TheinvestigationofantibacterialactivitiesofCloveessentialoilon

Escherichiacoli
.

*Note:Thismightbechangedoraddedbecausethelabinstructorisnotexplainingallofthe
procedure,theywantustodowhiletheyareteaching.Theprocedureandmaterialwouldbe
completelyfinishonthisfridayornextmonday*

Materials

Reagents
Escherichiacoli
Cloveoil
Tryptone
YeastExtraction
BacteriologicalAgar
NaCl

Equipments
LaboratoryAutoclave
500mLFalcon
250mLFalcon
Plates
LabBalance

Procedure

GeneralNotes
Warmthemediabeforestartingcellculturing.

PartI:Preparethemediaand
E.
Coli.
Thepreparationof750mLofBrothconsistof
7.5gramsofTryptone
3.75gramsofYeastextraction
7.5gramsofYeastextraction
5gramsofNaCl
750mLofroomtemperaturewater
750mLofBrothisseparatedinto250mLofBroth,and500mLofMediabyadding7.5
gramsofBacteriologicalAgar.
Bothof250mLofBrothand500mLofMediasterilizebyLaboratoryAutoclaveat121
degreecelsiusfor20minutes.
Transfer30mLtoeachplate,inwhichthisexperimentused24platesintotal.

Clove

Cloveisanevergreentreethathasapinkflowerbud,whichwillturnbrownwhendried
[1,2].Itlengthis0.5inchlongandthediameteris0.25inch[2].Additionally,cloveisconsidered
asaherbbecausepeopleuseitsoils,flowerbuds,leaves,andstemsformedicinalpurposes
[3].However,themostessentialpartiscloveoilbecauseithasacharacteristicofantibacterial,
antifungal,antiseptic,andantiviral[1].Therefore,itisusedtothreatendisorderssuchas
toothaches,indigestion,cough,asthma,stress,bloodimpurities,andetc.

[2/3/2016]
[1]
https://www.organicfacts.net/healthbenefits/essentialoils/healthbenefitsofcloveoil.html
[2]
http://www.whfoods.com/genpage.php?tname=foodspice&dbid=69
[3]
https://www.nlm.nih.gov/medlineplus/druginfo/natural/251.html

Additionally,accordingtoNaturalMedicinesComprehensiveDatabase,cloveoilcan
alsousedfordiarrhea.Therefore,bysupportingthisstatement,theresearchDevelopmentof
antibacterialstructuresandfilmsusingclovebudpowderhasclaimedthatitcanpreventing
microorganismgrowthin24hours.Moreover,fromSynergeticantibacterialefficacyofcold
nitrogenplasmaandcloveoilagainst
EscherichiacoliO
157:H7biofilmsonlettuceresearch,
theymentionedthatcloveoilcoulddamagebacterialcellwallandtheoutermembrane,which
causesbacteriatobedead.

However,TheresearchThespecificantibacterialactivityofliposomeencapsulated
Clove
oilanditsapplicationintofuhasstatedabouttheantibacterialactivitieson
Escherichia
coli
and
Staphylococcusaureus
inTofu.Theresearchhasusedtheliposomeformulationto
increasestabilityandefficiencyofcloveoil,buttheliposomethatencapsulatedcloveoilhasno
effectonE.coli.

InordertoinvestigatetheantibacterialactivitiesinCloveoil,weset4different
concentrationofcloveoil,0.5mg/mL,1mg/mL,2mg/mL,and4mg/mL,toperceivewhetherthe
cloveoilcaneliminateorinhibittheantibacterialactivitiesornot.Itissignificanttomakeit
explicit,becausemisunderstandingofthedrugusagecanleadtotheharmfulnessonuser.
SincethecloveoilcontainslargeamountofEugenolwhichmightcausesintestinaldiscomfortin
aprimary,and,thefailureofliverandkidneyinaseriouscase,whenusingcloveoilinan
inappropriateway.Moreover,Eugenolisalsoapotentialinhibitorofplateletactivityand
sometimescauseddiseasestobemoreserioussuchasChronicUrticaria.

