Table 2. Zones of inhibition against S. aureus and E. coli by plant extract. Data in mm.

Against S. aureus Against E. coliDandelion MeOH EtOH Ace H2O Dandelion MeOH EtOH Ace H2O

flower 0 0 0 0 flower 0 0 0 0

leaf 11 0 11 0 leaf 11 0 11 0

stem 0 0 0 0 stem 0 0 0 0Plumeless thistle MeOH EtOH Ace H2O

Plumelessthistle MeOH EtOH Ace H2O

flower 0 0 0 0 flower 0 0 0 0

leaf 0 0 11 0 leaf 0 0 11 0

stem 0 0 0 0 stem 0 0 0 0Indianteasel MeOH EtOH Ace H2O

California buttercup MeOH EtOH Ace H2O

flower 0 0 11 0 flower 0 13 0 0

leaf 0 0 0 0 leaf 0 0 0 0

stem 0 0 0 0 stem 0 0 0 0

Jonathan Tolentino and Claudia BrionesBiology Department, Skyline College, San Bruno CA

Acknowledgements• Dr. Christine Case, Professor of Biology, Skyline College.• Tiffany Reardon, Assistant Director, California MESA.• Patricia Carter, Biology Technician, Skyline College.• Marc Anderson, Professor of Chemistry, San Francisco State University• Ulla Andersen, Chemistry Mass Spectrometry Facility, UC Berkeley• Funded by NIH/SFSU Bridges to the Baccalaureate.

Table 1. Native American Uses of Plants Tested

Passiflora incarnata

Passionflower CherokeeWoundsRef: 3

Taraxacum officinale

Common DandelionBella CoolaStomachacheRef: 6

Dipsacus sativus

Indian Teasel IroquoisAcneRef: 4

C. pycnocephalus

Plumeless Thistle OjibwaBowel tonicRef: 5

Sonchus oleraceus

Common Sowthistle HoumaAnti-diarrhealRef: 7

R. californicus

California Buttercup MiwokFood SupplementRef: 1

Materials & MethodsPlant extracts: 1. Plants (Table 1) were gathered and separated by flowers, stems, and

leaves.2. A food processor & mortar and pestle were used to grind plant parts to

be mixed with solvents.3. Methanolic, ethanolic, acetone, and aqueous extracts were made in

concentrations at a minimum of 1 mL/g. Disk diffusion assay:

Filter paper disks immersed in an extract or solvent (control) were applied to nutrient agar plates inoculated with the test bacteria: Staphylococcus aureus or Escherichia coli and incubated for 24 hr at 35°C.

Minimal Inhibitory Concentration (MIC) and Minimal Bactericidal Concentration (MBC):

1. Microdilutions of plant extracts (500 mg/mL to 15 mg/mL) were made in nutrient broth, inoculated with the test bacteria: S. aureus or E. coli, and incubated for 24 hr at 35°C

2. Dilutions showing no growth were subcultured in nutrient broth to determine the MBC.

Paper Chromatography Paper chromatography was performed on the extracts, which determined Rf values to help identify active compounds. Isopropanol was used as solvent, and sections of strips were assayed against test bacteria to locate antibacterial activity.

High performance Liquid Chromatography HPLC separated the crude extract samples into fractions, which were concentrated using high pressure vacuum. Fractions were assayed by the disk-diffusion method.

Nuclear Magnetic Resonance Spectrometry NMR data were gathered from active fractions to help determine chemical structure of the active compound.

Mass Spectrometry Mass spectrometry was performed to determine elemental composition of active samples.

Background• Indigenous cultures throughout the world use herbal medicine to cure

illness. • Many invasive plants are regarded as pests, but certain species are

valued in Native American folklore for healing properties (2). • Such a source of alternative medicine can provide an abundant supply

that is readily available, and potentially contribute to managing the health of the ecosystem.

• Traditional uses of the medicinal plants we tested are shown in Table 1.

AimEvaluate antimicrobial activity and identify active compounds in plants traditionally used to treat infections

AbstractDue to increasing numbers of bacteria strains resistant to antibiotics, there is a need for effective alternative sources of antimicrobial agents. Native American folklore recognizes various common plants for medicinal purposes, but their efficacy has not been tested. Several Native American plants were hypothesized to have antibacterial properties. Passiflora incarnata, Taraxacum officinale, Dipsacus sativus, Carduus pycnocephalus, Sonchus oleraceus, and Ranunculus californicus were evaluated for their antibacterial activity against gram-positive Staphylococcus aureus and gram-negative Escherichia coli bacteria.

