activity against human pathogens Carica papaya peel mediated synthesis of silver nanoparticles and itsantibacterial Technology Journal of Applied Researchand

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Availableonlineatwww.sciencedirect.com

Journal of Applied Research and Technology

www.jart.ccadet.unam.mx JournalofAppliedResearchandTechnology15(2017)413–422

Original

Carica papaya peel mediated synthesis of silver nanoparticles and its antibacterial activity against human pathogens

J. Balavijayalakshmi

^∗

, V. Ramalakshmi

DepartmentofPhysics,PSGRKrishnammalCollegeforWomen,Coimbatore,Tamilnadu,India Received4August2016;accepted21March2017

Availableonline28September2017

Abstract

Metallicnanoparticlesaretraditionallysynthesizedbywetchemicaltechniques,inwhichthechemicalsusedarequiteoftentoxicandflammable.

Ripecaricapapayapeelisfound tobeasuitablesourceforgreensynthesisofsilvernanoparticles.Inthepresentwork,acosteffectiveand environmentalfriendlytechniqueforthegreensynthesisofsilvernanoparticlesfrom1mMsilvernitrate(AgNO3)solutionthroughtheextract ofripeCaricapapayapeelofvariousconcentrations(5ml,10ml,15ml,20ml,25ml)isdescribed.Thesynthesizedsilvernanoparticlesare characterizedbyusingtheUV–visabsorptionspectroscopy,FT-IR,XRD,SEMandTEM.Theformationofsilvernanoparticlesisconfirmedby surfaceplasmonresonance,determinedbyUV–visspectraat400–435nm.Theshiftintheabsorptionbandsandvariationinthecalculatedoptical bandgapsforthevariousconcentrationsofpapayapeelsextractsarealsoobserved.TheFT-IRspectrarevealthatanincreaseintheconcentration ofthepapayapeelextractshiftsthebandstohigherwavelengths.Theaveragecrystallitesizeforvariousconcentrationsofpapayapeelextractis observedfromXRDspectralanalysisandisfoundtobearound16–20nm,whichisingoodagreementwiththeTEManalysis.TheSEManalysis showsthesphericalstructureofthesilvernanoparticleswithsomeagglomerationforhigherconcentrationsofpapayapeelextract.Thesynthesized silvernanoparticlesshowgoodantibacterialactivityagainsthumanpathogenssuchasEscherichiacoliandStaphylococcusaureusandithasmany medicalapplications.

Keywords:Silvernanoparticles;UV–vis;FT-IR;XRD;SEM;Antibacterialactivity

1. Introduction

Therecentresearchbasedonnoblemetalnanoparticles(silver,gold,etc.)hasbecomemorefocussed,duetotheuniqueness oftheoptical,electrical,mechanical,magnetic,sizedependent, chemicalproperties of thesenanoparticles. The nanoparticles are very much different from those of the bulk materials (Forough & Fahadi, 2011). Due to these size dependent properties,metalnanoparticleshavesignificantapplicationsin electronics,optoelectronics,magnetic,biomedicalandinforma- tionstorage systems(Banerjee, Satapathy,Mukhopahayay,&

Das,2014).Anumberofapproachesareavailableforthesyn- thesisofmetalnanoparticles,suchaschemical,electrochemical,

∗Correspondingauthor.

E-mailaddress:[email protected] (J.Balavijayalakshmi).

PeerReviewundertheresponsibilityofUniversidadNacionalAutónomade México.

photochemical andradiation. The chemicalmethod produces toxic chemicals that may have an adverse effect in medical applications.Hencethereisaneedforbiosynthesisofnanopar- ticles. The biosynthesis of metal nanoparticles is a widely accepted technology and it is a kindof bottom up approach wherethemainmechanismbehindisreduction.Thenanoparti- clesproducedbythismethodaresafe,costeffectiveandmore environmentalfriendlywhencomparedtothechemicalmethods (Johnson&Prabu,2015).Comparetoothergenialbiological processes, the useof agriculturalwaste like peelextracts for theproductionofnanoparticlesisexpedient.Thefruitpeelsare especially easily available, efficient, affordable, eco-friendly, naturalandalsoveryrichinbioactivecompound.Thesebioac- tivecompoundscanbeusedasantioxidantsandantimicrobial agents,causingmostresearcherstoaimatidentifyinganeffi- cientwaytoextract thesebioactivecompoundsfromthefruit peels.