[9/3/2016]
http://www.motherearthliving.com/plantprofile/eugenol.aspx
http://www.ncbi.nlm.nih.gov/pubmed/2767290

[16/3/2016]
https://books.google.co.th/books?id=rGQps9fQX1YC&pg=PA172&lpg=PA172&dq=eugenol+cau
tion&source=bl&ots=Bo0wLAj9m6&sig=3EbwrojhArgntOOjpejn7Oflqw&hl=en&sa=X&ved=0ah
UKEwiLze_UrcXLAhUCxI4KHQ78DIMQ6AEIJTAC#v=onepage&q=eugenol%20caution&f=false
http://www.ncbi.nlm.nih.gov/pubmed/2767290
https://pubchem.ncbi.nlm.nih.gov/compound/eugenol#section=MechanismofAction
http://www.livingnaturally.com/ns/DisplayMonograph.asp?StoreID=E500FDE33212420DA51DB
85BA6C3F8BA&DocID=bottomlineclove

Eugenol
E.Coli

Edatabase:

Developmentofantibacterialstructuresandfilmsusingclovebud
powder
Theuseofnaturalantimicrobial(AM)agentsinthefoodpackagingindustryhasbeenthefocusofprovidingasafe
andeffectivemethodinprotectinganddeliveringfoodtotheconsumer.Thisstudyinvestigatesnaturalmaterialssuch
asclovebudpowder(
Syzygiumaromaticum
)todetermineitsAMactivityagainstGramnegative
Escherichiacoli
(
E.
coli
)(DH5)andGrampositive
Listeriainnocua
(
L.innocua
)(LSPQ3284)and
Staphylococcusaureus
(
S.aureus
)
(5473)microorganisms.Thelowestconcentrationofclovebudpowderpreventingthegrowthofamicroorganism
after24hincubationwasconsideredastheminimuminhibitoryconcentration(MIC)andthevaluesoftheminimum
bactericidalconcentration(MBC)wasdeterminedaswell.Theclovebudpowderwaspreparedandcoatedonto
LDPEfilmstoevaluateitsAMactivityforfoodpackagingapplications.Itwasfoundthattheclovebudpowder
inhibitedthegrowthofthetestedmicroorganisms.AnLDPEfilmembeddedwiththeclovebudpowderbycoating
twiceshowedthebestAMeffectsagainst
E.coli
bacteria.

Thespecificantibacterialactivityofliposomeencapsulated
Clove
oiland
itsapplicationintofu

Inthisstudy,theantibacterialactivitiesof
Clove
oilandliposomeencapsulated
Clove
oilwereinvestigated.First,the
antibacterialactivityof
Clove
oildemonstratedthattheessentialoilexhibitedfavorableantimicrobialactivityforboth
Escherichiacoli
and
Staphylococcusaureus
.However,asetbackofusing
Clove
oilasadisinfectantisitslow

chemicalstability.Then
Clove
oilwasincorporatedintoaliposomeformulationtoincreaseitsstability.Theoptimal

polydispersityindex(PDI)(0.196),Zetapotential(24.5mV)andentrapmentefficiency(20.41%)ofliposomewere
obtainedattheconcentrationof
Clove
oilto5.0mg/mL.Inaddition,selectiveantimicrobialactivityfor
S.aureus
by
utilizingporeformingtoxins(PFTs)toactivate
Clove
oilreleasefromliposomewasobserved.Bycontrast,

liposomeencapsulated
Clove
oilhasnoeffecton
E.coli
thatdoesn'tsecretePFTsbecauseantimicrobialcomponent
can'treachbacteria.Gaschromatography(GC)assayfoundthatwhenliposomemet
S.aureus
thatsecretePFTs,
PFTswouldinsertintotheliposomemembranesandformpores,throughwhichtheencapsulated
Clove
oilwas

released.Besides,liposomeencapsulated
Clove
oilexhibitedefficientantimicrobialactivityfor
S.aureus
intofu.

