APEC SCHOOLS DASMARI​ÑAS

Final Output:
Persea Americana (Avocado) Leaves
as an Alternative for Slowing Down
Growth of Campylobacter jejuni,
Salmonella, Clostridium perfringens
and Staphylococcus aureus

Submitted by:

11PM-1 Group 1

Acosta, Rhayza Marie

Faminia, Keneth Adrian

Monton, Maridel

Pulga, Jazmin Shamaya

Realizan, Gemma Rose

Salvador, Jesus Miguel

Submitted to:

Ms. Rona Mae Artuz

December 05, 2019

 Problem Statement & Hypothesis

Persea Americana​ has long been used medicinally, with most parts of the plant
being employed(Chevallier. A.,1996). Research shows that there are anti-cancerous
activity and antihypertensive activity present in parts of ​P. Americana. ​It was also
proved that the leaves, bark, and fruit of ​P. Americana ​can be used to treat dysentery,
cough,​ ​lowering blood pressure, treating liver obstructions, promoting menstrual flow
and for clearing high uric acid levels in the body which could lead to gout(Chevallier. A.,
1996). The leaves of ​P. Americana ​contains flavanols, phenol, and tannins which are
very effective for inhibiting the growth of bacteria(Cowan, MM., 1999).

The present study aims to test the antibacterial activity present in the ​P.
Americana ​leaf extract and if it can be applied as an alternative for antibacterial soap.
The purpose of this is to know if the ​P. Americana ​leaf extract can be used as an
alternative for antibacterial soap which will benefit the community as there will be an
antibacterial agent at the reach of their hands and it also aims to know if the alternative
can be used as a form of business.

The researchers hypothesize that the presence of flavanols, phenol, and

tannins(Cowan,MM., 1999) will have a restrictive effect on the growth of Campylobacter
jejuni, Salmonella, Clostridium perfringens and Staphylococcus aureus(Rouger, A.,
Tresse, O., Zagorec, M. Aug., 2017).​The proposed antibacterial mechanisms of
flavonoids are as follows: inhibition of nucleic acid synthesis, inhibition of cytoplasmic
membrane function, inhibition of energy metabolism, inhibition of the attachment and
biofilm formation, inhibition of the porin on the cell membrane, alteration of the
membrane permeability, and attenuation of the
pathogenicity.(Xie,Yang,Tang,Chen,Ren. (2015)). ​Phenol will act on microorganisms in
2 totally different ways: growth inhibition (bacteriostasis, fungistasis) or fatal action
(bactericidal, agent or viricidal effects).(Sabbineni, J. (Dec, 2016))Tannic acid has a
much greater relative binding efficiency to iron than gallic acid. Tannic acid may work
like a siderophore to chelate iron from the medium and make iron unavailable to
microorganisms. Microorganisms growing under aerobic conditions need iron for a
variety of functions, including reduction of the ribonucleotide precursor of DNA,
formation of haem, and other essential purposes.17 Chung et al.17 reported that the
inhibitory effect of tannic acid on the growth of intestinal bacteria may be caused by its
strong iron-binding capacity. Chung et al (Hisanori et al, (Oct, 2001))
 Methodology

Materials

100 drops of P. Americana leaf extract

5 cups of Glycerin Soap Base
3.5 cups of water
¼ cup of P. Americana Leaf Extract

Independent Variable

- Persea americana (avocado) leaves soap

Dependent Variable

- Presence of bacteria colonies

Controlled Variable

- Size of petri dish, jar, amount of agar powder, amount of persea americana leaf
extract, blender, size of cloth, funnel, cotton buds, size of jar, temperature, and
amount of observation time.
 Results

The first attempt of the researchers to conduct the experiment ended in

failure. The basis for their procedure was incorrect at some point that is why it ended in
their failure. The first procedure indicated that we mix the agar powder and the chicken
broth to cultivate the bacteria that we needed. The said procedure resulted in the agar
powder not mixing well with chicken broth, although some of the mixtures were
successfully formed, the successful mixture showed no sign of bacterial activity. That is
why the researchers deemed the first series of experiment to be a failure. The
researchers review their resources and conduct another study to look for another
procedure or method that is more accurate than what they already had. They found and
validated another procedure and conducted another series of experiment according to
the said procedure.

The second procedure indicated that we should dissolve the agar powder first in
water and heat it through microwaving, which bring more desirable results, or by heating
using stove and fan. Unfortunately, the researchers have no access to a microwave so
they opted for the latter option. They dissolve the agar powder in water and heat it using
a stove and a pan. The results were good as they manage to produce a gelatinous
substance which they transferred into the petri dishes, however the gelatinous
substance quickly cools down and hardened. The researchers tried to melt it down
again by bringing it to fire however the results were undesirable as the gelatinous
substance was affected in color and consistency. The researchers proceeded to
conduct the experiments with what they have. They have conducted three trials in total
where there are two petri dishes per trial: one petri dish for the control where they let the
bacteria grow unhindered and the other is for the soap where they test the effectivity of
the soap by dropping a small amount at the center.

