Laboratory Investigation: Antimicrobial Action of Silver Nanoparticles
Purpose
Silver nanoparticles range from between 1nm to 100 nm in diameter. Silver
nanoparticles have been used in optics, electronics and for catalyzing chemical
reactions, but recently they have become popular because of their ability to inhibit
bacterial and fungal growth. Because an increasing number of microbes have
demonstrated resistance to traditional antibiotics, many scientists view silver
nanoparticles as an alternative approach for controlling the growth of microbes.
In this laboratory you will measure the effectiveness of silver colloid in controlling
bacterial growth.
Background information
Silver dollars were once placed in milk bottles to keep the milk fresh. Water tanks in
ships and planes were coated with silver to prevent bacterial contamination.
Although all metals have antimicrobial properties, silver was found to have the
greatest ability to control microbial growth without demonstrating damaging effects
on human cells.
Although the antimicrobial properties of silver have long been known, scientists
have only recently begun to understand the mechanism. Silver nanoparticles bind to
thiol (-SH) groups in microbial enzymes, preventing the enzymes from catalyzing
intended biochemical reactions. Silver binds to the enzyme found in cell membranes
that is involved in providing energy for the active transport of molecules across the
membrane.
Bandage fiber
Silver
nanoparticles
Bandage with silver nanoparticles 1500 X. Image taken with a scanning electron
microscope.
Problem
Can silver nanoparticles inhibit bacterial growth?
Compare the effectiveness of silver nanoparticles with traditional antibiotics for
inhibiting the growth of common bacteria.
Hypothesis
1. Create a hypothesis for your investigation.
Prediction
2. Make a prediction for your investigation.
Design
The microbes (bacteria and fungi) grow readily on nutrient agar in a Petri dish. Agar
is a gelatin-like substance obtained from red alga. It is a sugar compound that
adheres to the cell walls of the alga and serves to bind cell walls together.
In this experiment microbes are transferred from a desk surface to a Petri dish
containing agar that contains nutrients for microbial growth. The microbes
introduced to the nutrient begin to grow quickly and eventually, take up most of the
agar surface. The microbes can be seen with as a film over the agar and are visible
without a microscope. Should an antibacterial agent be introduced into the Petri
dish, growth near the agent is inhibited. A “kill zone” or clear area is seen around to
the antimicrobial agent. The larger the kill zone, the more effective is the agent.
Caution
Most microbes collected in the environment will not be harmful. However, once they
multiply into millions of colonies in a petri dish they become more of a hazard. Be
sure to protect open cuts with rubber gloves and never ingest or breathe in growing
microbes. Keep the inoculated petri dishes taped closed until your experiment is
done. Then you should safely destroy the fuzzy microbial growth using bleach.
Materials
Petri dish with nutrient agar
Graduated cylinder
Sterile cotton swabs
Antibiotic disks
Ruler (mm)
Filter paper and hole punch
10% bleach solution
Masking tape
Felt marker
*Silver colloid solution
Incubator
Latex Gloves
Plastic bag for disposal.
Forceps
* Silver colloid is readily available from most health food stores.
Procedure
1.
Put on the latex gloves and wear them for the entire experiment. Using a
felt marker, divide the bottom of a Petri dish into three sections and
number each section.
1
2
3
2.
Removes the sterile swab from the case, Moisten it with a drop of distilled
water and run it along your forearm or a desk to collect microbes that are
normally present in the environment. Now remove the upper lid of the
Petri dish and gently run the swab back and forth across the surface of
the agar several times to cover all areas of the Petri dish. Do not pierce
the agar.
3.
Place two different antibiotic disks in the centre of sections #1 and #2.
Using a forceps touch the disks to have them stick to the surface of the
agar. Now using a forceps, place a disk of filter paper onto the centre of
section #3 and add three drops of colloid silver.
4.
Invert the Petri dishes so that the small disk
containing the agar is on top and then place the
Petri dishes for 48 h in the incubator and set the
temperature to 37 oC. If you don’t have an incubator, place the disks in a
warm cupboard for three or four days.
5.
Wipe down your lab table with a small amount of bleach solution and
remove your gloves and place them into a plastic bag.
Evidence
6.
After 24h and 48 h measure the diameter of the clear zone (or kill zone)
around each of the disks.
3. Construct a data table to show results for 2 additional days.
4. Provide data in a graph. Consider the type of graph you will choose
to display your data.
7.
Seal the Petri dishes with masking tape and place them in a plastic bag for
disposal. Never open a Petri dish. Bleach will be used to destroy bacteria.
Analysis
5. Which antibacterial agent proved best at inhibiting growth of the
colonies?
6. Compare your prediction to your final data.
Evaluation
7. How would you go about designing a control for this experiment?
8. What limitations would you place upon your conclusion?
Synthesis
9. What factors might affect the ability of silver nanoparticles to
inhibit bacterial growth?
10. Provide three potential applications for using silver nanoparticles
to control bacteria growth.
11. Identify and explain a concern or potential negative consequence
for using the silver nanoparticles for each of your suggested
applications.