Chapter I. Introductionremeleven.weebly.com/uploads/1/3/5/1/13511732/chapters_i...Philippine Normal...

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Philippine Normal University National Center for Teacher Education

1 | R e s e a r c h P a p e r

Chapter I. Introduction

Plants produce a diverse range of bioactive molecules making them

a best source of different types of medicine. Nowadays, most of the drugs

are obtained from these natural resources because they can serve the

purpose without any side effect that is often associated with the synthetic

antimicrobials. Many plants have been used because of their antimicrobial

traits, which are due to compounds synthesized in the secondary

metabolism of the plant.

Different drugs, naturally and synthetically, are produced and

modified to fight different kinds of microorganisms like Escherichia coli,

which is commonly known as E. coli, a gram-positive kind species of

bacteria normally present in human intestine.

Two of the natural products that have been proven by different

researches are the Ixora coccinea, or the santan; and the Eichhornia

crassipes, or the water Hyacinth, having anti-microbial compounds which

have been proven to be a good drug against different bacteria like E. coli.

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Plants having an anti-microbial compounds when tested with the

bacteria are not that good compared to synthetic products but as they were

combined with other compounds of other plants, they work synergistically,

in which when they act together, their potency of fighting different

microorganisms increases compared to their individual potency.

Statement of the Problem

This study aimed to determine the synergistic effect of anti-

microbial property of Ixora coccinea and Eichhornia crassipes

extracts against Escherichia coli.

Specifically, this study sought to answer the following

questions:

1. Is there a synergistic effect between Ixora coccinea and Eichhornia

crassipes extracts?

2. What extract is the most ideal anti-microbial drug to be used against

Escherichia coli?

3. In what strain of Escherichia coli would be Ixora coccinea and

Eichhornia crassipes extracts would be good anti-microbial drug?

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Significance of the Study

Escherichia coli, commonly known as E. coli, is one of the

most common species of microorganisms that are normally present in the

human intestine that can cause diarrhea, blood poisoning that can even

lead to death. Anti-microbial drugs are usually at high cost and one of the

alternatives for anti-microbial drugs are plant extracts. Ixora coccinea

extract and Eichhornia crassipes extract are good anti-microbial against

Escherichia coli. Combining the extracts of these plants would be able to

have synergistic effect, thus providing a more effective and more potent

anti-microbial drug against Escherichia coli. This study will also be helpful

in pharmaceutical realms that can yield to a powerful drug against any

other microorganisms.

Scope and Limitations

This research is a test about synergistic effect of Ixora

coccinea and Eichhornia crassipes extracts against Escherichia coli

and is limited in ethanoic solvent; and in the bioassay that will be

used.

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Definition of Terms

Anti-Microbial

- A plant substance that acts to inhibit the growth of harmful

microorganisms, or acts to destroy them.

Bioactive Molecules

- Molecules that can have an effect on living tissue

Bioassay

- A technique for determining the concentration or potency of a

substance such as a drug by measuring its effect on an

organism

Potency

- Capacity to produce strong physiological or chemical effects

Synergism/Synergistic

- Working together of two or more drugs, muscles, etc., to

produce an effect greater than the sum of their

individual effects

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Chapter II. Related Literature

It is a well-known fact that medicinal plants are the resources of

promising drugs for many diseases. And also according to the international

respected health institute, the World Health Organization, one of the best

sources to obtain a variety of drugs would be the medicinal plants.1

Scientists worldwide are exploring the possibilities of finding out and

utilizing pharmacologically active compounds from medicinal plants.

Screening of medicinal plants for their phytochemicals, antioxidant,

anticancer and antimicrobial properties is the prime concern for finding out

an effective phytochemically active principle.2

Scientific experiments since the late 19th century have documented

the antimicrobial properties of some plants, herbs, and their components.3

The antimicrobial activity of plant oils and extracts has formed the basis of

many applications, including raw and processed food preservation,

pharmaceuticals, alternative medicine and natural therapies.4

1 (Santos et al., 1995) 2 Ayyanar and Ignacimuthu, 2008; Agbafor et al., 2011; Roy et al., 2011; Vinoth et al., 2011; Mishra and Tripathi, 2011 3 (Shelef, 1983; Zaika, 1988; Alzoreky and Nakahara, 2003; Kumral and Sahin, 2003; Park et al., 2009) 4 Hammer KA, Carson CF, Riley TV, Antimicrobial activity of essential oils and other plant extracts. J Appl Microbiol. 1999; 86: 985-990

