ANTI-ANGIOGENIC AND ANTIBACTERIAL PROPERTY OF BANGUS BILE RESEARCH PAPER.doc

8/14/2019 ANTI-ANGIOGENIC AND ANTIBACTERIAL PROPERTY OF BANGUS BILE RESEARCH PAPER.doc

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ABSTRACT

I/We hereby certify that the above is the result of work done in my/our research.

__________ _________

Signature of Team Leader Date

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Category

Pick and shade only one box

The milkfish is an important seafood in Southeast Asia and some Pacific slands.!ecause milkfish is notorious for being much bonier than other food fish" deboned milkfish"

called #boneless bangus# in the Philippines" has become popular in stores and markets" yet

bangus bile is not gi$en importance and is being thro%n a%ay. The proponents further

in$estigate the uses of this &left o$er' to pro$ide usage for this specimen" one of those is anti(

carcinogenic %hich %as conducted by the researchers by using Chorioallantoic )embrane*CA)+ Assay.

The proponents gathered !angus bile from the public market to be used for the

experiment. The !angus bile %ere then air dried to remo$e moisture. After drying" the

proponents took the extract of the bangus bile through methanol soaking. The proponents bought

,- Duck eggs from a professional breeder. uickly thereafter" the proponents made threetreatments" !angus bile extract" positi$e control *Sim$astatin+ and a negati$e control *%ater+.

Then" filter papers %ere soaked on each treatment and placed on the CA) layer of the egg. Theeggs %ere then /uickly sealed and incubated for another 011 hours. After the incubation process"

the blood $essels of the treated eggs %ere obser$ed under a microscope and compared to the

control unit. The anti(angiogenic screening of the bangus bile extract sho%ed a significant result

in decreasing the number of blood $essels in the embryo of the duck egg based on the re2ection

of the null hypothesis in the analysis of $ariance.

3or anti(bacterial screening" the proponents used three treatments( !angus bile extract"

negati$e control *%ater+ and positi$e control *Amoxicillin+. 4ach treatment has three

replications. 5esults sho%ed 6ones of inhibition in both Staphylococcus aureusandEscherichia

coli %hich can be compared to the positi$e control.

Thus" the proponents concluded that the methanol extract of !angus bile" has an

antibacterial and anti(angiogenic property7 therefore can be used as a much cheaper alternati$e

to commercial medicine.

!iochemistry

!otany

Chemistry

Computer

Science

4ngineering

4n$ironmental

Science

)edicine and

8ealth

)icrobiology

Physics

Title: Anti-angiogenic and Antibacterial Property of Bangus(Chanos chanos) Bile

Proponents: Jared Carlo O. nrile! Joegin Christian "i#! "loydJoshua $oAd%iser: &ogelio Ponte'o


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TABLE OF CONTENTS

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INTRODUCTION

This part includes the background of the study" statement of the problem" hypothesis" significance of the

study" operational definition of terms" and the scope and limitations that %ould lead the readers about the research.

1.1 Background of the stud

Angiogenesis simply means the formation of ne% blood $essels. n a more specific definition it is a

physiological process in$ol$ing the gro%th of ne% blood $essels from pre(existing $essels" especially blood $essels

that supply oxygen and nutrients to cancerous tissues. t is a normal process in gro%th and de$elopment of the

embryo and fetus" as %ell as in %ound healing. 8o%e$er" this is also a fundamental step in the transition of tumors

from a dormant state to a malignant state. =e can simply find out if an ob2ect can cure or lessen the effects of

certain cancers" by the means of blood $essel count. This is done due to the fact that %ithout the blood $essels" the

cancer tissues %hich are present in our body cannot and %ill not exist" these blood $essels supply the needed

nutrients for the cancer tissues. 3urthermore" angiogenesis cannot be done %ith non(fertile eggs because %ith these

kinds of eggs" there is no sperm cell and egg cell formation.

)ilkfish ha$e a generally symmetrical and streamlined appearance" %ith a si6able forked caudal fin. They

can gro% to but are most often about 0 meter *E.1 in+ in length. They ha$e no teeth and generally feed on algae and

in$ertebrates. They occur in the ndian


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The ob2ecti$e of this study is to e$aluate the anti(angiogenic property and anti(bacterial property of bangus

bile extract.

