Instructional Method:
The instructor gives a presentation on new products using nanoparticles and bacteria culturing techniques. Students then do an activity in which they make nanoparticles and culture bacteria in the presence of those particles. The students draw conclusions about the effectiveness of the nanoparticles as an antimicrobial agent using proper experimental design.
Materials:
Preparation:
Procure sterile bacterial media. Culture bacteria 1-2 days before activity. Get the PowerPoint presentation or overheads ready. Make a hot water bath using 100 mL beakers half filled with boiling distilled water. Set out the solutions and equipment.
Background Information:
Nanotechnology is an emerging industry which is bringing us exciting new products and promises to change the way we live and work in the future. Several new products are using silver nanoparticles to generate antimicrobial surfaces. Silver nanoparticles are integrated into fabrics to prevent clothes from developing foul odors, doorknobs have silver nanoparticles embedded in their surfaces, even silver nanoparticle treated pacifiers are on the market. For a list of hundreds of nanotechnology products using silver nanoparticles see http://nanotechproject.org/44. With hundreds of new products claiming antimicrobial properties we thought appropriate to put silver nanoparticles to the test.
Nanotechnology is a new field of engineering which will change the world during your lifetime. Progress in the field of miniaturization allows scientists to now think about making products smaller than a micron (1 millionth of a meter). This means these products can be measured in nanometers (1 billionth of a meter).
How can we hope to be able to tell the sizes of particles that are so small? Actually, the size of dispersed particles can be deduced by the physical properties of the mixtures. By experimenting on our mixture, we will be able to draw some conclusions about the sizes of the nanoparticles. We will be looking at the properties of settling, Tyndall effect, and filtering.
If a mixture of a solid and a liquid has large solid particles, gravity will cause the particles to settle out as we see in the top picture. If you have colloidal (1-1000 nm) dispersed particles, collisions with the dispersing medium’s particles will help the particles stay mixed. This may result in a colored and/or cloudy mixture. A solution in which the dissolved particles are either ions or small molecules ( Note the difference between the terms clear and colorless.
Dissolved particles are too small to be scattered by light, however colloidal (nanoparticles) with sizes near the wavelength of light, will interact with visible photons. When a laser or other beam of light is shown through a colloid, the light will be scattered and the beam will be visible in the mixture.
In summary, solutions have particles less than 1 nm, are clear, show no Tyndall effect and remain dissolved. The nanoparticles in colloids are larger than those in solutions (1-1000 nm) and will scatter light (Tyndall effect). However the particles are small enough to remain dispersed without settling out. A colloid will typically look cloudy. Suspensions have the largest particles (>1000 nm), will settle out over time, and will scatter light while temporarily suspended.
Adding salt to the gold nanoparticle mixture will destablized the colloid causing coagulation. This should be seen as a color change and will also change the filtering characteristics of the mixture.
What would it take to see these particles under a microscope? The light microscopes used in biology class would not have nearly enough magnification.
A scanning electron microscope is a tool used by research scientists to “see” things on a very small scale. An SEM will detect at the nanometer level, small enough to see nanoparticles.
Here is an SEM image of colloidal gold on the filter surface. The holes in the filter are clearly visible in the background.
What are the potential applications for gold or silver nanoparticles? Electrical engineers are interested in them for their electrical properties. One exciting area is the idea of using nanoparticles as a delivery mechanism for drugs. Gold nanoparticles are currently manufactured for targeted delivery of biomolecules and drugs to selected cells. Trials are underway using this method to treat cancer cells in mice. The drug appears to accumulate in the tumor but not in the healthy cells.
Instructor Information for Antimicrobial Silver Nanoparticles Lab Activity
Overview: In this lab, students will make a silver colloidal mixture. Silver solution will be reduced and stabilized with sodium citrate in a boiling water bath. The colloidal dispersions formed will be poured into Petri dishes, soaked up into filters, and placed on top of a bacterial agar plate. The silver colloid should be yellow in color. Silver colloidal particles are 20-50 nm in size. If the silver nanoparticles prevent bacterial growth, there should be a ring on inhibition around where the filters soaked with nanoparticles are placed on the bacterial agar plate.
Background:
Nanotechnology is an emerging industry with many potential applications. Scientists are very interested in nanoparticles as it pertains to nanotechnology. Nanoparticles are often defined as having dispersed particles in the size range 1-100 nm. Gold nanoparticles are finding applications in cancer treatment, and silver nanoparticles are found to have antimicrobial properties.
