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Screening of Phytochemical and Antibacterial Activity of Sedum Morganianum against

Escherichia coli, Staphylococcus aureus And Pseudomonas aeruginosa

An Investigatory Submitted to the Faculty and Staff of Cordillera Regional Science High School

in Partial Fulfillment of Requirements in Research 12

Jerileen P. Paleng

Rose Celine D. Macliing

Clarizze Jocel B. Saingan

Anjesca Judd P. Reroma

Dresani Dijka D. Deponio

Danio H. Mayos Jr.

Proponents

Amethyst Joy E. Taqued

Research Adviser

July 2019

Over the years, plants have served as sources of large amount of drugs such as

antispasmodics, emetics, anti-cancer, antimicrobials and many more. Many are extensively used

by tribal people worldwide since a large number of the plants are claimed to possess the

antibiotic properties (Tiwari, 2011).


The effectiveness of these plants as medicines lies within their chemical content, also

known as phytochemicals. They are natural components that are biologically active in plants.

The presence of these chemical influence the antibacterial or antifungal properties of a plant.

Examples of phytochemicals are alkaloid, phenolics and flavonoids which can all be used as

medical drugs (Science Learning Hub Organization, 2011). Aside from its medicinal properties,

scientists also reported having extracted higher concentrations/amount of phenolics from plants

as compared to acetone, water, and methanol (Altemimi, Lakhssassi, Baharlouei, Watson &

Lighfoot, 2017).

There are several researches that used Escherichia coli, Staphylococcus aureus, and

Pseudomonas aeruginosa. One of these is an invention carried out by Banso (2008) in which he

observed that the stem bark of Acacia niloticahas has phenolic constituents against five

microorganisms including Escherichia coli and Staphylococcus aureus. Antimicrobial assay

using diffusion method was performed and the components of the plant was tested. Results

showed the presence of active principles like alkaloids, saponins, glycosides, tannins, terpenoids,

and flavonoids. With this, the ethanolic extract of the plant mentioned was proven to exhibit

antimicrobial properties.

Another study that used Escherichia coli bacteria, and Staphylococcus aureus were

explored by Biswas, Rogers, McLaughlin, Daniels, and Yadav (2013). The researchers

investigated the antimicrobial properties of guava leaf- extracts against the two gram-negative

bacteria (Escherichia coli and Salmonella enteritidis) and gram positive bacteria (Staphylococcus

aureus and Bacillus cereus) through well- diffusion method. The guava leaves were extracted in

four different solvents such as hexane, methanol, ethanol and water. Findings revealed that the
ethanol and methanol extracts showed inhibitory activity on gram-positive bacteria and it showed

that Escherichia coli and Salmonella enteritidis are resistant against the said extracts.

Further studies are researched by Foroughi, Pournaghi, Zhaleh, Zangeneh and Moradi

(2016) whereas the essential oil of Foeniculum vulgare was used to find the chemical

composition and antibacterial activity in Escherichia coli and Staphylococcus aureus. Screen test

and diffusion methods were done to locate antibacterial properties of the F. vulgare oil. The

outcome showed that the essential oil of F. vulganere with 0.007 g/ml concentration had averted

E. coli and 0.003 g/ml has averted S. aureus.

Chowdhury, Kubra & Ahmed (2014) also used diffusion method to determine the

antibacterial activity of Lawsonia inermis against five-gram negative bacteria including E. coli,

P.aeruginosa and S. aureus, B. sibtilis and P. epidermis . Methanol, chloroform, acetone, and

water were used as extraction solvent while dimethyl sulfoxide were used as dissolution solvent.

Based from the observation, these bacteria have no sign of growth after 24 hours of exposure to

extracts except B. sibtilis and P. epidermis.