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Ixora coccinea is a species of flowering plant in the Rubiacceae

family. It is a common flowering shrub native to Southern India, where its

name was derived which means Indian deity.5 It has been used in the past

as a therapy or a remedy to treat different kinds of diseases such as

wounds and inflamed tissues.6 It has been proven to have an anti-microbial

property and an anti-inflammatory property.7

Figure2.1: Ixora coccinea plant, leaves and flowers.

Leaves, flowers, roots and stems of Ixora coccinea were utilized as a

medicine. Its leaves contained flavonols, kaemferol and quercetin,

proanthocyanidins, phenolic acids and ferulic acids. It is considered to be

5 Wikipedia, February 2012. http://www.wikipedia.com/ixora_coccinea 6 Effect of Ixora coccinea flowers on dead space wound healing in rats / B S Nayak et al / Fitoterapia • vol. 70, no3, pp. 233-236, 1999 / doi:10.1016/S0367-326X(99)00025-8 7 Antimicrobial activity of Ixora coccinea leaves / Fitoterapia Vol 74, Issue 3, April 2003, Pages 291-293

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antiseptic and anti-microbial,8 which was detected in ethanolic extract. The

effective inhibitory concentration of the extract against bacterial test

organisms was 125µg mL-1, beyond which the inhibitory activity declined

and the organisms started reviving from the effect of the antimicrobial

principle.9

Water hyacinth is an erect free-floating and stoloniferous perennial

herb, which belongs to the Family Pontederiaceae.10 It is the most

damaging aquatic plant worldwide.11

Figure 2.2: Eicchonira crassipes leaves..

8 Stuart Xchange, 2012. http://www.stuartxchange.org/Santan.html 9 Arshad, and Shahid Hussain. Evaluation of anti-mycobacterial activity of garlic (Allium sativum) against clinical isolates of non-MDR and MDR Mycobacterium tuberculosis. Planta Med. 2009; 75: 1073 10 Nyananyo, Bl; Gijo Ah; Ogamba, En; The Physico-chemistry and Distribution of Water Hyacinth (Eichhornia cressipes) on the river Nun in the Niger Nelta 11 Moody, K.; Munroe, C.; Lubigan, R.; Paller , E. (1984): Major weeds of the Philippines. University of the Philippines, Los Baños. Laguna, Philippines.

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The environmental hazards associated with large Eichhornia

crassipes populations is degraded water quality and drastic changes in the

plant and animal community, light and oxygen diffusion are severely

curtailed by this floating plant, reduction in water movement and the

decomposition of the plants, if large biomass of the plants are killed at

once, can use up all the 12oxygen in the water.13

However, different compounds such as tannins and flavonoids

(flavonols and kaempferol) which are antiseptic, astringent and

antimicrobial have been reportedly isolated from this plant.14

Ethanol extracts of Eichhornia crassipes root showed antimicrobial

activities against Escherichia coli. Extracts of Eichhornia crassipes leaves

are more effective than that of its roots.15

Our daily lives are closely intertwined with microorganisms, and the

health of our planet depends very much on their activities.16 Some

microorganisms can be mutual to us but some were not.

12 Moody, K.; Munroe, C.; Lubigan, R.; Paller , E. (1984): Major weeds of the Philippines. University of the Philippines, Los Baños. Laguna, Philippines. 13 (Gopal,1987) 14 Moody, K.; Munroe, C.; Lubigan, R.; Paller , E. (1984): Major weeds of the Philippines. University of the Philippines, Los Baños. Laguna, Philippines. 15 Antimicrobial activity of some macrophytes from Lake Manzalah (Egypt). Fareed MF, Haroon AM, Rabeh SA.