Though there is a serious lack of information to describe the mechanism in which the silver nanoparticles actually prevent bacterial growth, most research points to interactions with the bacterial cell wall, and regulation of materials across the membrane. This could also be why there are different results in gram positive and gram negative bacterial strains. E. coli is a common gram negative bacteria used in microbiology, and is safely cultured and maintained. For quick results E. coli should grow overnight if incubated at 37°C.
Preparation:
You will need to set up the following:
You will need to prepare stock solutions of:
Safety:
Goggles and aprons should be worn as in all chemistry laboratory activities. The hot water baths should be handled with care to avoid burns. Any liquids spilled on skin can be washed off with water.
This lab uses Escherichia coli. E. coli is a gram-negative rod-shaped bacterium that is part of the normal intestinal fauna in mammals. Some strains, particularly O157:H7, are pathogenic to humans; most strains, however, are benign. Because of the ease of culturing E. coli, it has become the “workhorse” of microbiology. Strains for use in laboratories and classrooms are derived from E. coli that grows very well on Petri dishes but very poorly in intestines. The E. coli used in this lab is non-pathogenic and likely wouldn’t live in a human intestine even if large amounts of the bacteria were ingested because it couldn’t out-compete the naturally occurring bacterial fauna, including benign forms of E. coli, already present in the gut.
References:
3. Sondi, Ivan; Salopek-Sondi, Branka. Journal of Colloid and Interface Science. 2004, 275
177-182.
4. Song, H.Y; Ko, K.K; Oh, I.H; Lee, B.T; European cells and Materials. 2006, 11, 58.
5. Cushing, Brian L; Kolesnichenko, Vladimir L; O’Connor, Charles J. Chem. Rev. 2004,
104, 3893-3946.
6. Pal, Sukdeb; Tak, Yu Kyung; Song, Joon Myong. Applied Environmental Microbiology. 2007, 73, 1712-1720.
Useful Website:
National Nanotechnology Initiative
http://www.nano.gov
Filtering Nanoparticles Student Lab Activity
Overview: This lab was developed to test claims that colloidal silver has anti-microbial effects. Many products are on the market that have nanoparticles of silver in them. These include socks, clothing and food containers. One of their advertising claims is that the silver nanoparticles inhibit the growth of bacteria and hence reduce odors and/or spoilage.
Background: Nanotechnology is an emerging industry with many potential applications. Scientists are very interested in nanoparticles as it pertains to nanotechnology. Nanoparticles are often defined as having dispersed particles in the size range 1-100 nm. Gold nanoparticles are finding applications in cancer treatment, and silver nanoparticles are found to have antimicrobial properties.
Though there is a serious lack of information to describe the mechanism in which the silver nanoparticles actually prevent bacterial growth, most research points to interactions with the bacterial cell wall, and regulation of materials across the membrane. This could also be why there are different results in gram positive and gram negative bacterial strains. E. coli is a common gram negative bacteria used in microbiology, and is safely cultured and maintained. For quick results E. coli should grow overnight if incubated at 37°C.
Safety: Goggles and aprons should be worn as in all chemistry laboratory activities. The hot water baths
should be handled with care to avoid burns. Any liquids spilled on skin can be washed off with water.
Materials:
1 mM silver nitrate
38.8 mM (1%) sodium citrate
Hot plate
Small test tubes
250 mL beaker
Disposable transfer pipets
Agar plates
Scissors
Experimental Procedure:
11. use a 1 mL disposable transfer pipet and spread 1-2 drops of the bacteria culture on the agar plate using a Q-tip swab.
12. After 10 minutes of soaking, place filter paper squares in the designated areas.
13. The teacher then incubates petri plates for 24 hours at 37 degrees Celsius.
14. On the second day of this activity, the petri plates should be removed from the incubator and examined, student results discussed and evaluated through classroom discussion.
For more information, contact: Center for Nanoscale Chemical-Electrical-Mechanical Manufacturing Systems; University of Illinois at Urbana-Champaign, 4400 Mechanical Engineering Laboratory, 105 South Mathews Avenue, MC-244, Urbana, IL 61801
Phone: 217.265.0093 Email: nano-cemms@uiuc.edu Website: http://www.nano-cemms.uiuc.edu