Silver nanoparticles (AgNPs) play a profound role in biosynthesis because of their distinctive properties such as

https://doi.org/10.1016/j.jart.2017.03.010

1665-6423/©2017UniversidadNacionalAutónomadeMéxico,CentrodeCienciasAplicadasyDesarrolloTecnológico.Thisisanopenaccessarticleunderthe CCBY-NC-NDlicense(http://creativecommons.org/licenses/by-nc-nd/4.0/).

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Fig.1.Caricapapayapeelextract.

goodconductivity,chemicalstability,catalyticandantibacterial activity(Vadlapudi,Kaladhar,Behara,Naidu,&Sujatha,2013).

Silvernanoparticleshavestronginhibitoryandbacterialeffects, which have been used to prevent and treat various diseases (Veerasamyetal.,2011).Silvernanoparticleshavebeenassorted into more than 200 consumer products, including clothing, medicinesandcosmeticsduetoitsmedicinalandantimicrobial properties. The papaya, papaw,or pawpaw isthe fruitof the plantCaricapapaya,theonlyspeciesinthegenusCaricaofthe plantfamilyCaricaceae.ItisnativetothetropicsoftheAmeri- casbuthasnowspreadallovertheworld.Thefreshpapayafruit hasattractivestrikingodours,highvitamincontent(i.e.,vitamin AandC)andhighfibrecontent.Theotherpartsofpapayasuch asskin,pulp,seeds,stemsandleavesalsocontainhighlevels ofproteins,vitaminsandvarietyofphytochemicals,including naturalphenolswhichareusedintheelaborationofcosmetics and medications (Aravind, Bhowmik, Duraivel, & Harish, 2013;Khan,Yadav,Srivastava,&Pal,2012;Kokila,Ramesh,

&Geetha,2016).

Here, we report facileand cost effective biogenic synthe- sisofAgNPsusingaqueousextractofcaricapapayapeeland investigationoftheantimicrobialofthesynthesizedAgNPs.

2. Materialsandmethods

Thecaricapapayafruitiscollectedfromthelocalmarketin Coimbatoreandthe chemicalsilvernitrate ispurchasedfrom Hi-media.

2.1. Preparationofpeelextract

Thepapayapeelsarewashedthoroughlywithdistilledwater toremovevisibledustparticlesandairdriedatroomtemperature toremovethewatermolecules.

About25gofdriedandfinelyslicedpeelsaretakenandadded into100mlofdistilledwaterandheatedfor30min.Thecrude extractisthenfilteredoutusingWhatmanNo.1filterpapertoget aclearbioextract(Fig.1).Silvernanoparticlesaresynthesized

fromsilvernitratebyusingaqueousbioextractofcaricapapaya peelsasreducingagent.

2.2. Activeconstituentsandsynthesismechanismsofsilver nanoparticles

Papaya peel has various photochemicals especially, phe- nolic compoundswhichhaveantioxidantproperties. The two important biologicallyactivecompoundsincaricapapaya are chymopapainandpapain,whichareextensivelyusedfordiges- tivedisorders(Huetetal.,2006).Caricapapayaderivedpapain, caricain,chymopain,andglycerineendopeptidasecanimprove acidicpHconditionsandpepsindegradation.Lipase,orCPLand ahydrolaseare the otheractivecompoundsof CaricaPapaya, which are firmly bonded to the water-insoluble fraction of crudepapain.Thisisconsideredasa“naturallyimmobilized”

biocatalyst(DeMaría,Sinisterra,Tsai,&Alcántara,2006;Kok- ilaetal.,2016).

