How chemical defences work
Definition
Antimicrobial substance
A chemical produced by an organism that kills microorganisms or slows their growth.
Definition
Pathogen
A microorganism that causes a communicable disease, such as a virus, a bacterium, a fungus or a protist.
- A chemical defence is a substance the plant makes that attacks the pathogen itself, rather than a barrier that keeps the pathogen outside.
- This matters because a barrier is useless once it has been broken, and a chemical can still act on a pathogen that has already got in.
- Some of these substances are present in the tissues all the time, so a pathogen meets them the moment it arrives.
- Others are made only after the plant detects damage or infection, which saves the energy and raw materials that making them all year would cost.
- Because the substances are soluble they diffuse through the tissue around the infected area, so they reach the pathogen wherever it has spread to.
Key Idea
- Chemical defences fall into two jobs: killing microorganisms, and making the plant unpleasant or dangerous to eat.
- Both jobs reduce disease, because an animal that stops feeding also stops making the wounds that let pathogens in.
Antibacterial and antifungal substances
Definition
Fungus
A eukaryotic organism that is either single-celled or built from threads called hyphae and feeds by releasing enzymes onto its food.
Definition
Zone of inhibition
The clear area of agar surrounding a disc of antimicrobial substance in which bacteria have been prevented from growing.
- Antibacterial substances damage the cell wall or the cell membrane of a bacterium, so it either bursts or can no longer take in what it needs to reproduce.
- Garlic produces allicin when its tissue is crushed, and mint and witch hazel produce oils that act in the same way.
- Antifungal substances stop fungal threads growing into the tissue, usually by breaking down the membranes of the growing hyphae.
- Oat roots contain a substance that punches holes in fungal membranes, and the bark of many trees is rich in compounds that do the same.
- Many plants make both kinds at once, which is why plant tissue rots far more slowly while it is alive than it does after it dies.
- The effectiveness of an extract is compared by growing bacteria on agar and measuring the clear zone of inhibition it produces, since a larger zone means the substance has diffused further and still killed bacteria.
Practical
Comparing the antibacterial effect of plant extracts
- Aim: to find out which of several plant extracts is the most effective at inhibiting the growth of bacteria, by measuring the zones of inhibition they produce on a seeded agar plate.
- Apparatus: sterile agar plates already seeded with a harmless culture of bacteria, sterile filter paper discs, sterile forceps, a pestle and mortar, fresh garlic, mint and onion, distilled water, a measuring cylinder, a filter funnel and filter paper, a Bunsen burner, adhesive tape, a marker pen, an incubator set to 25 ∘C25\ ^\circ\text{C}25 ∘C, a 30 cm30\ \text{cm}30 cm ruler and eye protection.
- Method, making the extracts:
- Weigh out 5 g5\ \text{g}5 g of the first plant material and grind it in the mortar with 10 cm310\ \text{cm}^310 cm3 of distilled water until it forms a smooth paste.
- Pour the paste through filter paper in a funnel to remove the solid fragments, because a lump of tissue sitting on the agar would block the disc rather than release the extract evenly.
- Repeat with the same mass and the same volume of water for each of the other plants, so that concentration is kept the same throughout.
- Method, setting up the plate:
- Turn the plate over and draw four evenly spaced sectors on the base with the marker, labelling one for each extract and one for the control.
- Soak a filter paper disc in the first extract for the same length of time you will use for all of them, then hold it against the side of the beaker to drain off the surplus.
- Pass the forceps through the Bunsen flame and let them cool, then lift the lid of the plate just far enough to reach in at an angle and lay the disc flat in its sector.
- Press the disc down gently so its whole surface touches the agar, since a disc resting on one edge releases extract into the agar unevenly.
- Repeat for the other extracts, flaming the forceps between each one, and place a disc soaked only in distilled water in the fourth sector as the control.
- Method, incubating and measuring:
- Fix the lid with two short pieces of tape on opposite sides, leaving the rest of the rim open so that air can still reach the culture.