A study of Nwankwo, Ezekoye, and Igbokwe (2014) examined the antibacterial activity

of honey against Escherichia coli, Staphylococcus aureus, and Candida albicans using diffusion

method. They concluded that honey has strong inhibiting activity on E. coli and S. aureus,

having a clearance zone of 34 mm and 45 mm respectively. In addition, Monte, Abreu, Borges,

Simoes, and Simoes (2014) also used Escherichia coli and Staphyloccus aureus. They

investigated the antimicrobial properties of selected phytochemicals such as hydroxycoumarin,

indole-3-carbinol, salicylic acid, and saponin against the two bacterium. Monte et al. (2014)

discovered that the phytochemicals of hydroxycoumarin and indole-3-carbinol were the most

promising phytochemical.
On the other hand, Ahmad and Beg (2000) studied 45 Indian medicinal plants against

certain drug-resistant bacteria and yeast. The ethanolic extracts from the different plants were

tested on Candida albicans, Staphyloccus aureus, Escherichia coli, Bacillus subtilis, Shigella

dysentriae,and Salmonella paratyphi and among 45, only 40 was proven to show antimicrobial

properties. Using TLC bioautography, thin layer chromatography, and qualitative phytochemical

tests, the researchers were able to determine the phenolic constituents of the said medical plants

which include tannins, phenols, and flavonoids.

Other related journals also explored solutions as an alternative to the commercial

antimicrobials which is proven to have adverse side effects. Two of the researchers were Ajayi

and Fadeyi (n.d.) who analyzed the phytochemical constituents and antimicrobial property of

Moringa oleifera leaves on Strepptococcus species and Staphycoccus aureus. Using diffusion

method, 0.6 g/ml inhibition zone was observed and results on the phytochemical screening

displayed that the petroleum extract obtained from the said plants is proven to contain bioactive

activity like tannins and flavonoids against the specified clinical microorganisms.

Another phytochemical screening was shown in the work of Bukar, Mukhtar and Hassan

(2009). The researchers used the leaf of Senna siamea to obtain chloroform, ethanolic and

aqueous extracts against the microorganism Pseudomonas aeruginosa. The sample extracts were

evaluated using agar diffusion method and findings revealed that the extracts were active at high

concentration levels. It was recorded that aqueous extract was found to be the most active in

terms of inhibition zone and this was followed by ethanolic extract. To sum up, S. siamea

exhibits phytochemical constituents like tannins, steroids and saponnins which can be a

treatment against the said disease causing microorganism.


Similarly, Kubmarawa, Ajoku, and Okorie (2007) also used Pseudomonas aeruginosa

together with against Bacillus subtilis, Escherichia coli, Staphylococcus aureus, and Candida

albicans. The researchers investigated the antimicrobial activity of fifty medicinal plants and

results showed that 28 plants had prevented the growth of one or more bacteria. Moreover, it has

revealed the content of the plants are tannins, saponins, alkaloids, glycosides, flavonoids and

essential oils. This proves that with appropriate chemical, plants can turn into effective medical

drugs.

Sedum morganianum is a tropical species that has long been grown as an ornamental for

its distinctive, evergreen foliage. It has several common names including burro’s tail, donkey’s

tail. It is a tender perennial plant that has trailing stems and succulent, blue-green leaves with a

silvery bloom that rubs off when handled (Mahr, 2016).

According to the Master Gardener Program (2016), the genus Sedum is a large group

with numerous species in the stonecrop family (Crassulaceae). As stated by the Herbal

resource(n.d.), Sedums contain tannins, rutin (citrus flavanoid glycosine), several alkaloids,

sedinon and sedridin and several organic acids but the plant’s potency is reduced when dried.

They’re grown indoors rarely blooms but mooving it outdoors during summer and keeping them

at cooler temperatures rather that room temperatures may encourage flowering and are best

grown in sturdy containers where the stems cam hang down (Master Gardener Program, 2016).

Since Sedum morganium is under the Crussalaceae family, they exhibit a carbonfixation called

the Crussalaceaen Acid Metabolism (CAM). In this metabolism, the stomata of the plant are

closed throughout the day and open during the night to collect carbon dioxide, and will be set

into malic and organic acid. Consequently, it will be accumulated and stored in the cellular

vacuoles until the sun’s energy is available for photosynthesis, which reduces water loss so these
plants survive longer on conditions where they aren’t watered frequently especially during hot

seasons (Encyclopedia Britannica, 2008 as cited by Delmas et al., 2017).

In the 16th century, healers started prescribing Sedums as a treatment for intestinal

parasites because of the poisonous properties of its flower and were also used as powerful

diuretics and laxatives but excessive intake could result to gastrointestinal pain sunch as diarrhea.

