Introduction The synthesis and functionalization of silver nanoparticles AgNPshavebeenextensivelyinvestigatedinpastdecadesdueto theirremarkablepropertiesandpotentialapplications.Anumber
Trang 1j ou rn a l h o m e pa g e : w w w e l s e v i e r c o m / l o c a t e / c a r b p o l
Do Nu Bich Duyenb, Nguyen Quoc Hiena
a Research and Development Center for Radiation Technology, VINATOM 202A, Street 11, Linh Xuan Ward, Thu Duc District, Ho Chi Minh City, Vietnam
b Ho Chi Minh City University of Technology, 268 Ly Thuong Kiet, Ward 14, District 10, Ho Chi Minh City, Vietnam
a r t i c l e i n f o
Article history:
Received 21 March 2013
Received in revised form 17 October 2013
Accepted 20 October 2013
Available online 26 October 2013
Keywords:
Silver nanoparticles
Cotton fabrics
Chitosan
Antibacterial
␥- irradiation
a b s t r a c t
Silvernanoparticles(AgNPs)withdiameterabout12nmwereimmobilizedonthesurfaceofcotton fabricsby␥-irradiationoffabricsintheAgNO3chitosansolution.Effectsofabsorbeddose,concentration
ofAgNO3solutiononimmobilizationoftheAgNPsonfabricswereinvestigated.Theoptimaldosewas selectedtobe13.8kGyandthesuitableconcentrationofAgNO3was1.5mMin1%chitosansolution.The contentofAgNPsonfabricswasof1696±80mg/kgfortheseconditions.ThepresenceofAgNPsonfabrics wasconfirmedfromscanningelectronmicroscopy(SEM)imagesandXraydiffraction(XRD)patterns TheantibacterialactivityofAgNPs/cottonfabricsafter40washingsagainstStaphylococcusaureusand Escherichiacoliwasfoundtobe99.99%.Inaddition,theAgNPsfabricswereinnoxioustotheskin(k=0) after1,5,10,20,30and40washingsbyskin-irritationtestingtoanimal(rabbit)
© 2013 Elsevier Ltd All rights reserved
1 Introduction
The synthesis and functionalization of silver nanoparticles
(AgNPs)havebeenextensivelyinvestigatedinpastdecadesdueto
theirremarkablepropertiesandpotentialapplications.Anumber
ofreportsareavailableonthesynthesisofmetalnanoparticlesin
solutionbydifferentmethods(Hannan&Subbalaxmi,2011;Sáez
&Mason,2009;Szczepanowics,Stefanska,Socha,&Warszynski,
2010).Bolge,Dhole,andBhoraskar(2006)describedthe
synthe-sisofdispersed nanoparticulatesilverusing gamma-irradiation
Chen,Song,Liu,andFang(2007)alsosynthesizedsilver
nanopar-ticlesby␥-ray irradiationinacetic aqueoussolutioncontaining
chitosan.Thebroad-spectrumofantimicrobialpropertiesofAgNPs
encourageitsuseinbiomedicine,waterandairpurification,food
production, cosmetics, numerous household products (Li et al.,
2008;Mritunjai,Shinjini,Prasad,&Gambhir,2008).Becauseoftheir
effectiveantimicrobialpropertiesandlowtoxicitytoward
mam-maliancells,AgNPshavebecomeoneofthemostcommonlyused
nanomaterialsin consumerproducts(Choietal., 2008).Kokura
etal.(2010)provedthatsilvernanoparticleswereabletobeused
forpreservation ofcosmeticsagainstmixed bacteriaandmixed
fungi.AgNPscanbeimmobilizedonthefibers,bringingnew
prop-ertiestothefinaltextileproduct,especiallyforantibacterialeffect
Theantibacterialfabricscanbeusedtomakebandage,gauze,bed
∗ Corresponding author Tel.: +84 8 62829159; fax: +84 8 38975921.
E-mail address: truongthihanh05@yahoo.com (T.T Hanh).