After 4 hours, the researchers observed the experiment to see the change that
occurs within the petri dishes. There was no visible change that occured within the petri
dishes that is labeled control, however in the petri dishes that is labeled soap, zone-like
areas formed at the center where we dropped the soap. After 8 hours, there were no
other visible changes that occured. After 12 hours, the researchers noticed that there
was a little change in the zone-like area, it seems to grow a little bit. After 16 hours,
there was no visible change. After 20 hours, the chicken broth that the researchers put
have reappeared and seems to be showing bacterial activity but there was no visible
formation of bacteria colony. After 24 hours, there was no visible change that occured.
The researchers continued to observe for another day and there was no visible change
that occurred.
 Analysis

The first series of experiment ended in failure due to the lack of knowledge of the
researchers about cultivating bacteria. It may also be due to the misconception of the
researchers since they didn’t have prior knowledge about cultivating bacteria. It can also
be that the procedure that they research is erred, that is why the experiment failed.

The lack of bacterial activity in the second series of experiment cannot be

attributed to the same reason as the former experiment. The procedure for the
experiment has been validated with numerous sources and were presented with actual
video. The lack of bacterial activity on the second series of experiments may be due to
the following reasons: the bacteria that we introduced to the nutrient agar are not
compatible with it, the way how the researchers did the procedure may have altered the
desire output of the nutrient agar, perhaps there was no bacteria at all that was
introduced to the nutrient agar, or the observation time might not be enough.

We have included the possibility that the bacteria may not be compatible with the
nutrient agar because according to research most bacteria will grow well using nutrient
agar, but some more fastidious bacteria (those with more complex nutrient requirements
like Bacillus stearothermophilus, Branhamella catarrhalis, and Bacillus coagulans)
prefer tryptic soy agar. That is why the type of bacteria that we introduced to the nutrient
agar may be the factor why there is no bacterial activity within the petri dishes.

The researchers steps in following the procedure may also be the factor why
there is no bacterial activity within the petri dishes. Perhaps the ratio of the water to the
nutrient agar was not right that may have resulted in a failed product that’s why it cannot
support the growth of the bacteria. It can also be that the researchers over boil the agar
powder and causes it to lose the nutrients to support the growth of the bacteria.
Whether the former or the latter, they will cause the nutrient agar to be incapable of
supporting the growth of the bacteria.

No bacteria was introduced to the nutrient agar was taken into consideration as
well. The chicken broth that we used came from a chicken that was boiled for 30
minutes, the researchers suspected that the bacteria has long been gone and was not
transferred to the chicken broth. If this is the case, then there is no really bacteria that
was introduced to the nutrient agar which resulted to the lack of bacterial activity within
the petri dishes.

Lastly, the cause of the lack of bacterial activity within the petri dishes can be due
to the limited time that the researchers have to observe. According to research, different
bacteria needed different time to grow due to different environment that they were
subjected to. The researchers suspected that the bacteria that was introduced to the
nutrient agar needed a lot of time for it to reproduce on a scale that was visible to the
naked eye, however this is very unlikely because even though it may need a lot of time
to be visible to the naked eye there should be some visible changes that should have
taken place.
 Conclusion

As there is not enough evidence to prove the researcher’s hypothesis, the

scientific study should not concluded. The researchers would need to conduct a series
of experiments to further assess the antibacterial activity of the ​P. Americana leaf
extract.
 Recommendation

As the researchers were limited by their knowledge and experience, the

researchers failed to assess the antibacterial activity present in the leaf extract of ​P.
Americana.​ The researchers were hindered due to having no prior knowledge about
cultivating bacteria, they suggest that future researchers that will conduct similar study
should first research about the cultivation of bacteria. They should first run tests that will
ensure that there will be no mishaps when cultivating bacteria. The researchers are also
shackled by the lack of devices and materials, they suggest that prepare the materials
and add some extra as a contingency plan. They also suggest using microwave in
preparing the nutrient agar. The biggest hindrance that the researchers encountered is
the lack of time to run further experiments and observe the experiments. This prevent
the researchers in further assessing the experiment.
 References
Cowan M. M. (1999). Plant products as antimicrobial agents. Clinical microbiology reviews,
12(4), 564–582. ​https://www.ncbi.nlm.nih.gov/pmc/articles/PMC88925/

Hisanori Akiyama, Kazuyasu Fujii, Osamu Yamasaki, Takashi Oono, Keiji Iwatsuki,
Antibacterial action of several tannins against Staphylococcus aureus, Journal of
Antimicrobial Chemotherapy, Volume 48, Issue 4, October 2001, Pages 487–491,
https://doi.org/10.1093/jac/48.4.487

Joshita Sabbineni, Phenol-An effective antibacterial Agent, Research & Reviews:

Journal of Medicinal & Organic Chemistry,Volume 3, Issue 2, December, 2016
http://www.rroij.com/open-access/phenol-an-effective-antibacterial-agent-.pdf

J. HOWARD BROWN, PH.D. Feb., 1947. Retrieved from

https://ajph.aphapublications.org/doi/pdfplus/10.2105/AJPH.37.2.206

Rouger, A., Tresse, O., Zagorec, M. Aug., 2017. Retrieved from

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5620641/

Okorie, F. Jan., 2019. Retrieved from

https://www.legit.ng/1215747-health-benefits-avocado-leaves.html