Thepapayapeelextractsarerichinvitamins,aminoacids, carbohydrates, ␤-carotene, lycopeneandpolyphenols.Papaya peelextractsactsasanacesourceofriboflavinwhichhelpsto form the bound co-enzyme namely,Falvin MonoNucleotide (FMN)andflavinadeninedinucleotide(FAD),whichactsasa catalystforvariousreductionandoxidationreactions.Theother possiblemechanismsforthesynthesisofsilvernanoparticlesare nitratereductasesandshuttlequinineprocess(Durán,Marcato, Alves,DeSouza,&Esposito,2005;Kokilaetal.,2016).Inthis study,thesimilarmechanismiscarriedoutforthesynthesisof silvernanoparticlesusingtheextractofcaricapapaya.

2.3. Preparationofsilvernanoparticles

About5mloffilteredbiopeelextractistakenandaddedinto 1mMofpureaqueoussilvernitratesolutionandstirredforan hourforthereductionofsilvernitrateintosilvernanoparticles.

The reduction process Ag⁺ to Ag⁰ nanoparticlesis followed bythecolourchangeofthesolutionfromyellowtobrownish- yellow to deep browndepending on parametersstudied such as the extract concentration. The formation of brown colour

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Fig.2.Preparationofsilvernanoparticles.

confirmsthesynthesesofsilvernanoparticlesarecompleted.A similarmethodisfollowedforthepreparationofsilvernanopar- ticlesbyusingvariousconcentrationsofthecaricapapayapeel extract(Fig.2).

3. Resultsanddiscussion 3.1. Visibleobservation

Figure 3(a–e) shows the mixture of synthesized silver nanoparticlesusingpapayapeelextractsofvariousconcentra- tions(5ml,10ml,15ml,20mland25ml).Thecolour ofthe reactionmixtureafter1hfromtheinitiationofthereactionisdif- ferentfordifferentconcentrationsofcaricapapayapeelextract asevidencedfromFigure3.Thecolourofthereactionmixture forthe5mlconcentrationofpapayapeelextractshowsareddish browncolour,whichincreaseswithincreaseinthepeelextract concentrationfrom5mlto25ml.

Fig.3.Silvernanoparticlesusing(a)5ml,(b)10ml,(c)15ml,(d)20ml,(e) 25mlofcaricapapayapeelextractafter1hofincubation.

Fig.4.Visibleobservationofsilvernanoparticlesusing(a)5m,(b)10ml,(c) 15ml,(d)20ml,(e)25mlofpapayapeelextractafter24hofincubation.

Themaximumcolourintensity(deepbrown)isobservedfor the15mlconcentrationofpapayapeelextract.Theintensityof thecolourisfoundtodecrease(shallowbrown)withtheincrease intheextractconcentrationfrom20to25ml.Thedeepbrown colourofthereactionmixturesindicatestheformationofmore numberofsilvernanoparticlesfortheconcentrationsupto15ml ofpapayapeelextract.Similarly, theshallowbrowncolourof thereactionmixturesindicatetheformationoflessnumberof silvernanoparticlesfortheconcentrationsabove15mlofpapaya peelextract.Itisobservedthatthesynthesizedsilvernanoparti- clesareaggregatedforthehigherconcentrationofpapayapeel extract.Theaggregationofsilvernanoparticlesleadstodesta- bilizationofsilvernanoparticles(Mohapatra,Kaintura,Singh, Kuriakose,&Mohapatra,2015).

Figure4showsthereactionmixtureofsilvernanoparticles after24hofincubationfromtheinitiationofreactionatroom temperature.Itisobservedthatthecolourofallthereactionmix- turesbecomesmoreintense(deepbrown),therebyconfirming the formation of morenumber of silvernanoparticles, as the reactiontimeincreasesexponentially.

3.2. UV–visiblespectralanalysis

Figure5showstheUV–visibleabsorptionspectraofsilver nanoparticles with five different concentrations (5ml, 10ml, 15ml,20mland25ml)oftheaqueouspapayapeelextractat roomtemperatureafter24hofincubation.Theabsorption(or) surfaceplasmonresonance(SPR)bandfor5ml,10ml,15ml, 20mland25mlconcentrationsofpapayapeelmediatedsilver nanoparticlesareobservedat410nm,420nm,435nm,422nm, 418nm respectively.The SPRpeak withmaximumandmin- imum intensity is observed at 435nm and 418nm for 15ml and 25ml concentration of papaya peel extract respectively (Mohapatraetal.,2015).