sheets, surgicalclothes (Dubas, Kimlangdudsana,&Potiyaraj, 2006;Gupta,Bajpai,&Bajpai,2008).TheAgNPsinteractwiththe bacterialmembraneandareabletopenetrateinsidethecell.The mechanismsofinteractioninvolvedAgNPsattachedtobacterial cellmembranesincreasepermeabilityanddisturbrespiration.The AgNPs may catalyze reactionswith oxygen leading to reactive oxygenspecies(ROS)productionwhichcancauseDNAdamage, protein andcell membranebreakdown Thecatalytic silvercan destroybacteria’sdisulfidebondstocounteractthesynthesisof bacterialcell.AgNPsareoxidizedgeneratingsilverionsthatcan disrupt ATP production (adenosine triphosphate) to inhibit the adsorptionofphosphateofprotein(Jone&Hoek,2010)
TheAgNPsintheenvironmentaswastewatertreatmentplant effluentwereassociatedwithreducedsulfurfromorganicthiols groupsandinorganicsulfidestoAg2S.Sulfideexhibitsamuchlower toxicitythanotherformofAg(Kaegietal.,2011)
Inthisstudy,AgNPsimmobilizedoncottonfabricsbyinsitu synthesis.Silverionswerereducedtosilveratomsby␥-irradiation andsimultaneouslyimmobilizedonthefabrics.Thedurabilityof AgNPslinkedwithcottonfabricsandantibacterialeffectsaswellas skinirritationtestwerealsoinvestigatedafterrepeatedwashings
2 Experimental
2.1 Materials Cotton fabrics (100%) were provided by VICOTEX Company (Vietnam)withweighing120g/m2.Theywerewashedtoremove 0144-8617/$ – see front matter © 2013 Elsevier Ltd All rights reserved.
Trang 20.5m×0.5m,beforeusing.Shrimpshellchitinwassuppliedbya
factoryinVungTauprovince,Vietnam.Chitinwastreatedin3%
sodiumhydroxide(w/v)at100◦Cfor3hfordeproteinizationand
in3%hydrochloricacid(v/v)fordecalcification.Chitosanwitha
degreeofdeacetylation(DD%)about80%waspreparedby
deacety-lationofchitinin50%sodiumhydroxideat100◦Cfor1h.Thisvalue
wasdeterminedbasedonFT-IRspectraaccordingtothefollowing
equation(Brugnerottoetal.,2001):
A1320
A1420=0.3822+0.0313(100−DD%) (1)
where A1320, A1420 are absorbance of chitosan at 1320 and
1420cm−1,respectively
TheMwofchitosan(7.63×104)wasmeasuredbyanAgilent
1100 gel permeation chromatography (GPC; Agilent
Technolo-gies,USA)withdetectorRIG1362Aandthecolumnultrahydrogel
model250fromWaters(USA).Thestandardsforcalibrationofthe
columnswerepullulan(Mw780–380×103).Allotherchemicals,
includingsilver nitrate(AgNO3), (S)– lactic acid(90%),sodium
hydroxide(NaOH)wereofreagentgrade.Distiledwaterwasused
inallexperiments
2.2 PreparationofAgNPs/cottonfabricsby-irradiationof
cottonfabricsinAgNO3solution
Cottonfabricsabout60gafterwashingwereirradiatedin500ml
of0.5to5.0mMAgNO3solutionusingthestabilizerof1.0%chitosan
inthedoserangefrom5to20kGy.Thegamma-irradiationdosewas
determinedbyusingtheethanol-chlorobenzene(ECB)dosimetry
systemfrommeanvalueofabsorbeddosesofthreedosimetersat
30◦C.TheAgNPs/cottonfabricswerewashedaccordingtoa
mod-ifiedPN-ENISO105-C06:2010(AS1)(MarekKozEl ˙zbietaetal.,
2013)in ordertoassessresistancetothewashing process.The
maximumnumberofwashingwas40cycles Anapproximately
3gweightsamplewasplacedin150cm3 ofwashingliquorand
waswashedat40◦Cfor30min(onecycle).Theconcentrationof
theactivesurfaceagentwasequalto1g/dm3.Afterwards,samples
wererinsedwithwateranddried at40◦C for40min.Asimilar
procedurewasapplied for40 repeated washes Thecontent of
AgNPswasevaluatedbyinductivelycoupledplasmaatomic
emis-sionspectroscopy(ICP-AES),modelOptima5300DV(PerkinElmer)
after1,5,10,20,30and40washings
Allcollecteddatawereexpressedasmean±SE–standarderror,
inthisstudy.Thedifferencesbetweensamplevalueswereassessed