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One of the microorganisms that are very disadvantageous to us, a

microorganism that can cause food poisoning is Escherichia coli that

produce high level of toxins that can cause kidney malfunction, blood-

poisoning, diarrhea and can even lead to death.17 Different plant extracts

are already been studied which is proven to kill this microorganism like C.

sativum.18

Plants are been investigated to better understand their properties,

safety and efficiency.19 Different studies determined that combinations of

plant extracts against pathogenic bacteria resulted synergistically.20

Synergism is interaction of discrete agencies (as industrial firms), agents

(as drugs), or conditions such that the total effect is greater than the sum

of the individual effects.21

Individually, extracts of Ixora coccinea is slightly not favorable anti-

microbial extract against Escherichia coli;22 compared to Eichhornia

crassipes23, but then some plant extract like C. auriculata and C.

16 Microsoft ® Encarta ® 2009. © 1993-2008 Microsoft Corporation. All rights reserved. 17 Microsoft ® Encarta ® 2009. © 1993-2008 Microsoft Corporation. All rights reserved. 18 Toroglu, 2010. Journal of Environmental Biology 19 (Ellof, 1998; Nascimento et al., 2000; Mothana et al., 2009) 20 Natchimuthu Karmegan, Mani Jauakumar and Subbiah Karuppusamy, 21 Merriam-Webster. http://www.merriam-webster.com/dictionary/synergism 22H.O. Oladimeji, P.M. Ubulom, E.I. Akpabio, I.E. Etim and E. Nyong 23 Antimicrobial activity of some macrophytes from Lake Manzalah (Egypt). Fareed MF, Haroon AM, Rabeh SA.

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quadrangularis which is not good anti-microbial extract against Escherichia

coli, but their combined extract resulted synergistically.24

Tannins and flavonoids, which are said to be anti-microbial

compounds,25 can be found in the extract of Ixora coccinea and Eichhornia

crassipes, which can be synergistic same as what happened to the extract

of C. auriculata and C. quadrangularis.26

Some of plants that were combined do not exhibit synergism, thus it

is not ideal to act together; some exhibit antagonism, which when they

interact, their potency decreases compared to their individual effectiveness

like what happened to A. amara and A. marmelos against Escherichia

coli.27

This research tends to prove any synergistic effect of the anti-

microbial property of Ixora coccinea extract and Eichhornia crassipes

extract against Escherichia coli. Also, the solvent that will be used will be

assessed; on what solvent would the anti-microbial compounds of these

extracts be more potent against Escherichia coli.

24 Natchimuthu Karmegan, Mani Jauakumar and Subbiah Karuppusamy, 25 Lamikanra et al., 1990; Burapadaja and Bunchoo, 1995; Adesina et al., 2000) 26 Natchimuthu Karmegan, Mani Jauakumar and Subbiah Karuppusamy, 27 Natchimuthu Karmegan, Mani Jauakumar and Subbiah Karuppusamy,

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Chapter III. Methodology

Phase 1 - Collection of Plant Samples

The plant samples were collected in some parts of the

Philippines, particularly in Caloocan City for Ixora coccinea

and Binan City, Laguna for Eichhornia crassipes. The

collected samples were verified at the Botany Division of

National Museum.

Phase 2 - Cleaning and Drying of Plant Samples

The collected plant samples were washed and dried at the

oven for 24 hours. The leaves were left inside the oven for 24

hours so that the moisture will be gone and cannot hinder in

the extraction process.

Phase 3 - Extraction of Santan and Water Hyacinth

leaves

The leaves of Santan and Water Hyacinth were pounded

using a mortar and pestle. Plant sample were soaked in

ethanol solvent for 48 hours. Solvents will be removed through

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rotary vacuum evaporator and collected extract were stored at

4oC in air tight bottles.

Phase 4 - Anti-Microbial testing of Santan and Water

Hyacinth Extracts

Antibacterial testing were conducted in DOST-ITDI Standards

and Testing Division by disc diffusion method. The test was

tested against Escherichia coli. The plant extracts were tested

using organic solvent, Ethanol.

1. Ixora coccinea in Ethanol

2. Eichhornia crassipes in Ethanol

3. Ixora coccinea and Eichhornia crassipes in Ethanol

This had been determined which of the three assays was the

most effective anti-microbial agent specifically against

Escherichia coli

*The first two solvents were the control samples.