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3.5

2.5

1.5

0.5

0.0

400 500 600 700

Wavelength(nm)

Absorbance

AgNps-peel

a b c d e

800 3.0

2.0

1.0

Fig.5.UV–visiblespectraofsilvernanoparticlesusing(a)5ml,(b)10ml,(c) 15ml,(d)20ml,(e)25mlofpapayapeelextract.

Table1

Opticalbandgapvalueofsilvernanoparticles.

Sample Opticalbandgap(eV)

5ml 4.9

10ml 4.7

15ml 4.6

20ml 4.7

25ml 4.8

FromFigure5,itisalsoobservedthattheabsorptionpeak wavelengthof AgNPs shiftstothe higherwavelength sideas theconcentrationofthepapayapeelextractincreasesfrom5to 15ml,correspondingtotheredshift.Thisredshiftindicatesa gradualincrementinthemeandiameterofthesilvernanoparti- cles.Itisfurtherobservedthat,astheconcentrationofpapaya peelextractincreasesto20and25ml,theabsorptionwavelength ofAgNPsshiftstolowerwavelengthsidecorrespondingtoblue shift.Thisblueshiftindicatesagradualreductioninthemean diameterofthesilvernanoparticles.

The opticalband gapvalue for the variousconcentrations of caricapapayapeelextracts arecalculatedandarelisted in Table1.Theopticalbandgapdecreasesastheconcentrationof papayapeelextractincreasesfrom5to15ml,whichconfirms that the size of the particle increasesand therebythe energy band gapdecreases. On furtherincrease inthe concentration of papaya peel extract to 20ml and 25ml, the optical band gapisfoundtoincreasetherebyconfirmingthedecreaseinthe sizeofthesilvernanoparticlesfortheseconcentrations,which couldalso beevidenced fromXRDanalysis(Ghobadi, 2013;

Kamat,Meisel,Weller,&Eychmuller,1996;Schön&Simon, 1995).

3.3. TimedependentUV–visiblespectralanalysis

The timedependentUV–visible spectraof silver nanopar- ticleswithdifferentconcentrations(5ml,10ml,15ml,20ml,

and 25ml) of papaya peel extract isshown inFigure 6. The UV–visible spectra of silver nanoparticles are recorded after timeintervalsof0min,15min,30min,45min,60minand24h from theinitiation ofreaction.It isobservedthat theincuba- tionperiodofthereactionmixtureshowsagradualincreasein absorbancespectrumwithsurfaceresonancePlasmonresonance band.Itisobservedthattheabsorptionpeakintensityislowerfor 0minofreactiontime,whichconfirmsthatlessnumberofsilver nanoparticlesisformedinthereactionmixture.Theabsorption peakintensitygraduallyincreaseswithincreaseinthereaction timefrom0minto60minforalltheconcentrationsofpapaya peelextract,whichindicatesanincreaseintheconcentrationof silver nanoparticlesinthereaction mixture(Mohapatraetal., 2015).Thehighestabsorptionpeakintensity isobservedafter 24hofincubationfromtheinitiationofthereaction,whichindi- catesthe completeformationof silvernanoparticlesinallthe reactionmixtures.

3.4. FT-IRanalysis

The FT-IR transmissionspectra of the carica papaya peel extract is shown in Figure 7. The absorption bands around 2924.09cm⁻¹,2862.36cm⁻¹and1458.18cm⁻¹,979.84cm⁻¹ may correspond to stretching or bending vibrations of C–H alkenes. The bands around 3726.47cm⁻¹and 918.12cm⁻¹ respectively are attributed to stretching and bending vibrations of carboxylic acids. The bands around 1188.15cm⁻¹ and 1658.78cm⁻¹ may be assigned to amide I and II N–H bending arising due tocarbonyl stretching and peptide link- agesof proteins respectively(Khalil,Ismail,El-Baghdady,&

Mohamed, 2014; Venkatesan, Subramanian, Tumala, & Vel- laichamy, 2014). From the FT-IR analysis, the presence of carboxyl andamidegroupsinthecaricapapaya peelextracts isconfirmed.Thesegroupsareresponsibleforthebio-reduction ofsilverionsintosilvernanoparticles.