using two-tailed unpairedStudent’s t-tests The standard error
shouldbe<±5%ata95%confidencelevel,numberofsamplesthat
wereanalyzedperconditiontobethree(N=3)
2.3 CharacterizationoftheAgNPs/cottonfabrics
ThepresenceofAgNPsonfabricswasconfirmedbySEMimage,
usingaJSM-6480LVscanningelectronmicroscopywasoperated
at10kVandusedat1,300and5,500magnifications.Xray
diffrac-tionpatternsweremeasuredonaShimadzu5Adiffractometerwith
CuK␣radiation(35kVand25mA),scanningataspeedof0.5◦/min
from5–80◦(2).Thecrystallinesilvernanoparticlehasbeen
esti-matedbyusingDebye–Scherrerformula:
D=K·
whereK=0.89,isthewavelengthofX-ray(1.5405 ˚A),ˇisthefull
widthathalfmaximum(FWHM),isthediffractionangle(Bragg)
andDistheparticlediametersize(Baker,Pradhan,Pakstis,Pochan,
&Shah,2005)
ThemechanicalpropertiesweredeterminedusingaStrograph V10-C (Toyoseiki Co., Japan) testing instrument at a constant crossheadspeedof50mm/min.Fivespecimenswithadumbbells shapewerepreparedaccordingtoASTMD1882-Landwereused forthemeasurement
2.4 Antibacterialefficacyandskinirritationtestofthe AgNPs/cottonfabrics
Antibacterial properties of resultant fabrics were verified accordingtoAATCCTestMethod100–2004 against Staphylococ-cusaureus No 6538, a Gram-positive organismand Escherichia coliNo.10229,aGram-negativeorganism(AATCCTestmethod: 100–2004) The inoculum concentration of bacteria in a germ containingnutrientsolutionwas1×107 to2×107CFU/ml Test specimenswerecutin4.8±0.1cmdiametersandabsorbed1ml
ofinoculuminsterilePetriplates.Then,specimenswereplacedin thejarcontained100mlneutralizingsolution.Jarswereshaken vigorously for 1min, serial dilutionswere made From each of threesuitabledilutions,0.1mlliquidwasdrawnandtransferred
tonutrientagar,thenincubatedallplatesfor48hat37◦C.Percent reductionofbacteriawascalculatedfromthenumberof bacte-riarecoveredinthejarimmediatelyafterinoculationandthejar incubatedoverdesiredperiod
SkinirritationtestwascarriedoutbyInstituteofDrug Test-ingfollowingISO10993-10(2002).Thistestwascarriedouton grown and healthy rabbits weighing at least 2kg The rabbits usingfortheirritationtest wererearedat25◦C, withhumidity
of30–70%.TheAgNPs/cottonfabricswereappliedtoshavedskin about10cm×15cminthebackofrabbit.Theskinsituationofthe rabbitswasobservedafter12,24,48and72h
3 Results and discussion
3.1 PreparationofAgNPs/cottonfabrics TheAgNPs/cottonfabricswerepreparedby␥-irradiationof cot-tonfabricsinsilvernitratesolution.Theinfluentialfactorssuch
astheinitialAg+concentrationandabsorbeddoseonthe immobi-lizationofAgNPsontofabricswereinvestigatedbyICP-AES.During
␥-irradiationofcottonfabricsintheAgNO3solution,thehydrated electron(eaq −)and radicalofH• that weregeneratedby water radiolysiscanreduceAg+toAgasshowninEqs.(3)–(5)(Long,Wu,
&Chen,2007).Continuousreductionof theAg+ solutioncauses theaggregationofclustersintoAgNPs.In chitosansolution,the
•OHgroupscanreactwithchitosantoabstracthydrogenandform macromoleculefreeradicalsasEq.(6).Theseradicalscanreducethe clusterofAg+ionsformingtheclusterofAgatomsonfabric sur-face(Eq.(7)).Thebindingofsilverclustersbychitosanisachieved throughtheAg Obond(Chen,Wu,&Zeng,2005).Thereactions canbeproposedasfollows:
H2O(␥-rays)→•OH+e aq+•H+H2O2+H2+ (3)
[C6H11O4N]n(chitosan)+•OH
→[C6H11O4N]n−1[C6H10O4N]• +H2O (6)
Ag++[C6H11O4N]n−1[C6H10O4N]•
→Ag0-[C6H11O4N]n−1[C6H9O4N]+H+ (7)
Trang 3400
800
1200
1600
2000
20 15
10 5
0
Dose (kGy)
Fig 1. Relationship between the content of AgNPs on the cotton fabrics and
irradi-ation dose.