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Schematic Diagram

Phase 3Extraction ofPlant Samples

Plant leaves samples were pounded and were soaked in ethanol solvent for 48 hours

Solvents were removed through rotary vacuum evaporator and

collected extract were stored at 4oC in air tight bottles

Phase 4 Anti Microbial

Testing

The extracts were tested using organic solvent, Ethanol.

Antibacterial testing were conducted in DOST-ITDI

Standards and Testing Division by disc diffusion method. The test was tested against Escherichia

coli.

Phase 1 Collection of Plant

Samples

Ixora coccinea and Eichhornia crassipes

were collected

Plant samples were verified by the National

Museum or BPI

Phase 2 Cleaning and Drying

the Plant Sample

Collected plant samples were washed.

Plant leaves samples were dried in an oven

for 24 hours

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Chapter IV. Data Analysis

Data were gathered from the observations and experiments done in

the extracts of Ixora coccinea and Eichhornia crassipes against

Escherichia coli. In this research, tables were used for visualization of the

data that can be seen in the appendices of this paper.

The data that were gathered and recorded were divided into three

experiments, one is for the Ixora coccinea extract, another is for

Eichhornia crassipes extract and the last is the combination of two

extracts. Each experiment was further divided into three trials; and there

were one petri dish in each of the extract and the combination. Every day,

the results has been gathered, recorded and tabulated (Refer to Tables

6.1 and 6.2 in Appendix).

Tables were divided into two, first column is the Single Extract

Effect which is about the individual effect of the two leaves extracts; the

second column is the Combination Extract Effect which is about the

combined leaves extracts of Ixora coccinea and Eichhornia crassipes.

In the first trial of Ixora coccinea extract in the strain-2 of

Escherichia coli, the zone of inhibition showed on the 1st day of the

observation period, which is 4 millimeters; on the 3rd day of the

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observation period, the zone of inhibition became 9 millimeters. The zone

of inhibition remained the same up to the 5th day of the observation

period.

In the second trial of Ixora coccinea extract in the strain-2 of

Escherichia coli, the zone of inhibition also showed on the 1st day of the


observation period, the zone of inhibition increases to 7 millimeters and

remained the same up to the 5th day of the observation period.

In the third trial of Ixora coccinea extract in the strain-2 of

Escherichia coli, same observations were gathered, the zone of inhibition

was seen on the 1st day of the observation period which is 4 millimeters

and it increases to 7 millimeters on the 3rd day and up to the 5th day of

observation period.

In the first trial of Eichhornia crassipes extract in the strain-2 of

Escherichia coli, the zone of inhibition showed on the 1st day of

observation period, which is 5 millimeters. It increases to 6 millimeters on

the 2nd day and still increases on the 3rd day to 10 millimeters. It increases

to 15 millimeters on the 5th day of the observation period. In the second

trial of Eichhornia crassipes extract in the strain-2 of Escherichia coli, the

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zone of inhibition was observed on the 1st day of the observation period

with 6 millimeters. On the 2nd day, it increases to 8 millimeters and

continue to increase on the 3rd day up to 9 millimeters. It remained the

same up to the 5th day of the observation period. In the third trial of

Eichhornia crassipes extract in the strain-2 of Escherichia coli, it was on

the 1st day of the observation period when the zone of inhibition appeared

which is 5 millimeters. It increases to 6 millimeters on the 2nd day,

continue to increase on the 3rd day which is 8 millimeters and remained

the same up to the 5th day of the observation period.

In the first trial of the combination extracts of Ixora coccinea extract

and Eichhornia crassipes extract in the strain-2 of Escherichia coli, the

zone of inhibition was observed on the 1st day with 2 millimeters and 5

millimeters in Ixora coccinea extract and Eichhornia crassipes extract,

respectively. On the 2nd day, inhibition was observed in Ixora coccinea

extract with 4 millimeters and for Eichhornia crassipes extract with 5

millimeters. The trend increases until on the 5th day, the inhibition was

observed with 7 millimeters and 9 millimeters for Ixora coccinea extract

and Eichhornia crassipes extract, respectively.

