Core Practical 9 - 09/02/22
Core practical 9 - investigating the antimicrobial properties of plants including aseptic
techniques for the safe handling of bacteria
Aim : To investigate and compare the antimicrobial properties of garlic and mint on bacterial
growth using agar plates and measuring the zones of inhibition.
Introduction
Dental caries are localised, transmissible infectious processes that end up in the destruction
of hard dental tissue. It results from accumulation of plaque on the surface of the teeth and
microbes due to poor oral hygiene. For example Streptococcus mutans is a main pathogen
of dental caries which has a role in the fermentation of carbohydrates resulting in acid
production and demineralisation of tooth enamel. Therefore, chemical agents with
antimicrobial activity were incorporated into dental products as a method of reducing levels
of oral bacteria. (Prasanth, 2011)
Garlic (Allium sativum) is a herbal medicine that has been demonstrated to possess a variety
of antibacterial properties against Gram-negative and Gram-positive bacteria (including
species of Salmonella, Staphylococcus, Streptococcus, Klebsiella). The principal
phytochemicals that exhibit these antibacterial properties are organosulfur compounds that
include allicin, ajoenes, and allyl sulphides. The reactive organosulfur compounds form
disulfide bonds with free sulfhydryl groups of enzymes and compromise the integrity of the
bacterial membrane therefore killing them (Bhatwalkar et al., 2021). Allicin is produced from
the amino acid alliin by enzyme alliinase when garlic is crushed. In a dose-dependent
manner allicin can inhibit the reproduction of both bacteria and fungi or kill cells outright by
interfering with lipid synthesis and RNA production. This also and RNA production. This also
includes antibiotic-resistant strains like methicillin-resistant Staphylococcus aureus (MRSA).
(Borlinghaus et al., 2014)
Although mint has been shown to have an antimicrobial effect (Parham et al., 2020), it is
more often used as a flavouring agent in toothpaste and mouthwashes to get rid of
unpleasant smell and to mask the taste of the other ingredients. (Vranic et al., 2004) Menthol
which is naturally found in peppermint and other mint plants is also used in toothpaste for its
ability to trick your brain into sensing that the food is cold giving a cooling and numbing effect
to toothpaste. (Ghosh, n.d.)
Hypothesis : The control will show no inhibition effect on the bacteria. Garlic will have the
most effective antibacterial properties due to the presence of Allicin and organosulfur
compounds. Mint should in theory have little or no effect on the bacteria used in this
experiment
Variables
Independent variable : substance whose antimicrobial properties are being tested (garlic or
mint)
Dependent variable : zone of inhibition made by substance (area measure in cm2)
, Core Practical 9 - 09/02/22
Control variables
- Same concentration of plant material used
- Size of paper discs
- Volume of bacteria (evenly spread)
- All petri dishes should be incubated at the same temperature (25°C) for the same
amount of time (24 hours)
- Agar jelly used in each case must have consistent, neutral pH as it affects the growth
of bacteria
- Humidity and light intensity also affect the growth of bacteria so must be controlled to
be the same for each plant sample
- Size of paper discs
- Prevent contamination of culture using aseptic techniques and sterile equipment
Control
Have a paper disc soaked in distilled water placed on the petri dish. This will act as a
negative control enabling us to compare the results with the antibiotics and the distilled water
to show that the formation of the inhibition zone is due to antibiotics and not because of the
presence of water. In this case, there should not be a zone of inhibition around the distilled
water as it should not inhibit the growth of bacteria therefore the presence of a clear region
around the other paper discs must be due to the antimicrobial properties of the antibiotics.
Risk Assessment
The microorganisms are a potential biological hazard. Although the cultures should be safe
and we know what it is, they may have become contaminated so we use aseptic techniques
and treat them as if they were dangerous. We use aseptic techniques to prevent the culture
from becoming contaminated and to prevent the lab and ourselves from becoming
contaminated from the culture.
Bunsen Burner Keep the flame low, hair tied back and work carefully around the flame. Wear goggles.
Ethanol is highly Make sure ethanol is kept far away from the bunsen burner. Make sure the room is well
flammable ventilated.
Glassware Keep glassware in the centre of the bench and handle carefully to prevent it falling and
breaking.
Microorganisms - Wear a clean lab coat or overall to ourselves and things around from microbes on
the skin, clothing, etc. Wear eye protection and cover any cuts or broken skin with a
waterproof plaster
- Put nothing in the mouth (such as eating, drinking, licking labels, chewing gum,
pencils or pens) as it could be contaminated with the potentially hazardous bacteria
- Start the operations only when all apparatus and materials are within
immediate reach. Complete all operations as quickly as possible, but without
any hurry. Vessels must be open for the minimum amount of time possible.
- Use aseptic techniques and sterile equipment before and after coming in
contact with microorganisms to prevent contamination. Flaming the neck of the
bottle produces a convection current away from the opening and ensures no
microorganisms enter.