Clove
oilcoulddamagetheintegrityofcellmembrane,leadingtothelossesofintracellular
constituentsandcelldeath.Controlledreleaseof
Clove
oilfromliposomewassuccessfully
achievedbyPFTssecretedfrom
S.aureus
.Liposomeencapsulated
Clove
oilcould

efficientlyinhibit
S.aureus
intofu.

Synergeticantibacterialefficacyofcoldnitrogenplasmaandcloveoil
against
Escherichiacoli
O157:H7biofilmsonlettuce
Thisstudyaimstoevaluatetheantimicrobialeffectsofcoldnitrogenplasma(CNP)andcloveoilagainst
Escherichia
coli
O157:H7(
E.coli
O157:H7)biofilmonlettuce.Bothcloveoil(1mg/mL,2mg/mLand4mg/mL)andCNP

(400600W)displayedsignificanteradicationeffecton
E.coli
O157:H7biofilmsinvitro(
p
<0.001).Subsequently,

theantibiofilmeffectofcombinedtreatmentwasstudiedaswell.Comparedwiththerespectivetreatment,combined
treatmentexhibitedremarkablesynergisticeffectoneradicating
E.coli
O157:H7biofilms.Theconfocallaserscanning
microscopy(CLSM)andscanningelectronmicroscopy(SEM)hadalsovisuallytestifiedthattheantibacterialeffects
ofcloveoilon
E.coli
O157:H7biofilms(invitroandonlettuce)wereenhancedbyCNPat400Wforshorttreatment
duration.Theresultsofsensoryevaluationindicatedthatcombinedtreatmenthasmildnegativeeffectonlettuce
quality.Moreover,thesynergeticantibacterialmechanismofcloveoilandCNPagainst
E.coli
O157:H7was

concludedasthattheycoulddamagethebacterialcellwallandtheoutermembrane,leadingtoleakageofcellular
components,suchasnucleicacidandATP.

CloveoilandCNPexhibitedsignificanteradicationeffecton
Escherichiacoli
O157:H7
biofilmsinvitro.TheantibiofilmactivityofcloveoilonlettucewasenhancedbyCNP

assistance.ThesynergeticantibiofilmmechanismofcloveoilandCNPwasexplored.

Antitermiticandantifungalactivitiesofeugenolanditscongenersfrom
theflowerbudsof
Syzgiumaromaticum
(clove)

Theinvestigatedessentialoilfrom
Syzygiumaromaticum
isapotenttermiticideandfungicide.
S.aromaticum
essentialoil:eugenolwasfoundasthemajorcompound.Eugenolwastestedandshowhighantitermiticand

antifungalactivities.Thecolveoilandeugenolhavepotentialforthedevelopmentofnaturalwoodpreservatives.

Biodegradationofwoodbyfungiandtermitesisrecognizedasoneofthemostseriousproblemsforwooden
structures.Inthisstudy,weanalyzedtheantitermiticandantifungalactivitiesofflowerbudsessentialoilfrom
Syzygiumaromaticum
anditsdominantconstituents.Essentialoilcomponentswereanalyzedbygas

chromatographyandthemajorcompoundofessentialoilwaseugenol(90.6%).Theresultsdemonstratedthatatthe
dosageof50g/g,cloveessentialoilhad100%mortalityto
Reticulitermeschinensis
aftertestingfor5d.Furthermore,
wefoundeugenolkilledalltermitesaftertestingfor3d,withanLC50valueof12.1g/g.Meanwhile,wealsofound
cloveessentialoilandeugenoldisplayedactivityagainstthreefungi,
Trameteshirsuta
,
Schizphylhlscommne
,and
Pycnoporussanguineus
.Wefoundeugenolwasprincipalantifungalconstituentofcloveoil.TheIC50valuesof
eugenolagainst
T.hirsuta
,
S.commne
,and
P.sanguineus
were83.6,116.1,and102.4g/mL,respectively.In
addition,comparisonsofthetermitemortalityandantifungalindicesofeugenolanditscongenersprovedthat

eugenolexhibitedthestrongestantitermiticandantifungalactivities.Thecloveoilandeugenolhavepotentialforthe
developmentofnaturalwoodpreservatives.