In the second trial of the combination extracts of Ixora coccinea

extract and Eichhornia crassipes extract in the strain-2 of Escherichia coli,

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the zone of inhibition was observed on the 1st day with 3 millimeter and 5







In the third trial of the combination extracts of Ixora coccinea extract

and Eichhornia crassipes extract in the strain-2 of Escherichia coli, it was

observed on the 1st day when the zone of inhibition was observed with 4

millimeter and 6 millimeters, for Ixora coccinea extract and Eichhornia

crassipes extract, respectively. On the 2nd day, inhibition was observed in

Ixora coccinea extract with 5 millimeters and for Eichhornia crassipes

extract with 6 millimeters. The trend increases until on the 5th day, the

inhibition was observed with 7 millimeters and 8 millimeters for Ixora

coccinea extract and Eichhornia crassipes extract, respectively.

The summary of the data gathered in the zone of inhibition of strain-

2 of Escherichia coli is tabulated in table 4.1.

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Table 4.1: Inhibition zone of strain-2 Escherichia coli in single extract effect of two extracts and the combination extract effect of two extracts.

Data were gathered from the experiment for the strain-78 of

Escherichia coli. In the first trial of Ixora coccinea extract in the strain-78

of Escherichia coli, the zone of inhibition showed on the 2nd day of the


observation period, the zone of inhibition became 8 millimeters. The zone

of inhibition remained the same up to the 5th day of the observation

period.

In the second trial of Ixora coccinea extract in the strain-78 of

Escherichia coli, the zone of inhibition also showed on the 2nd day of the


observation period, the zone of inhibition increases to 6 millimeters and


Day 05

Single Extract Effect (mm)

Combination Extract Effect (mm)

Ixora coccinea

Eichhornia crassipes

Ixora coccinea


Trial 01 9 15 7 9

Trial 02 7 9 7 9

Trial 03 7 8 7 8

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In the third trial of Ixora coccinea extract in the strain-78 of

Escherichia coli, the zone of inhibition was seen on the 2nd day of the

observation period which is 8 millimeters and remained the same up to

the 5th day of observation period.

In the first trial of Eichhornia crassipes extract in the strain-78 of

Escherichia coli, the zone of inhibition showed on the 1st day of

observation period, which is 2 millimeters. It increases to 6 millimeters on

the 2nd day and still increases on the 3rd day to 10 millimeters. It remained

the same up to the 5th day of the observation period.

In the second trial of Eichhornia crassipes extract in the strain-78 of

Escherichia coli, the zone of inhibition was observed on the 1st day of the

observation period with 6 millimeters. On the 2nd day, it increases to 8

millimeters and continue to increase on the 3rd day up to 10 millimeters. It


In the third trial of Eichhornia crassipes extract in the strain-78 of

Escherichia coli, it was on the 1st day of the observation period when the

zone of inhibition appeared which is 5 millimeters. It increases to 6

millimeters on the 2nd day, continue to increase on the 3rd day which is 8

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millimeters and remained the same up to the 5th day of the observation

period.

In the first trial of the combination extracts of Ixora coccinea extract

and Eichhornia crassipes extract in the strain-78 of Escherichia coli, the

zone of inhibition was observed on the 1st day with 0 millimeter and 3







In the second trial of the combination extracts of Ixora coccinea

extract and Eichhornia crassipes extract in the strain-78 of Escherichia

coli, the zone of inhibition was observed on the 1st day with 0 millimeter

and 6 millimeters in Ixora coccinea extract and Eichhornia crassipes

extract, respectively. On the 2nd day, inhibition was observed in Ixora

coccinea extract with 4 millimeters and for Eichhornia crassipes extract

with 8 millimeters. The trend increases until on the 5th day, the inhibition

was observed with 7 millimeters and 13 millimeters for Ixora coccinea

extract and Eichhornia crassipes extract, respectively.

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In the third trial of the combination extracts of Ixora coccinea extract

and Eichhornia crassipes extract in the strain-78 of Escherichia coli, it was

observed on the 1st day when the zone of inhibition was observed with 0

millimeter and 6 millimeters, for Ixora coccinea extract and Eichhornia

crassipes extract, respectively. On the 2nd day, inhibition was observed in

Ixora coccinea extract with 5 millimeters and for Eichhornia crassipes

extract with 6 millimeters. The trend increases until on the 5th day, the

inhibition was observed with 8 millimeters and 9 millimeters for Ixora

coccinea extract and Eichhornia crassipes extract, respectively.