- A jar/beaker of disinfectant should be available at each workstation for disposal
Core practical 9 - investigating the antimicrobial properties of plants including aseptic
techniques for the safe handling of bacteria
Aim : To investigate and compare the antimicrobial properties of garlic and mint on bacterial
growth using agar plates and measuring the zones of inhibition.
Introduction
Dental caries are localised, transmissible infectious processes that end up in the destruction
of hard dental tissue. It results from accumulation of plaque on the surface of the teeth and
microbes due to poor oral hygiene. For example Streptococcus mutans is a main pathogen
of dental caries which has a role in the fermentation of carbohydrates resulting in acid
production and demineralisation of tooth enamel. Therefore, chemical agents with
antimicrobial activity were incorporated into dental products as a method of reducing levels
of oral bacteria. (Prasanth, 2011)
Garlic (Allium sativum) is a herbal medicine that has been demonstrated to possess a variety
of antibacterial properties against Gram-negative and Gram-positive bacteria (including
species of Salmonella, Staphylococcus, Streptococcus, Klebsiella). The principal
phytochemicals that exhibit these antibacterial properties are organosulfur compounds that
include allicin, ajoenes, and allyl sulphides. The reactive organosulfur compounds form
disulfide bonds with free sulfhydryl groups of enzymes and compromise the integrity of the
bacterial membrane therefore killing them (Bhatwalkar et al., 2021). Allicin is produced from
the amino acid alliin by enzyme alliinase when garlic is crushed. In a dose-dependent
manner allicin can inhibit the reproduction of both bacteria and fungi or kill cells outright by
interfering with lipid synthesis and RNA production. This also and RNA production. This also
includes antibiotic-resistant strains like methicillin-resistant Staphylococcus aureus (MRSA).
(Borlinghaus et al., 2014)
Although mint has been shown to have an antimicrobial effect (Parham et al., 2020), it is
more often used as a flavouring agent in toothpaste and mouthwashes to get rid of
unpleasant smell and to mask the taste of the other ingredients. (Vranic et al., 2004) Menthol
which is naturally found in peppermint and other mint plants is also used in toothpaste for its
ability to trick your brain into sensing that the food is cold giving a cooling and numbing effect
to toothpaste. (Ghosh, n.d.)
Hypothesis : The control will show no inhibition effect on the bacteria. Garlic will have the
most effective antibacterial properties due to the presence of Allicin and organosulfur
compounds. Mint should in theory have little or no effect on the bacteria used in this
experiment
Variables
Independent variable : substance whose antimicrobial properties are being tested (garlic or
mint)
Dependent variable : zone of inhibition made by substance (area measure in cm2)
, Core Practical 9 - 09/02/22
Control variables
- Same concentration of plant material used
- Size of paper discs
- Volume of bacteria (evenly spread)
- All petri dishes should be incubated at the same temperature (25°C) for the same
amount of time (24 hours)
- Agar jelly used in each case must have consistent, neutral pH as it affects the growth
of bacteria
- Humidity and light intensity also affect the growth of bacteria so must be controlled to
be the same for each plant sample
- Size of paper discs
- Prevent contamination of culture using aseptic techniques and sterile equipment
Control
Have a paper disc soaked in distilled water placed on the petri dish. This will act as a
negative control enabling us to compare the results with the antibiotics and the distilled water
to show that the formation of the inhibition zone is due to antibiotics and not because of the
presence of water. In this case, there should not be a zone of inhibition around the distilled
water as it should not inhibit the growth of bacteria therefore the presence of a clear region
around the other paper discs must be due to the antimicrobial properties of the antibiotics.
Risk Assessment
The microorganisms are a potential biological hazard. Although the cultures should be safe
and we know what it is, they may have become contaminated so we use aseptic techniques
and treat them as if they were dangerous. We use aseptic techniques to prevent the culture
from becoming contaminated and to prevent the lab and ourselves from becoming
contaminated from the culture.
Bunsen Burner Keep the flame low, hair tied back and work carefully around the flame. Wear goggles.
Ethanol is highly Make sure ethanol is kept far away from the bunsen burner. Make sure the room is well
flammable ventilated.
Glassware Keep glassware in the centre of the bench and handle carefully to prevent it falling and
breaking.
Microorganisms - Wear a clean lab coat or overall to ourselves and things around from microbes on
the skin, clothing, etc. Wear eye protection and cover any cuts or broken skin with a
waterproof plaster
- Put nothing in the mouth (such as eating, drinking, licking labels, chewing gum,
pencils or pens) as it could be contaminated with the potentially hazardous bacteria
- Start the operations only when all apparatus and materials are within
immediate reach. Complete all operations as quickly as possible, but without
any hurry. Vessels must be open for the minimum amount of time possible.
- Use aseptic techniques and sterile equipment before and after coming in
contact with microorganisms to prevent contamination. Flaming the neck of the
bottle produces a convection current away from the opening and ensures no
microorganisms enter.
- A jar/beaker of disinfectant should be available at each workstation for disposal
