Plant Substances as Potential Medicines Lab - 6d
Purpose: To see if local plants can inhibit bacterial growth.
Materials:
-Balance -LB broth base -250 mL media bottles -sterilizer -laminar flow hood and disinfectant -safety glasses -Bunsen burner/gas lighter -inoculating loop -petri dishes -E. coli JM109 -plant specimen -mortar and pestle |
-10 mL pipet
-plastic funnels -5 mm filter paper disks -100 mL beakers -10 mL syringe -1.7 mL reaction tubes/rack -Methanol -1 mL pipet -heat block -forceps -Ampicilin -Glass spreader -Incubator oven |
Procedure:
Part 2:
1. Grind 2 g plant material with 10 ml of H2O, let sit for 3 minutes
2. Filter through paper lined funnel, then sterilize
3. Repeat, but with Methanol instead of water, afterwards put in heat block for 1 day with lid open
4. Add 1 ml sterile water and then vortex
5. Using sterile forceps, put 2 filter disks in each tube and leave for another day
Part 3:
1. Transfer 1 ml of E. coli into each quadrant of petri dish with a 1 ml pipet
2. Spread with glass spreader sterilized by the flames of purgatory (cause science)
3. Cover, let soak in agar for 15 mins.
4. Place disk filters in designated quadrants using sterilized forceps
5. Incubate at 37 degrees Celsius for 24 hours
Part 2:
1. Grind 2 g plant material with 10 ml of H2O, let sit for 3 minutes
2. Filter through paper lined funnel, then sterilize
3. Repeat, but with Methanol instead of water, afterwards put in heat block for 1 day with lid open
4. Add 1 ml sterile water and then vortex
5. Using sterile forceps, put 2 filter disks in each tube and leave for another day
Part 3:
1. Transfer 1 ml of E. coli into each quadrant of petri dish with a 1 ml pipet
2. Spread with glass spreader sterilized by the flames of purgatory (cause science)
3. Cover, let soak in agar for 15 mins.
4. Place disk filters in designated quadrants using sterilized forceps
5. Incubate at 37 degrees Celsius for 24 hours
Results:
H2O Plant Sample: Positive, ~0.5-1 mm clearing around both filters Methanol Plant Sample: Negative, no clearing around either filter Positive Control: Positive (weakly), ~0.5 mm clearing Negative Control: Negative, no clearing Data Analysis I have found that the plant I tested does indeed have anti-bacterial qualities. My controls did work although quite weakly, this may be due to a dilution in the ampicilin or just a bad batch of ampiicilin. To expand on this experiment we could look into the materials of the plant to try and identify the exact ingredient that causes the anti-bacterial qualities. After this (if we were a biotech firm) we'd probably repeat the experiment a few more times and if it still worked like it did then we'd try to identify the exact molecule that causes this occurrence. |
Thinking Like a Biotechnician Questions:
1. If an extract gives a negative result, it means that the extract does not have antimicrobial agents.
2. If the methanol extract smells of alcohol that means the alcohol will kill the bacteria, not the plant extract.
3. To determine the exact compound that is antimicrobial, you'd have to extract all the compounds in the plant and then test them in a similar method as this lab in order to see which one is the compound that kills bacteria.
1. If an extract gives a negative result, it means that the extract does not have antimicrobial agents.
2. If the methanol extract smells of alcohol that means the alcohol will kill the bacteria, not the plant extract.
3. To determine the exact compound that is antimicrobial, you'd have to extract all the compounds in the plant and then test them in a similar method as this lab in order to see which one is the compound that kills bacteria.