The summary of the data gathered in the zone of inhibition of strain-

78 of Escherichia coli is tabulated in table 4.2

Table 4.2: Inhibition zone of strain-78 Escherichia coli in single extract effect of two extracts and the combination extract effect of two extracts.

Day 05

Single Extract Effect (mm)

Combination Extract Effect (mm)

Ixora coccinea


Ixora coccinea


Trial 01 8 10 8 11

Trial 02 6 10 7 13

Trial 03 8 8 8 9

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Different statistical tools were used to analyze the data gathered and

to measure any significance of this study in the field of science.

Percentage was used to measure the percent increase of the zone

of inhibition of the combined Ixora coccinea and Eichhornia crassipes

extracts against the 2 strains of the bioassay.

Table 4.3: Percentage increase of the combined extracts compared to the individual extracts in strain-2 Escherichia coli. Percentage increases is calculated as (B-A) / A x 100.

In table 4.3, referring to the first trial, the percentage increase of the

combined Ixora coccinea extract effect compared to the single Ixora

coccinea extract effect against the strain-2 of Escherichia coli, the result is

negative which means that instead of a synergistic effect that should be

Percentage Increase in Strain-2 Escherichia coli

(A) Single Extract Effect (mm)

(B) Combination Extract Effect (mm)

Percentage Increase

Ixora coccinea


Ixora coccinea


Ixora coccinea


Trial

01

9 15 7 9 -22.0% -40.0%

Trial

02

7 9 7 9 0.0% 0.0%

Trial

03

7 8 7 8 0.0% 0.0%

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expected, an antagonistic effect was observed. From 9 mm zone of

inhibition in the single extract effect, it decreases to 7 mm when combined

with Eichhornia crassipes extract. For the percentage increase of the

combined Eichhornia crassipes extract effect compared to the single

Eichhornia crassipes extract effect against the strain-2 of Escherichia coli,

the result is -40.0%, a negative result, which means an antagonistic effect

is observed instead of a synergistic effect that is expected. The Ixora

coccinea greatly affects in the inhibition growth of the Eichhornia crassipes

extract. From 15 mm zone of inhibition in the single extract effect, it

decreases to 9 mm when combined to the Ixora coccinea extract.

For second and third trials, the percentage increase of the combined

Ixora coccinea extract effect compared to the single Ixora coccinea extract

effect against the strain-2 of Escherichia coli is zero, which means no

change in the zone of inhibition is observed. Same with the Eichhornia

crassipes extract effect compared to the single Eichhornia crassipes

extract effect.

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Table 4.4: Percentage increase of the combined extracts compared to the individual extracts in strain-78 Escherichia coli. Percentage increases is calculated as (B-A) / A x 100.

In table 4.4, referring to the first trial, the percentage increase of the

combined Ixora coccinea extract effect compared to the single Ixora

coccinea extract effect is zero which means that Eichhornia crassipes

extract does not have an effect to Ixora coccinea extract. However, the

percentage increase of the combined Eichhornia crassipes extract effect

compared to the single Eichhornia crassipes extract effect is a positive

10.0% which means the a synergistic effect is observed; thus, Ixora

coccinea extract affects the zone of inhibition of Eichhornia crassipes.

Percentage Increase in Strain-78 Escherichia coli

(A) Single Extract Effect (mm)

(B) Combination Extract Effect (mm)

Percentage Increase

Ixora coccinea


Ixora coccinea


Ixora coccinea


Trial

01

8 10 8 11 0.0% 10.0%

Trial

02

6 10 7 13 14.3% 30.0%

Trial

03

8 8 8 9 0.0% 12.5%

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For the second trial, the percentage increase of the combined Ixora

coccinea extract effect compared to the single Ixora coccinea extract effect

is 14.3% which means that Eichhornia crassipes extract has an effect to

Ixora coccinea extract. The percentage increase of the combined

Eichhornia crassipes extract effect compared to the single Eichhornia

crassipes extract effect is 30.0% which means that Ixora coccinea extract

has an effect to Eichhornia crassipes extract.

