Testing Plant Substances as Potential

Medicines Lab
Background: Plants have been and still are in a constant war for survival. However,
some of their enemies are not other plants, but bacteria and viruses. Plants have
developed their own defense against these enemies, which scientists try to harvest.
One must collect the sample in far-away places, then create procedures for testing all
the individual samples. They create petri dishes with their extract and see if the extract
will kill off an invading microbes.

Objective/Purpose: To find what plant materials, found locally, contain active

ingredients that will inhibit the growth of bacteria.
Using a mortar and pestle, grind up 2 grams of plant tissue with 10 mL of
deionized water.
Let it sit for 3 minutes
Filter the sample through an 11 cm filter funnel
Filter sterilize the extract using a syringe filter
Collect 1 mL of the filter-sterilized extract into a 1.7 microfuge tube
Label the sample
Reconstitute Methanol Filtrate with 1 mL sterile water
Place 3 sterile pieces of filter paper in each filter. (6 total)
Label and pour agar into plate

Plant extracts, syringe/ prefilter, pipet

Label microfuge tube: Water and Methanol
Attach sterile filter to prefilter
Load 1.7 ml of extract into syringe, using pipet
Depress plunger 1.0 ml filter sterilized extract
Snap cap on microfuge tube

Label agar plates with a cross, label quadrants

Using sterile forceps, add the appropriate number of sterile disks to each tube of filtered
extract
Prepare negative control disks:
2 - sterile water

2- Ampicillin
Place the disks in the appropriate solution
Sterile disks were added to microfuge tubes containing one mL sterile water
10-20 mL of warmed nutrient agar was poured into 2 petri dishes using sterile technique
After allowing agar to solidify, plates were turned upside down and stored at 4 degrees
celsius overnight
One mL of Ecoli colony was added to each plate. A flame-sterilized spreading loop was
used to spread the bacteria throughout the surface of the agar
Using flame-sterilized forceps, filter disks were placed in separate quadrants onto the
plate in the following sequence: Water, plant extracts, ampicillin. Plants were left on lab
bench for 20 min. to allow both bacteria and filter filter disks to adhere to the agar
Plants were incubated upside down, overnight at 37 degrees celsius. Plate were
photographed and observed for clearance around the filter disks after 24 hours, 48
hours, and 72 hours

Results: Our results were not as groundbreaking as we had hoped they would be.
Over the course of our two days looking at the agar plates, nothing happened. To start
with our water dish, the ampicillin didnt really do its job. It only had a small ring around
it, not a clearance of the agar. Our plant-water soaked papers only had this small ring as
well, not a true clearance. This maintained for both of the two days (We only had two
days instead of three because we were a day behind). With our methanol plate,