Figure 8 shows the FT-IR spectra of carica papaya peel mediated silver nanoparticles (AgNPs), for various concentrations (5ml,10ml, 15ml,20mland25ml)of papaya peel extract.TheFT-IRspectrashowcharacteristicabsorptionbands around 700–750cm⁻¹, 950–980cm⁻¹, 1000–1150cm⁻¹, 1200–1300cm⁻¹, 1330–1370cm⁻¹, 1500–1550cm⁻¹, 1600–1650cm⁻¹ and 3100–3400cm⁻¹. The bands around 700–750cm⁻¹ may be attributed to stretching vibrations of C–Cl alkyl halides (Khalil et al., 2014). The observed bandsaround 950–980cm⁻¹ mayarise duetobendingvibra- tions of C–H group alkenes. The absorption band around 1000–1150cm⁻¹maybeassignedtoC–Ostretchingvibrations of carboxylic acids. The bands around 1200–1300cm⁻¹ are associated with N–H bend amines (Sivakumar, Nethradevi,

& Renganathan, 2012). The bands around 1330–1370cm⁻¹ may be due to C–N stretch amine group.The bands around 1500–1550cm⁻¹correspond toN–Oasymmetricstretchnitro compounds. The bands around 1600–1650 may correspond to stretching vibrations of primary and secondary amines (Khalil et al., 2014; Shanmugavadivu, Kuppusamy, & Ran- jithkumar, 2014; Sivakumar et al., 2012). The bands around

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5 ml

15 ml 20 ml

25 ml

10 ml 15 min

15 min

15 min 15 min 30 min

30 min

30 min 30 min

60 min

60 min 60 min 24 hr

24 hr

24 hr 24 hr 45 min

45 min

45 min 45 min 0 min

0 min

0 min 0 min 4

4

4 4

3

3 3

2

2 2

1

1 1

0

0 0 400

400

400 600

600

600 500

500

AbsorbanceAbsorbance

Wavelength(nm) Wavelength(nm)

Wavelength(nm)

Wavelength(nm) Wavelength(nm)

AbsorbanceAbsorbance

Absorbance

500

500 700

700

700 800

800

Fig.6.TimedependentUV–visiblespectraofsilvernanoparticlesusing(a)5ml,(b)10ml,(c)15ml,(d)20ml,(e)25mlconcentrationofpeelextract.

3100–3400cm⁻¹maybeduetoC Ostretchofcarboxylicacids (Khaliletal.,2014).

The FT-IR spectrum of silver nanoparticles is shown in Figure8,confirms thepresenceof carboxylgroupandamine groupinthereactionmixturebythepresenceoftheappropri- ate bands.Itis observedthat onincreasing the concentration ofpapayapeelextracts,thebandsareshiftedtohigherwave- length. It isfurther observedon comparing Figures7 and8, thatthebandsaround3100–3400cm⁻¹and1600–1650cm⁻¹, dueto carboxylic and aminegroups respectively, are shifted

tohigher wavelengthsduetothebindingof silverionswhen compared with respect to pure carica papaya peel extract and the depth of the band decreases (Khalil et al., 2014;

Sivakumaretal.,2012).Thesecarboxylandamidegroupsindi- cate the presence of secondaryamines, whichis a signature markerofproteins,thusconfirmingthebio-fabricationofsilver nanoparticles,bytheactionoftheproteinsorphytochemicals.