For the last trial, the percentage increase of the combined Ixora

coccinea extract effect compared to the single Ixora coccinea extract effect

is zero which means that Eichhornia crassipes extract does not have an

effect to Ixora coccinea extract. However, the percentage increase of the

combined Eichhornia crassipes extract effect compared to the single

Eichhornia crassipes extract effect is a positive 12.5% which means the a

synergistic effect is observed; thus, Ixora coccinea extract affects the zone

of inhibition of Eichhornia crassipes.

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Chapter V. Summary of Findings, Conclusion and Recommendations

Data that were gathered was analyzed to know if the results showed

a significant effect in the combination of Ixora coccinea and Eichhornia

crassipes extracts against Escherichia coli.

Analysis of Variance (ANOVA) was used to know any significant

differences from the individual effect of Ixora coccinea and Eichhornia

crassipes extracts with respect to the combined Ixora coccinea and

Eichhornia crassipes extracts against the 2 strains of the bioassay.

Table 5.1: Analysis of variance of the individual effect of Ixora coccinea and Eichhornia crassipes extracts and the combined effect of Ixora coccinea and Eichhornia crassipes extracts to strain-2 Escherichia coli.

SUMMARY Groups Count Sum Average Variance

Single Extract

Ixora coccinea 3 23 7.666667 1.333333

Eichhornia crassipes 3 32 10.66667 14.33333

Combination Extract

Ixora coccinea 3 21 7 0


ANOVA Source of Variation SS df MS F P-value F crit

Between Groups 23 3 7.666667 1.916667 0.205416 4.066181

Within Groups 32 8 4

Total 55 11

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In table 5.1, ANOVA of strain-2 Escherichia coli, F-critical is greater

than F, thus it showed no significant difference between the zones of

inhibition in the individual extracts of Ixora coccinea and Eichhornia

crassipes and with the combined extracts of Ixora coccinea and Eichhornia

crassipes. In summary, there is no synergistic effect of antimicrobial

property of Ixora coccinea and Eichhornia crassipes against strain-2 of

Escherichia coli.

Table 5.2: Analysis of variance of the individual effect of Ixora coccinea and Eichhornia crassipes extracts and the combined effect of Ixora coccinea and Eichhornia crassipes extracts to strain-78 Escherichia coli.

SUMMARY Groups Count Sum Average Variance

Single Extract

Ixora coccinea 3 22 7.333333 1.333333


Combination Extract

Ixora coccinea 3 23 7.666667 0.333333

Eichhornia crassipes 3 33 11 4

ANOVA Source of Variation SS df MS F P-value F crit

Between Groups 25.66667 3 8.555556 4.888889 0.032324 4.066181

Within Groups 14 8 1.75

Total 39.66667 11

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In table 5.2, ANOVA of strain-78 Escherichia coli, F is greater than

F-critical, thus it showed significant difference between the zones of

inhibition in the individual extracts of Ixora coccinea and Eichhornia

crassipes and with the combined extracts of Ixora coccinea and Eichhornia

crassipes. In summary, there is a synergistic effect of antimicrobial

property of Ixora coccinea and Eichhornia crassipes against strain-78 of

Escherichia coli.

Conclusion

This data suggested that the extracts of Ixora coccinea and

Eichhornia crassipes do not have a synergistic effect in the anti-microbial

property against strain-2 of Escherichia coli; however the extracts have

synergistic effect in the anti-microbial property against strain-78 of Ixora

coccinea and Eichhornia crassipes.

This suggest that even though statistically there is a synergistic

effect that is observed in strain-78 of the bioassay, practically there is none

because it is only the Eichhornia crassipes extract which has a percentage

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increase while the Ixora coccinea extract do not have a consistent percent

increase in zone of inhibition, referring to table 5.2.

In conclusion, there is no synergistic effect of anti-microbial property

of Ixora coccinea and Eichhornia crassipes against the strain-2 and strain-

78 of Escherichia coli.

Recommendation

The researchers recommend to use other strains of Escherichia coli

or other gram positive bacteria or even other bioassays for more precise

and accurate results.

The researchers also recommend to use disc diffusion assay and

other assay test that will vary the concentrations of each of the plant

leaves extracts.

.

`



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