Theabsorbeddoseplaysanimportantrolein formationand
growthofAgNPsbyreductionofAg+toAg0atomby␥-irradiation
Atalowdose,nanoparticleshavenotappearedyet,while
nanopar-ticleswillbeformedatahigherdose.Inthisstudy,thepreparation
oftheAgNPs/cottonfabricsbyinsitusynthesisinwhichAg+ions
werereducedtoAgatomsandsimultaneouslydepositedoncotton
fabrics.TheinteractionbetweenfibersandmetallicAgNPswhich
werestabilized by chitosan resultsfromformation of chemical
bondbetweensilverandalcoholicgroupsofcottonandphysical
adsorptionofAgNPsonthefabricsurface(Perelshtein,Applerot,&
Perkas,2008).Fig.1clearlyindicatesthatthecontentofAgNPson
0 1000 2000 3000 4000 5000
6 5 4 3 2 1 0
AgNO3 Conc (mM)
Fig 2. Effect of AgNO 3 concentration on the content of AgNPs on the cotton fabrics.
thecottonfabricshaschangedinsignificantlyintherangeoflow dosesfrom4to10kGy,butincreasedathigherdoses.Amaximal valueoftheAgNPscontentonfabricswasfoundtobe1696mg/kg forthecottonfabricsin1.5mMAgNO3solutionirradiatedatthe doseof13.8±0.6kGy
TheeffectofAg+ concentrationonthecontentoftheAgNPs whenirradiationofthecottonfabricsinAgNO3solutionwasalso investigated.InFig.2,thecontentofAgNPsonthefabricsincreased
asusingtheinitialAg+ concentrations from0.5to5mMat the doseof13.8kGy.Inordertocontrolthereleaseofsilver nanoparti-clesintoenvironmentbutkeepingtheantibacterialpurposeafter washings,thecottonfabricswhichwereimmobilizedwithAgNPs
atthecontentof1696±80mg/kgcorrespondingtoAg+salt con-centrationof1.5mMwasselectedforfurtherinvestigation
Fig 3.SEM images of: original cotton fabrics (a) ×1300 and (c) ×5500; AgNPs cotton fabrics (b) ×1300 and (d) ×5500.
Trang 4Table 1
The tensile strength and elongation at break of cotton fibers against irradiation
doses.
Fig 4.UV–vis spectra of colloidal AgNPs from 1.5 mM AgNO 3 solution
correspond-ing to irradiation doses of: (a) 4.3 kGy, (b) 7.5 kGy, (c) 10.6 kGy, (d) 13.8 kGy, (e)
17.0 kGy, and (f) 20.3 kGy.
3.2 CharacterizationofAgNPs/cottonfabrics
Thepresenceofsilvernanoparticlesontofabricscanbeseen
bySEM images (Fig 3).The images in Fig.3a and c showthe
smoothstructureoftheoriginalcottonfabricsbeforecoatingwith
silvernanoparticles.Aftergamma-irradiationofcottonfabricsin
AgNO3solution,thenanoparticlesthatweredispersedonthe
sur-faceoftheAgNPs/fabricsdemonstratedtheimmobilizationofsilver
nanoparticlesonthefabrics(Fig.3b).Thehomogeneousdeposition
isobservedinahighermagnificationimage(Fig.3d).UV–vis
spec-trawererecordedforthecolloidalAgNPsfrom1.5mMAgNO3in
1.0%chitosansolutionirradiatedbygamma-irradiationwith
cot-tonfabrics,inthedoserangefrom4to20kGy(Fig.4).Thechitosan
concentrationwasmaintainedat1%solutionasthisprovided
suffi-cientchitosanforstabilizationoftheAgNPsandalsoallowedforan
acceptablesolutionviscosityat30◦C.TheUV–visabsorbance
spec-traofAgNPsdepictpeakmaximaintherangefrom416–420nm
withrespectiveincreaseinopticaldensity (OD)ingammadose
from4to13.8kGy.Theseresultsdemonstrateaproportionalyield
increaseofAgNPswithgamma-irradiationdosebelow13.8kGy
Above13.8kGy(17.0and20.3kGy),theopticaldensitychanged
insignificantly.Therefore,thedoseof13.8kGyisoptimalfor con-versionofthe1.5mMAgNO3toAgNPsin1%chitosansolutionat
30◦C
Mechanical properties of cotton fabrics were determined in thedoserange from0 to20kGy Theresultsshowed that ten-sile strength () and elongation at break (ε) of cotton fabrics slightly lower atthe dosesbelow 13.8kGy in comparison with non-irradiatedcottonfabrics(Table1).However,thesevalueswere muchlowerthanatthehigherdoses,sothattheirradiationdose wasselectedtobe13.8kGyforirradiationofcottonfabrics TheXRDpatternsinFig.5ashowspeaksat12◦and21◦,which can beattributed to thecrystalline structure of cellulose Four peaksat38.25◦,43.28◦,62.46◦ and76.32◦ inFig.5 assignedto diffractionsfromthe(111),(200),(220)and(311)planesof face-centeredcubicsilver,respectively.Thisresultisinagreementwith reportsfromotherauthorswhousedpolysaccharidesfor stabiliz-ingAgNPs(Phuet al.,2010;Staszewski,Kopyto,BeckerWrona, Dworak,&Kwarcinski,2008).Thefullwidthathalf-maximumof diffractionpeaks (111)in Fig.6 is employedtodeterminethe averagecrystallinesizeusingDebye–Scherrerasformula(2).The crystallinesizeofAgNPswascalculatedtobeabout12nm
