Transcript C. pycnocephalus

Evaluation of Native American Herbal Extracts
As Food Preservatives
Jonathan Tolentino
Biology Department, Skyline College, San Bruno CA
Abstract
Results
Foodborne illness is an important public health problem
worldwide. Risks with continued use of chemicals as food
preservatives have prompted a search for safer and more natural
alternatives. Several California native plants exhibiting
antibacterial properties were hypothesized to help prevent bacterial
spoilage caused by Escherichia coli. Taraxacum officinale,
Dipsacus sativus, Carduus pycnocephalus, and Ranunculus
californicus extracts were evaluated for their ability to prevent E.
coli growth on lettuce and ground beef kept at 25°C. Ethanolic and
acetone extracts were evaporated and rehydrated with sterile
distilled water before being assayed. Bacterial growth was
determined using dilution and plate counting.
Bacterial growth in lettuce with extracts was not significantly
different compared to the control over the seven day observation.
Bacterial growth was too many to count for all meat trials with
extracts and the control. Results demonstrated that extracts are not
effective against E. coli food spoilage as prepared. Future studies
of the extracts may involve steps to increase its effectiveness.
Effective extracts can provide a source of much needed food
preservatives and antibacterial washes.
• The extracts are not effective as prepared. E. coli was able to
grow in lettuce and ground beef treated with extracts (Figure 2).
Evaluate the effectiveness of selected Native American plants
against food spoilage bacteria and foodborne pathogens in vitro.
Background
• Food contamination results in 76 million foodborne diseases in
the U. S. annually, and costs the U. S. economy $10-$83 billion
in 2008 (3).
• The rise of foodborne illnesses and concern about chemicals as
food preservatives has created interest in finding safe microbial
inhibitors.
• Propolis, a resinous mixture collected by bees, reduces bacteria
in oriental sausage (1). Rosemary extract and grapefruit seed
extract inhibit bacteria in marinated pork (2).
• Native American plants used in traditional medicine have been
investigated for antimicrobial properties. Taraxacum officinale,
Dipsacus sativus, Carduus pycnocephalus, and Ranunculus
californicus extracts inhibit growth of E. coli (4).
• Meat assays with extracts and control had too much bacterial
growth to count (Figure 3).
Carduus pycnocephalus
Italian thistle
Taraxacum officinale
Common dandelion
Dipsacus sativus
Indian teasel
Ranunculus californicus
California buttercup
Figure 1. Plants used in food preservative assays.
Materials & Methods
Extract preparation
1. Flowers of R. californicus and D. sativus and leaves of T.
officinale and C. pycnocephalus (Figure 1) were gathered and
weighed. Stems and midveins were removed.
2. Plant parts were ground with mortar and pestle.
163 mg/mL ethanolic extracts were made from R. californicus
flowers, and 500 mg/mL acetone extracts from T. officinale, C.
pycnocephalus leaves and D. sativus flowers. The two solvents
used correspond to previous assays that confirmed antimicrobial
activity of extracts (4).
3. Extracts were air-evaporated for 5 days at 35°C then rehydrated to
the original volume with sterile distilled water, and stored at 5°C.
Bacterial growth in food
1. 3 g of lettuce were cut and placed in sterile Petri dishes.
2. Samples were inoculated with 100 E. coli cells.
3. 10 ml extract (500 mg/mL T. officinale, D. sativus and C.
pycnocephalus extracts, 163 mg/mL R. californicus extract) was
added to test plates. 10 mL sterile water was added to controls. 1
mL sterile water was added each day to compensate for volume
lost due to sampling. Plates were incubated at 25°C for 7 days.
4. Samples were taken after 72 and 168 hrs. and bacterial growth was
determined by plate counts.
5. Steps 2 through 4 were repeated using 10 g ground beef per assay.
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Discussion & Conclusions
• The decrease in volume of the R.californicus extract after
evaporation was minimal, suggesting that the ethanol did not
completely evaporate after five days at 35°C. The volume of T.
officinale. C. pycnocephalus and D. sativus extracts after
evaporation was approximately equal to their initial rehydration
with sterile water, suggesting that the acetone was removed from
these extracts.
8
Log Bacterial Count
Aim
• The 5-day evaporation at 35°C was neither long nor hot enough
to completely remove the ethanol in the R.californicus extract.
Inhibition of E. coli growth in lettuce with R.californicus extract
is attributed to residual alcohol.
• Future attempts to test the extracts as a food preservative may
involve increasing concentration of the extracts by prolonging
evaporation and using more plant per volume.
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6
5
4
3
Control
2
1
Literature Cited
T. officinale
0 S.E.
C. pycnocephalus
0
0
50
100
150
200
Time (hours)
Figure 2. Bacterial growth over 7 days in lettuce treated with extracts.
R. californicus results were inconclusive, and therefore omitted.
1. Ali, F. H. et al. 2010. “Propolis as a natural decontaminant and
antioxidant in fresh oriental sausage.” Vet. Ital. 46(2):167-72.
2. Schirmer, B. C. et al. 2010. “Evaluation of natural
antimicrobials on typical meat spoilage bacteria in vitro and in
vacuum packed pork meat.” J. Food Sci. 75(2):M98-M102.
3. Nyachuba, D. G. 2010. “Foodborne illness: is it on the rise?”
Nutr. Rev. 68(5):257-69.
4. Tolentino, J. 2009. “Antimicrobial properties of plants used in
Native American Traditional Medicine.” SACNAS National
Conference Abstracts 2009, 110.
Acknowledgements
Figure 3. Extract assays against E. coli in lettuce. Bacterial inhibition by
R. californicus extract may be due to its residual ethanol. Bacterial
growth in lettuce treated with C. pycnocephalus extract was too many to
count.
Dr. Christine Case, Professor of Biology, Skyline College.
Rhia Bordon, Biology Student, Skyline College.
Tiffany Reardon, Assistant Director, California MESA.
Stephen Fredericks, MESA Director, Skyline College.
Patricia Carter, Biology Technician, Skyline College.