- Incubate the plate upside down at 25 ∘C25\ ^\circ\text{C}25 ∘C for 484848 hours, so that condensation collects in the lid instead of dripping onto the agar and smearing the colonies.
- Without opening the plate, measure the diameter of each clear zone in millimetres across two directions at right angles and take the mean.
- Set up three plates for each extract and calculate a mean for every one, so that a single unusual plate does not decide the result.
- Results: a cloudy lawn of bacteria covers the agar apart from a clear circle around each disc that released an antibacterial substance, and the control disc has no clear zone at all.
- Maths: convert each mean diameter to a radius with r=d2r=\dfrac{d}{2}r=2d and find the area of the zone with A=πr2A=\pi r^{2}A=πr2, because area is proportional to the number of bacteria killed while diameter is not.
- A disc giving a zone of diameter 18 mm18\ \text{mm}18 mm has an area of A=π×92=254 mm2A=\pi \times 9^{2}=254\ \text{mm}^{2}A=π×92=254 mm2 to three significant figures.
- Watch out: the control disc is what proves the water itself killed nothing, discs must be identical in size and soaked for equal times, plates must all be incubated together at one temperature, and a zone measured through a closed lid is measured accurately only if the ruler is held flat against the base.
- Safety: wear eye protection, keep the incubator at 25 ∘C25\ ^\circ\text{C}25 ∘C so that bacteria which grow at human body temperature are not favoured, never reopen an incubated plate, tie long hair back near the Bunsen burner, wash your hands after handling plates and hand every used plate in for sterilising before disposal.
Chemicals that deter animals
Definition
Herbivore
An animal that feeds only on plant material.
- Poisons kill or sicken an animal that eats the plant, so the rest of that population learns to leave the species alone.
- Foxglove leaves contain a substance that disturbs the rhythm of the heart, deadly nightshade berries contain one that blocks nerve signals, and tobacco leaves contain nicotine, which is toxic to insects.
- Bitter-tasting chemicals work without harming the animal, because a mouthful that tastes unpleasant is spat out and the plant is left alone.
- Tannins in oak and tea leaves bind to proteins in the mouth and gut, which makes the leaf taste harsh and makes its protein harder to digest.
- Fewer animals feeding means fewer holes bitten in the leaves, and since almost every infection starts at a wound this cuts the rate of disease as well.
- Some plants release scent chemicals into the air as soon as a caterpillar starts to feed, and these attract wasps that lay their eggs in the caterpillar.
Common Mistake
- Do not describe a thorn or a thick cuticle as a chemical defence.
- A chemical defence is a substance the plant produces, so the answer must name a poison, a bitter compound or an antimicrobial substance.
Plant defence chemicals as medicines
- A substance that kills a bacterium in a leaf may also kill the same bacterium in a person, so plant defences have long been a starting point for medicines.
- Aspirin is based on a compound from willow bark, where the plant uses it as part of its response to infection, and it is now used to relieve pain and fever.
- Quinine comes from the bark of the cinchona tree and was the first effective treatment for malaria.
- Digitalis, the foxglove poison that deters herbivores, is used in carefully measured doses to steady a failing heart.
- The dose is what separates a poison from a medicine, which is why these substances are extracted, purified and measured precisely rather than taken as raw plant material.
Exam technique
- Say what the chemical does to the pathogen or to the animal, because naming the compound alone rarely earns the mark.
- In a zone of inhibition question, compare the treated discs with the control before drawing any conclusion about which extract worked.
- A larger clear zone shows the substance is more effective, and it is worth adding that this is because it stopped growth even where it had diffused further and become more dilute.
Self review
- State the difference between a physical and a chemical plant defence.
- Explain how an antibacterial substance kills a bacterium.
- Give two ways a plant uses chemicals to stop animals eating it.
- Why does deterring herbivores also reduce the amount of disease a plant suffers?
- Explain why a disc soaked only in distilled water must be included when testing plant extracts on an agar plate.
