 2. Oxidising reduced NAD →NAD regenerated

  3. So glycolysis can continue (which needs NAD) allowing continued production of ATP
• Suggest why anaerobic respiration produces less ATP per molecule of glucose than aerobic respiration

  ● Only glycolysis involved which produces little ATP (2 molecules)

  ● No oxidative phosphorylation which forms majority of ATP (around 34 molecules)
• What happens after glycolysis if respiration is aerobic?

  Pyruvate is actively transported into the mitochondrial matrix
• Describe the link reaction

  1. Pyruvate oxidised (and decarboxylated) to acetate

  ○ CO2 produced

  ○ Reduced NAD produced(picks up H) 2. Acetate combines with coenzyme A, forming Acetyl Coenzyme A

  
  

  Products per glucose molecule: 2 x Acetyl Coenzyme A, 2 XCO2 and2XreducedNAD
• Describe the Krebs cycle

  1. Acetyl coenzyme A (2C) reacts with a 4Cmolecule

  ○ Releasing coenzyme A

  ○ Producing a 6C molecule that enters the Krebs cycle

  2. In a series of oxidation-reduction reactions, the 4C molecule is regenerated and:

  ○ 2xCO2 lost

  ○ Coenzymes NAD & FAD reduced

  ○ Substrate level phosphorylation (direct transfer of Pi from intermediate compound to ADP) →ATP produced

  
  

  Products per glucose molecule: 6 x reduced NAD, 2 x reduced FAD, 2 x ATP and4xCO2
• Describe the process of oxidative phosphorylation

  1. Reduced NAD/FAD oxidised to release H atoms → split into protons (H+) and electrons (e-)

  2. Electrons transferred down electron transfer chain (chain of carriers at decreasing energy levels)

  ○ By redox reactions

  3. Energy released by electrons used in the production of ATP from ADP + Pi (chemiosmotic theory):

  ○ Energy used by electron carriers to actively pump protons from matrix → intermembrane space ○ Protons diffuse into matrix down an electrochemical gradient, via ATP synthase (embedded) ○ Releasing energy to synthesise ATP from ADP+Pi

  4. In matrix at end of ETC, oxygen is final electron acceptor(electrons can't pass along otherwise)

  ○ So protons, electrons and oxygen combine to form water
• Give examples of other respiratory substrates

  Break down products of lipids and amino acids,which enter the Krebs cycle. For example:

  ● Fatty acids from hydrolysis of lipids→ converted to Acetyl Coenzyme A

  ● Amino acids from hydrolysis of proteins→ converted to intermediates in Krebs cycle
• Describe how are spirometer can be used to measure the rate of aerobic respiration(by measuring oxygen up take)

  1. Add set mass of single-celled organism eg. yeast to set volume/conc. of substrate eg. glucose 2. Add a buffer to keep pH constant

  3. Add a set volume/conc. of a chemical that absorbs CO2 eg.sodium hydroxide

  4. Place in water bath at a set temperature and allow to equilibrate 5. Measure distance moved by coloured liquid in a set time
• Explain why the liquid moves. (4)

  ● Organisms aerobically respire so take in O2

  ● CO2 given out but absorbed by sodium hydroxide solution

  ● So volume of gas and pressure in container decrease

  ● Fluid in tube moves down pressure gradient towards organism
• Explain why the respirometer apparatus is left open for 10 minutes.(1)


  ● Allow apparatus to equilibrate

  ● Allow for overall pressure expansion/change throughout

  ● Allow respiration rate of organisms to stabilise
• Explain why the apparatus must be air tight.(2)

  ● Prevent air entering or leaving

  ● Would change volume and pressure, affecting movement of liquid
• Describe a more accurate way to measure volume of gas.(1)

  ● Use a gas syringe
• Suggest a suitable control experiment and explain why it is necessary.(2)

  ● No organisms OR use inert objects OR use dead organisms AND all other conditions/apparatus/ equipment the same

  ● To show that(respiring) organisms are causing liquid to move/ taking up oxygen/causing the change in volume/pressure
• Describe how are spirometer can be used to measure the rate of anaerobic respiration(by measuring carbon dioxide release)

  ● Repeat experiment as above but remove chemical that absorbs CO2

  ● Make conditions anaerobic, for example: ○ Layer of oil/liquid paraffin above yeast to stop O2 diffusing in

  ○ Add a chemical that absorbs O2

  ○ Leave for an hour to allow O2 to be respired and used up
• Explain why the liquid moves.(3)

  ● Yeast anaerobically respire so release CO2 ● So volume of gas and pressure in container increase

  ● So fluid in capillary tube moves down a pressure gradient away from organism
• Explain why the apparatus is left for an hour after the culture has reached a constant temperature. (1)

  ● Allow time for oxygen to be used/respired
• Describe how rate of respiration can be calculated

  1. Calculate volume of O2 / CO2 consumed / released (calculate area of a cylinder)

  a. Calculate cross-sectional area of capillary tube using π r²

  b. Multiply by distance liquid has moved

  2. Divide by mass of organism and time taken

  3. Units- unit for volume per unit time per unit mass eg. cm3min-1g-1
• Describe how redox indicator dyes such as Methylene blue can be used to measure rate of respiration

  ● Redox indicators (eg. methylene blue) change colour when they accept electrons becoming reduced

  ● Redox indicators take up hydrogens and get reduced instead of NAD/FAD →modelling their reactions

  
  

  1. Add a set volume of organism eg. yeast and a set volume of respiratory substrate eg. glucose to tubes

  2. Add a buffer to keep pH constant

  3. Place in water bath at a set temperature and allow to equilibrate for 5 mins

  4. Add a set volume of methylene blue, shake for a set time (do not shake again)

  5. Record time taken for colour to disappear in tube

  6. Rate of respiration(s-1) = 1 / time (sec)
• Give two examples of variables that could be controlled. (2)

  ● Volume of single-celled organism

  ● Volume/conc./type of respiratory substrate ● Temperature(with a water bath)

  ● pH(with a buffer)

  ● Volume of redox indicator (only control)