1.+ S)gn)f)cance of the stud

The significance of this study is to see the anti(bacterial and anti(angiogenic characteristics of !angus bile

and be able to find out if it could be an alternati$e and ne% source of antibiotic. Also" %ith this study" the researchers%ill be able to help and contribute additional information to the medical field of science.

1., Sco#e and L)")tat)on

This study is only limited in the experimentation on the anti(angiogenic property of the !angus bile extract"

and the anti(bacterial property of the said extract. The anti(angiogenic property can be studied through the use of

sim$astatin as the positi$e control" %ater *negati$e control+" bangus bile extract" the combined property of the

bangus bile extract and sim$astatine and by solely depending on the blood $essel formation of the medium. The

anti(bacterial effect %ill be tested by usingEscherichia coli, Staphylococcus aureus and Pseudomonas aureginosa as

the specimens in :irby(!auer Testing. There %ere three treatments used in this study J Positi$e Control

*Amoxicillin+" ;egati$e Control *%ater+ and the bangus bile extract.

The said experiments %ere conducted from uly to September in the Science Laboratory of 9eneral Santos

8ope Christian School.

1.- Re*)e of re%ated L)terature

This chapter presents the re$ie% of related literature and the operational definition of terms used in this

research. Literatures %ere cited on the bases of their importance in the conduct of the study.

Ant)/Ang)ogen)c

a drug or substance used to stop the gro%th of tumors and progression of cancers by limiting the pathologic

formation of ne% blood $essels *angiogenesis+. *)osbyKs Dictionary of Complementary and Alternati$e )edicine+

Ant)$acter)a%

Antibacterial is the destroying or suppressing the gro%th or reproduction of bacteria.

*)iller(:eane 4ncyclopedia and Dictionary of )edicine" ;ursing" and Allied 8ealth+

Chor)oa%%anto)c "e"$rane

Also called the chorioallantois or abbre$iated to CA) is a $ascular membrane found in eggs of

someamniotes" such as birds and reptiles. t is formed by the fusion of the mesodermal layers of t%o de$elopmental

structuresH the allantois and the chorion. n mammals" this structure forms the placenta. *)osbyKs Dictionary of

Complementary and Alternati$e )edicine+

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Escher)ch)a co%)

Escherichia coli is found in large numbers as a normal inhabitant of the large intestine of %arm(blooded

animals. =hene$er they lea$e their usual habitat" these organisms can cause urinary tract infections" peritonitis"

endocarditis" and other diseases. Some strains cause se$ere gastroenteritis.

(9rolier 4ncyclopedia of :no%ledge)

0)r$/Bauer Ant)$)ot)c Test)ng D)sc D)ffus)on Test)ng2

:irby !auer Testing is a test %hich uses antibiotic(impregnated %afers to test %hether particular bacteria

are susceptible to specific antibiotics. :no%n /uantities of bacteria are gro%n on agar plates in the presence of thin

%afers containing rele$ant antibiotics. f the bacteria are susceptible to a particular antibiotic" an area of clearing

surrounds the %afer %here bacteria are not capable of gro%ing *called a 6one of inhibition+.

*en.%ikipedia.orgM%ikiM:irby(!auer_antibiotic_testing+

A"o3)c)%%)n

Amoxicillin is a moderate(spectrum" bacteriolytic" N(lactamantibiotic used to treat bacterialinfections

caused by susceptible microorganisms. t is usually the drug of choice %ithin the class because it is better absorbed"

follo%ing oral administration" than other N(lactam antibiotics. Amoxicillin is one of the most common antibiotics

prescribed for children.

*httpHMM%%%.medicinenet.comMnorfloxacin+

Sta#h%ococcus aureus

Staphylococcus is a genus of spherical bacteria" some species of %hich are normal inhabitants of the skin

and others of %hich causes disease. The most common pathogen" S.aureus" is fre/uently responsible for boils"

carbuncles" abscesses" osteomyelitis" and" sometimes" meningitis an pneumonia" and is the most common cause of

bacterial food poisoning and hospital infections.

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*9rolier 4ncyclopedia of :no%ledge+

Snerg

Synergy is the correlation or concurrence of action bet%een different organs in health or disease" or bet%een

different drugs. *=ebsterOs Dictionary of the 4nglish Language+

4ater)a%s and 4ethods

!.1 Research Des)gn

This study %ill follo% a simple research design. t attempts to sho% the importance and the medicinal effectof !angus *Chanos chanos)bile specifically the anti(angiogenic property of the said !angus bile.The study %ill

ha$e $arious tests conducted at the Science Laboratory of 9eneral Santos 8ope Christian School. The first part is%here the proponents extract the substance from the !angus bile *Chanos chanos).