Therefore, itmay be confirmedthat, thesebio-molecules are responsibleforthecappingandstabilizationofthesynthesized nanoparticles. The FT-IR analysis suggests that proteins are

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100.0

99.6

99.4

3,000 2,000 1,000

3,500 2,500

Wavenumber(cm^-1)

Papaya peel

Transmittance(%)

1,500 4,000

99.8

Fig.7.FT-IRspectrumofpapayapeelextract.

115

b c d e AgNps-peel

a 110

105

100

95

90

4,000 3,500 2,500 2,000 1,500

Wavenumber(cm^-1)

% Transmittance

1,000 3,000

Fig.8.FT-IRspectraofsilvernanoparticlesusing(a)5ml,(b)10ml,(c)15ml, (d)20ml,(e)25mlofpapayapeelextract.

involvedinthecappingandstabilizationofthesynthesizedsilver nanoparticles.

3.5. XRDstructuralanalysis

The XRD patterns of bio-synthesized silver nanoparticles of various concentrations of papaya peel extract are shown in Figure 9. The XRD patterns clearly show that, the silver

900 800 700 600

200 100

30 40

Two theta(deg) (200)

(222)

(220) AgNps-peel

e c b a d

(311) (111)

Intensity(counts)

50 60 70 80

20 300 400 500

Fig.9.XRDspectraofsilvernanoparticlesusing(a)5ml,(b)25ml,(c)20ml, (d)10ml,(e)15mlofpapayapeelextract.

nanoparticles formed by the bio reduction of silver ions by papayapeelbrotharecrystallineinnature.XRDanalysisisused todeterminethephasedistribution,crystallinityandpurityof thesynthesizedsilvernanoparticles.

TheXRDpatternsofsilvernanoparticlesshowfivedistinct peaks in the spectrum of 2θ value ranging from 30^◦ to 80^◦. The peaksare indexedwithreferencetothe standardJCPDS card no. 04-783 for silver and are found to be (111), (200), (222),(220)and(311),correspondingtothecubicfaceofsilver nanoparticles (Saravanakumar, Ganesh,Jayaprakash, &Jang, 2015). The peaks observed around 27^◦ and 32^◦ may dueto the peel extract. TheseBraggs peak might haveresulted due tocappingagent,stabilizingthenanoparticles(Ibrahim,2015;

Roopanetal.,2013).Thecrystallitesizesofthesilvernanopar- ticlesusing variousconcentrations of papaya peelextract are calculatedusingtheDebye–Scherrer’sequation(Gnanajobitha, Rajeshkumar,Kannan,&Annadurai,2013).

D= Kλ

β cosθ

The averagecrystallite size ofsilver nanoparticlessynthe- sizedusing5ml,10ml,15ml,20mland25mlofpapayapeel extract are found to be 16.1nm, 16.3nm, 17.9nm, 17.8nm, 17.7nmrespectively.Itisobservedthattheaveragecrystallite size of the particlesincreaseswithincreaseinthe concentra- tionofpapayapeelextractfrom5mlto15mlandwithfurther increase inthe concentration to20ml and25ml,the crystal- lite size of the particlesdecreases gradually. Thedecrease in crystallitesizeoftheparticlecouldalsobeconfirmedfromthe UV–visiblespectralanalysis.Itisfurtherobservedthatthecrys- tallitesizeobtainedfromthesynthesizedsilvernanoparticlesis much smallerthanthat reportedbyKokilaetal.intheirarti- cle as28nmandthisreductionincrystallitesizeenhancethe propertiesofsilvernanoparticles.

3.6. SEManalysis

Scanningelectronmicroscopy(SEM)analysisisperformed for studying the surface morphology and shapes of silver nanoparticles. Figure 10(a–e)shows the SEMimages of various magnifications of silver nanoparticles synthesized using 5ml, 10ml,15ml, 20ml and 25ml concentrations of carica papaya peel broth respectively. It is observed that the silver nanoparticles are sphericalin shapeand the concentration of theextractaltersthesizeandshapeofnanoparticles.Theparti- clesareuniformlydistributedandnoaggregationsareobserved for 5–15ml of peel extract. But the particles get agglomerated as the concentration of papaya extract increases above 15mlascanbeevidencedfromFigure10(d–e).Theagglomer- ationofparticleleadstodestabilizationofsilvernanoparticles (Narmadha,Hemashenbagam,SathiyaVimal,&VasanthaRaj, 2013).