Discussion & Conclusions•The T. officinale leaf methanolic and acetone extracts of inhibited both

gram-positive and gram-negative bacteria (MIC=0.25 g/mL).•The C. pycnocephalus leaf-acetone extract inhibited gram-positive S.

aureus (MIC=0.25 g/mL) and gram-negative E. coli (MIC= 0.5 g/mL). •The D. sativus flower-acetone extract was bactericidal against gram-

positive bacteria (MBC=4.38 g/mL). •The R. californicus flower-ethanolic extract was bactericidal against

gram-negative bacteria (MBC=26 mg/mL). •Results confirm that these plants have antimicrobial properties.

Exploring folkloric herbal plants may provide rewarding natural resources for medicine. Continued testing should determine whether these plants can be processed into potential drugs to control certain infectious diseases.

Literature Cited1. Barrett, S. A. and E. W. Gifford. 1933. Miwok Material Culture. Bulletin of the Public

Museum of the City of Milwaukee 2(4):11. 2. Cowan, M. M. 1999. “Plant Products as Antimicrobial Agents.” Clinical Microbiology Review

12(4): 564–582.3. Hamel, P. B. and M. U. Chiltoskey. 1975. Cherokee Plants and Their Uses—A 400 Year

History. Sylva, N.C. Herald Publishing Co.4. Herrick, J. W. 1977. Iroquois Medical Botany. State University of New York, Albany, Ph.D.

Thesis. 5. Smith, H. H. 1932. Ethnobotany of the Ojibwe Indians. Bulletin of the Public Museum of

Milwaukee 4:327-525. 6. Smith, H. I. 1929. Materia Medica of the Bella Coola and Neighboring Tribes of British

Columbia. National Museum of Canada Bulletin 56:47-68. 7. Speck, F. G. 1941. “A List of Plant Curatives Obtained From the Houma Indians of

Louisiana.” Primitive Man 14:49-75.

Results• None of the aqueous extracts inhibit the test bacteria. Methanolic and

acetone extracts of T. officinale leaves and the acetone extract of C. pycnocephalus leaves inhibit both S. aureus and E. coli with zones of inhibition at 11 mm. Ethanolic extracts of R. californicus flowers and acetone extracts of D. sativus flowers inhibit E. coli (Table 2).

• The MIC of the methanolic and acetone extracts of T. officinale leaves against both bacteria is 0.25 g/mL. The MIC of C. pycnocephalus leaf in acetone against S. aureus is 0.25 g/mL, and 0.5 g/mL against E. coli (Figure 1).

• The MBC of the acetone extract of D. sativus flower against S. aureus is 4.50 g/mL. MBC of the ethanolic extract of R. californicus flower against E. coli is 15 mg/mL.

• Paper chromatographic segments of D. sativus flower extract and T. officinale leaf extract produced zones of inhibition against S. aureus (Figure 2).

• One HPLC fraction from D. sativus flower acetone extract (Figure 3) and another from T. officinale acetone leaf inhibited S. aureus. No other fractions showed inhibition from the two plants.

• NMR data from acetone extracts of D. sativus flower and T. officinale leaf indicate one or more compounds in each extract. A second HPLC analysis of the compounds determined that they are fairly pure, but are still unidentified.

Figure 1. Inhibitory concentrations of plant extracts against (a) S. aureus and (b) E. coli.

a. Inhibition of S. aureus. b. Inhibition of E. coli.

Figure 2. Paper chromatograms were cut to use in a disk diffusion assay. Rf =0.75 of the acetone extract of D. sativus flower inhibited growth of S. aureus (at arrow). Rf =0.05 of the acetone extract of dandelion leaf inhibited growth of S. aureus (not shown).

ANTIBACTERIAL ACTIVITY OF HERBAL EXTRACTS USED IN NATIVE AMERICAN TRADITIONAL MEDICINEANTIBACTERIAL ACTIVITY OF HERBAL EXTRACTS

USED IN NATIVE AMERICAN TRADITIONAL MEDICINE

Figure 3. One HPLC fraction from D. sativus flower acetone extract inhibited S. aureus. No other fractions produced inhibition.

Figure 6. Paper chromatograms were cut to use in a disk diffusion assay.

(a) Rf =0.05 of the acetone extract of dandelion leaf inhibited growth of S. aureus.

(a) Rf =0.75 of the acetone extract of Indian teasel flower inhibited growth of S. aureus.

Indian teasel flower fraction A Vs. S. aureus

Dandelion leaf fraction A vs. S. aureus

NMR data from acetone extracts of D. sativus flower and T. officinale leaf indicate one or more compounds in each extract. A second HPLC analysis of the compounds determined that they are fairly pure, but are still unidentified.

Figure 2. Bactericidal concentrations of plant extracts.