The proponents then conduct the anti(angiogenic screening %herein Duck eggs %ere used as the medium.

The proponents conducted the :irby(!auer testing to see if the extract is effecti$e in inhibiting the gro%th of

bacteria. The bacteria used areEscherichia coli, seudomonas aeru!inosa,and Staphylococcus aureus.

Data gathered %as obser$ed after the treatment using the solution then analy6ed through blood $essel

count for each of the concentration.

!.! 4ater)a%s and E5u)#"ents

This sub(section presents the ra% materials and laboratory e/uipments to be used in the experimental

setup. These %ere categori6ed based on the antibacterial susceptibility test and anti(angiogenic testing.

4ater)a%s and Instru"ents used for ant)/ang)ogen)c screen)ng

The said study used $arious instruments and e/uipments to perform the anti(angiogenic screening.

Ta$%e !.1 4ater)a%s used for Ant)/Ang)ogen)c Screen)ng Ta$%e !.! 4ater)a%s used for 0)r$/Bauer Test)ng

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!.& 6rocedure F%o Chart

This part sho%s the procedures undertaken for the study7 a more detailed chart of the procedure is sho%nbelo%.

8

Primary)aterials

Laboratory4/uipments


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!.+ E3#er)"entat)on and 'enera% 6rocedure

6rocedure of e3tract)ng the su$stance fro" Bangus $)%e Chanos chanos2.

!angus !ile *Chanos chanos+ %ere dried to remo$e unpleasant odor. Dried !angus bile %as crushed using

mortal and pestle and a 0GmL $olume of distilled %ater added to the po%dered bile then use for experimentation.

0)r$/Bauer Ant)$)ot)c Test)ng7D)sc D)ffus)on Test)ng

After mixing the )ueller 8inton Agar %ith distilled %ater" it %as heated on a hot plate. ;ine Petri Dishes"

along %ith the inoculating loop and the )ueller 8inton Agar %ere sterili6ed in the autocla$e. The Petri dishes %ere

di$ided into three treatments ( %ater"bangus bile and Amoxicilin. Also" the Petri dishes %ere further labeled as

positi$e for Staphylococcus aureus and negati$e for 4scherichia coli. As the agar is still in its li/uid state" the

proponent /uickly added the E. coli"S. aureus and . aeru!inosabacteria on the petri dishes. 3ilter papers %ere

soaked in the !angus bile" %ater and dissol$ed amoxicilin. =hen the agar solidified" these treated paper discs %ere

then placed on each of the three replications. After ,1 hours" the 6one of inhibition of each treatment %as then

measured %ith the use of the Bernier Caliper.

Ant)/Ang)ogen)c Screen)ng

3irst and foremost" the proponents bought fresh Duck eggs from a professional handler. After purchasing

the duck eggs" they %ere /uickly incubated using the incubator of the school. T%el$e filter papers %ere then used"

three for each treatment. t %as dipped onto the said concentration. The soaked filter papers %ere then placed on the

CA) layer of the eggs and %ere sealed tightly to a$oid unnecessary entry of bacteria. After placing the filter paper

on the CA) of the eggs" the eggs %ere then incubated for 011 hours and %ere closely obser$ed. The Duck eggs

%ere then remo$ed after the said time period and the contents %ere closely obser$ed on a Petri Dish. Samples of the

blood $essels of the eggs %ere then taken and obser$ed under the microscope. The data gathered %as then analy6ed

and compared.

6RESENTATION8 ANAL9SIS AND INTER6RETATION OF DATA

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This part presents the data gathered through actual obser$ation and the changes that happened to the blood

$essels of the Duck eggs incubated for - days.

Resu%ts of the 0)r$/Bauer Test)ng

The results sho%ed that the extracts did ha$e anti(bacterial properties.Escherichia coliand Staphylococcus

aureusboth sho%ed positi$e results %hile Pseudomonas aureginosa did not.

The analysis of $ariance sho%ed the re2ection of the null hypothesis and accepted the alternati$e hypothesis

of ha$ing differences. 8o%e$er" Tukey test of difference in the mean of each group sho%ed that the !angus bile

mean has no significant difference %ith that of the mean of amoxicillin. t sho%ed that !angus bile has comparable

inhibitory effect %ith that of amoxicillin in inhibiting the gro%th of Staphylococcus aureus.