3.3 Antibacterialeffectandskinirritationtest Thedurabilityofantibacterialsubstancelinkedwithfabricsis alsoanimportantfactorforusing.TheAgNPsmaybeimmobilized
onfabricsbythebondsoftheatomicsilverwithalcoholicgroups
incellulosestructureandchitosancappedAgNPs.Accordingtothe resultinTable2,theAgNPscontentonfabricsdecreasedafterthe firstwashingcyclebutchangedinsignificantlyfornextwashings ThesignsofAg/CF0,Ag/CF1,Ag/CF5,Ag/CF10,Ag/CF20,Ag/CF30and Ag/CF40weretheabbreviationsoftheAgNPs/cottonfabricsbefore washingandwashing1,5,10,20,30and40cycles,respectively Thelayer-by-layerlinksofthestabilizermoleculescoatingoutside wereformedfromelectrostaticsforce,sothattheywerebroken easilybyphysicalimpactordetergent.However,thebondsfrom thereactivecenterofcellulosetoAgNPscappingbychitosanmay
bedurable
ResultsinTable2alsodemonstratedthatantibacterialactivity wasinfluencedbythecontentofAgNPsonthefabrics.After40 washings,anamountofAgNPswasremainedonthefabricsofabout
858mg/kg,whichinhibitedthegrowthofS.aureusandE.colito
be99.95and99.98%,respectively.Guptaetal.(2008)alsoreported thattheAgNPscontentisakeyfactorincontrollingtheantibacterial activity.Inaddition,theAgNPsdiameterisalsoanimportantrole forantibacterialeffect.LeeandJeong(2005)provedthattheAgNPs sizeabout2–3nmhasabetterantibacterialeffectthan30nmone
Inthisstudy,theAgNPsdiameterabout12nmwasalsoeffective forantibacterialactivity
Testing skin-irritation was carried out for the fabric sam-ples, which weresynthesized insitu by ␥-irradiationof fabrics
in the 1.5mM AgNO3 solution at the dose of 13.8kGy The fabrics imbedded AgNPs were washed from 1, 5, 10, 20,
Table 2
Bacterial reduction of AgNPs cotton fabrics and numerical grades for testing skin irritation on the rabbit skin.
Trang 5Fig 5.XRD patterns of: (a) The cotton fabrics, and (b) AgNPs cotton fabrics.
Fig 6.Diffraction angle and the value FWHM at 2 = 38.25 o
30 and 40 cycles then were applied on the rabbit skin
The noxiousness on skin was determined from observing
the skin situation after contact times for 12, 24, 48 and
72hat25◦C Theerythemaandedemaontherabbit skinwere
recordedatthenumericalgradestobe0duringcoatingAgNPs
fabrics These results confirmed that AgNPs/cotton fabrics are
innoxioustoskinwithcoefficientofk=0(Table2).LeeandJeong
(2005)alsoreportedthattheAgNPsof2–3nmindiameterhavethe
numericalgradingtobe0forerythemaandedema.Theyincluded
AgNPswith2–3nmsizewasskin-innoxious
4 Conclusions
Thesilverionswerereducedeffectivelybygammairradiation and immobilizedonthecottonfabrics byinsitu synthesis.The AgNPscontentdepositedonthefabricswasof1696mg/kg,when thefabricsamplewasirradiatedin1.5mMAgNO3 and1.0% chi-tosansolutionatthedoseof13.8kGyat30◦C.TheAgNPswere confirmedbyUV–visspectraandSEMimages.Theaverage diam-eterofAgNPswasdeterminedbyXRDpatterntobeabout12nm ThepresenceofAgNPsonthefabricswasdepictedbySEMimages
Trang 620,30and40cycleswasabout99.99%forS.aureusandE.coli.The
AgNPs/cottonfabricswashingfrom1to40cycleswereinnoxious
toskin(k=0).Theseresultsdemonstratethatgammairradiation
ofcottonfabricsinthepresenceofaqueoussolutionofAgNO3and
chitosanisapromisingapproachforpreparationofstable,safeand
efficaciousantibacterialfabrics
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