Around 1830, healers believed that sedums increase blood flow and could cure epilepsy but

scientific reviews suggest that sedums if symptoms are acute, can be used to lower high blood

pressure and regular intake of sedum can induce hypertension. Sedums should not be used by

pregnant women because it can trigger miscarriage but there are hypothesis which claim that

sedums can be used to treat ringworms, scurvy and diptheria.

In the study of Bensouici et al. (2016), it is the first study on the phytochemistry,

antioxidant, anticholinesterase, and antibacterial activities Sedum caeruleum L. They isolated the

secondary metabolities and determined the anioxidant, anicholinesterase, and antibacterial

activities os S. caeruleum.

Since some plants under the genus of Sedum and Crassulaceae family are already proven

to have chemical compounds that are useful for medical purposes, the researchers intend to do a

phytochemical screening on Sedum morganium, which is also under the same genus and family,

in order to know more about the plant in terms of its antibacterial and chemical properties. The

researchers would like to observe if it has active compounds that can be used to make medicinal

drugs.

The study would add in the knowledge in botany. Moreover, it would also contribute in

addressing problems such as antibiotic resistant bacteria The study aims to determine some

chemical constituents found in the plant after the phytochemical screening process as well as
investigating its antibacterial properties. Specifically, the study aims to answer the following

questions:

1. What are the phytochemicals present in Sedum morganium?

2. Which of the following strains of bacteria is mostly affected by the Sedum

morganium extract basing on their means?

a. Escherichia coli

b. Staphylococcus aureus

c. Pseudomonas aeruginosa

3. Is there a significant difference between the antimocrobial effect of Sedum

morganium extract and the control (Ciprofloxacin) in terms of inhibition zone on

Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa?

a. 50% S. morganianum + 50% distilled water

b. 100% S. morganianum

c. Control (Ciprofloxacin)

Significance of the Study

Despite the abundance and easy propagation of S. morganianum in the locality, there are

limited researches, both locally and internationally, tackling on its benefits and possible uses

(Hodgson, 2007). The study would identify the phytochemical contents of Sedum morganianum

which is the first step in identifying if a plant can be possible source of medicine. Moreover, this

study can there as reference for future researches concerning the S. morganianum and

phytochemical analysis. The result of this study would also greatly benefit the environment, for it

will produce an antibacterial against Escherichia coli, Staphylococcus aureus and Pseudomona

aeruginosa which can be used an alternative for commercial antimicrobial that have adverse
effect on our environment. According to the Upsala Journal of Medical Sciences (2014),

antibiotics leave a residue that can pollute and has many risks that can harm the environment.
Methodology

The procedures would be divided into two parts which would be the phytochemical

screening test method and the antibacterial test method (Disc diffusion).

Gathering of Materials

1. The Sedum morganianum will be obtained from Cruz, La Trinidad, Benguet.

2. Chemical reagents such as potassium mercuric iodide, sodium hydroxide solution, ferric

chloride solution and others will be obtained from Natural Sciences Research Unit

(NSRU) Saint Luis University, where the phytochemical screening test will be

conducted.

3. The different laboratory apparatuses such as incubator, test tubes, evaporating plates,

cotton swabs, petri dishes and others will be obtained from NRSU-SLU, where the disc

diffusion test will be conducted.

Preparation of Sedum morganianum

First, the plant will be disinfected before applying the maceration method. After that, it

will be set aside and will be used for the phytochemical screening test and the disc diffusion

test.

Phytochemical screening test

The extracts will be treated using particular chemical reagents depending on the

phytochemicals that will be detected. This will be done from the standard procedures

provided by Tiwari et al. (2011).

Disc Diffusion Test


The extracts will be divided into two treatments. First will be the one hundred percent

sedum morganianum extract and second will be fifty percent sedum morganianum and fifty

percent distilled water. White paper discs will be soaked on the treatments. In the meantime,

pure bacterial culture will be rubbed gently with light pressure on the prepared agar media

using cotton swabs. Afterwards, the paper disc from the treatment along with the control will

be put on the swabbed agar media using forceps. Finally, the petri dishes will be incubated

under temperature 37 degrees Celsius for 18 hours.