The gro%th inhibition of Pseudomonas aureginosa %ith !angus bile sho%ed a greater difference of 0.@1

mm. The analysis of $ariance sho%ed re2ection of the null hypothesis and accepted the alternati$e hypothesis of

ha$ing differences. 8o%e$er" Tukey test for difference in the mean of each group sho%ed that the each treatment is

significantly different from each other" %hich sho%ed that !angus bile is not effecti$e in inhibiting the gro%th of

Pseudomonas aurigonosa.

=hile the gro%th inhibition of 4. coli %ith !angus !ile under study. t is clear that the gro%th inhibitin of

!angus !ile and amoxicillin sho%ed a slight difference of G.-@ mm. The analysis of $ariance sho%ed the re2ection

of the null hypothesis and accepted the alternati$e hypothesis of ha$ing differences. 8o%e$er" Tukey test for

difference in the mean of each group sho%ed that the each !angus !ile and amoxicillin has no signicantly different

from each other" %hich sho%ed that !angus bile is effecti$e in inhibiting the gro%th of 4. coli

AN'IO'ENIC SCREENIN'

The test sho%ed the number of blood $essel formed on the choreoallantoic membrane assay of the fertile

duck egg. =ater %ith the greatest number of blood $essels of 1,. !angus bile %ith ,,.,,, and sim$astatin %ith

,0.GGG.

The analysis of $ariance sho%ed the re2ection of the null hypothesis and accepted the alternati$e hypothesis

of ha$ing differences. 8o%e$er" Tukey test for difference in the mean of each group sho%ed that the !angus bile

mean has no significant difference %ith that of the mean of sim$astatin. This only sho%ed that !angus bile has

comparable anti(angiogenic effect %ith that of sim$astatin in inhibiting the gro%th and formation of abnormal blood

$essels.

SU44AR9 OF FINDIN'S AND CONCLUSIONS

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5uptured $isible blood $essel formed

sho%ing branched $essels under the

naked eye treated %ith !angus !ile.


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This portion presents the summary of findings and conclusions made from the study and recommendations

gi$en by the researchers.

F)nd)ngs

The researchers started %ith the collection of the bangus bile . The collected samples %ere air dried for @

days. After drying" they soaked %ith )ethanol in 4rlenmeyer flasks in the science laboratory of the said school.

There %ere t%o screenings used. After each screening" results %ere again recorded despite the positi$e or negati$e

outcomes. The results on some of the screening %ere positi$e and some are negati$e.

Antibacterial screening %as conducted using three treatments. A pure culture of a gram positi$e and a gram

negati$e bacterium %ere cultures in the schoolOs science laboratory. 9lass %ares %ere then prepared for the

antibacterial testing. The testing %as then follo%ed using aseptic techni/ue. The Petri dishes used in the testing %ere

then put in the incubator in ,1 hours.


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B)$%)ogra#h

Internet

httpHMMen.%ikipedia.orgM%ikiMChorioallantoic_membrane

en.%ikipedia.orgM%ikiM:irby(!auer_antibiotic_testing

httpHMM%%%.medicinenet.comMnorfloxacin

en.%ikipedia.orgM%ikiMScreening

%%%.scientificcommons.orgMstatinManti(angiogenic

!ooks

)osbyKs Dictionary of Complementary and Alternati$e )edicine

The American 8eritage Dictionary of the 4nglish Language

)iller(:eane 4ncyclopedia and Dictionary of )edicine" ;ursing" and Allied 8ealth

=ebsterOs Dictionary of the 4nglish Language

9rolier 4ncyclopedia of :no%ledge

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http://www.scientificcommons.org/statin/anti-angiogenichttp://www.scientificcommons.org/statin/anti-angiogenic


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6)ctor)a%s

Soak)ng of

Bangus B)%e )n A%coho%

6re#ar)ng the

6etr) D)shes

Ster)%):at)on of 6etr) D)shes

6etr) D)shes )th Agar

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4ak)ng a ho%e )n the $%unt end of the egg Incu$at)ng the Treated Egg

ANAL9SIS OF ;ARIANCES

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ANTI-ANGIOGENIC AND ANTIBACTERIAL PROPERTY OF BANGUS BILE RESEARCH PAPER.doc

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