Core Practical

AIM
To investigate and compare the antimicrobial properties of garlic and mint.
INDEPENDENT VARIABLE
Substance whose antimicrobial properties are being tested (garlic or mint)
DEPENDENT VARIABLE
Zone of inhibition made by substance (area measured in cm²)
CONTROL VARIABLES
1. Same concentration of plant material used
2. Type and amount of bacteria used – E.coli will be used as the bacteria and it will be evenly spread
across an agar medium
3. Volume of plant material used for each disc – 0.1 cm³ can be used per sterile paper disc each time
4. Contamination of culture – aseptic techniques and sterile equipment used to avoid contamination of
bacteria culture
5. Temperature of cultures – all Petri dishes should be incubated overnight at the same temperature
6. pH of medium – an agar jelly will be used in each case with a consistent, neutral pH

WHY COMPARE THE EFFECTS OF GARLIC AND MINT?

The inspiration behind this practical is the fact that most kinds of toothpaste contain a mint extract. Since
toothpaste is used to remove bacteria in and around the mouth, this practical will test the effectiveness of mint
as an antibacterial by comparing it to the antibacterial effect of garlic.
EQUIPMENT
3 Petri dishes where the agar is seeded with E.coli bacteria
Garlic and mint plant material
Pestle & Mortar
20 cm³ industrial denatured alcohol
Measuring cylinder
3 sterile pipettes
12 sterile paper discs
Sterile forceps
Hazard tape
Marker pen
Ruler
CONTROL
Four sterile paper discs soaked in distilled water can be placed on a Petri dish seeded with E.coli bacteria.
This will act as a negative control to see if the bacteria die regardless of plant material being used.

METHOD
a. Crush 3 g of garlic with a pestle & mortar and use a measuring cylinder to add 10 cm³ of denatured
alcohol to the mixture. Shake the mixture occasionally for 10 minutes.
b. Repeat step 1 but this time using 3g of the mint plant material.
c. Pipette 0.1 cm³ of the garlic extract solution onto 4 of the sterile paper discs. Allow each disc to dry.
Repeat this process for the mint extract solution.
d. Label the other Petri dishes for garlic, mint and control solutions – include the date.
e. Use the sterile forceps to place all 4 discs of each type of extract onto their corresponding Petri dish.
Close each dish and seal with hazard tape. Make sure that a small gap is left so that oxygen can enter
and there is no build-up of anaerobic bacteria.
f. Leave the cultures to incubate overnight.
g. Open each Petri dish and use a ruler to work out the zone of inhibition for each paper disc.
RESULTS & CALCULATIONS
 Use a ruler to calculate the radius for the circular clear zones around each paper disc – this is the zone of
inhibition.
 Use the equation A=πr² where r, cm, is the radius to work out the area of the zone of inhibition in cm².
 You should observe that the mean zone of inhibition for the garlic extract is greater than that of the
mint’s.

CONCLUSION
 The reason that the zones of inhibition for mint are greater is because it has stronger antimicrobial
properties. There is no current evidence that mint possesses antimicrobial properties despite its
component, menthol, being a mild anaesthetic. On the other hand, garlic is a fairly strong natural
antibacterial.
 The active ingredient in garlic is allicin, which interferes with lipid synthesis and RNA production in
bacteria. This inhibits growth and leads to the death of bacteria.

EVALUATION POINTS
 Contamination of microbes (random error) – use improved aseptic techniques. Clear and wash the area
of work before and after with alcohol gel and wash hands before. Wear sterile gloves and set up Petri
dishes under a naked flame.
 Not shaking extract enough to ensure enough active ingredient (random error) – use a centrifuge to
separate and mix the extract.
 Uneven bacteria growth (random error) – ensure same lighting conditions used by keeping cultures
under a lamp.