3.7. TEManalysis

The TEMimagesof silvernanoparticlessynthesizedusing aqueousextractofcaricapapayapeelareshowninFigure11.

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Fig.10.SEMimagesofsilvernanoparticlesusing(a)5ml,(b)10ml,(c)15ml, (d)20ml,(e)25mlofpapayapeelextract.

The concentration of the extract alters the shape and size of silvernanoparticlesinthesolution.Theseimagessuggestthat the morphology of the nanoparticles is mostly spherical in shape. A few agglomerated nanoparticles are also observed in someplaces, thereby indicating possible sedimentation at a later time. The average particle size of silver nanoparticles is found to be from 15nm to20nm. The results are in

good agreement withthe crystallitesize obtained fromXRD analysis.

3.8. Saedanalysis

Figure12 showsthe selectedareaelectron diffractionpat- tern(SAED)ofthesilvernanoparticlessynthesizedusingcarica papayapeelextractasreducingagent.Theringpatternsindicate that the particles are crystalline in nature. Each ring corre- spondstothe differentlatticeplanes obtained from theXRD analysis.

3.9. Antibacterialactivity

Antibacterial property of silver nanoparticles synthesized usingfivedifferentconcentrationsofpapayapeelextractagainst two different human pathogens Escherichia coli (gram neg- ative) and Staphylococcus aureus (gram positive) are shown in Figure 13. The labels S, D, P, 50 and 100 represent the inhibition zones for standard antibiotic, distilled water, pure carica papaya peel extract, 50␮l AgNPs and 100␮l AgNPs respectively.Itisobservedthat,theantimicrobialactivityofsil- vernanoparticleswithdifferentpapaya peelextracts isfound to be significant against both the bacterial strains, while the plant extract and distilled water does not show any such activity.

Theinhibitionzonesforthesilvernanoparticleswithdifferent concentrationsofpapayapeelextractarelistedinTable2.Ampi- cillinisusedasastandardantibiotictocomparetheresultswith thezoneobtainedusingsilvernanoparticles.Thezoneobtained using 100␮l of silver nanoparticles is greater than the zone formedusing50␮lofsilvernanoparticles.Itisobservedfrom Figure13that,increaseintheconcentrationofsilvernanopar- ticleswill resultintheformation ofawelldefined inhibition zone.ItisevidentfromTable2thattheinhibitionzonesforboth thebacterialstrainsdependontheconcentrationofpapayapeel extract.Agradualincrementinthediameter ofthe inhibition zoneisobservedfor silvernanoparticlespreparedusing5ml, 10mland15mlconcentrationsofpapayaextract.Thismaybe duetotheincreaseintheconcentrationofsilvernanoparticles inthereactionmixture.Further,increaseintheconcentrations of the peel extracts to 20ml and 25ml,the diameter of the inhibition zone obtained for E.coli andS. aureus strains get decreased.Thedecreaseinthediameteroftheinhibitionzone forhighconcentrationpeelextractsilvernanoparticlesmaybe duetothepresenceoflowconcentrationofsilvernanoparticles inthe reactionmixture. Itis confirmedthatthe antimicrobial activityofsilvernanoparticlesstronglydependsontheconcen- trationofthesilvernanoparticlespresentinthereactionmixture.

Theantimicrobialactivityforgramnegativebacteriaisgreater thanthatforgrampositivebacteriaforalltheconcentrationsof peelextracts(Banala,Nagati,&Karnati,2015;Ibrahim,2015;

Saravanakumaretal.,2017).Itisfurtherevidentthat thebest resultshavebeenachievedinourstudy,thatthezoneof inhibition hasbeen foundtobe 0.75cm (75mm) for E.coli and 0.65cm(65mm)forS.aureusfor100␮lisverymuchhigher

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50 nm

10 nm 5 nm

20 nm

Fig.11.TEMimagesofsilvernanoparticlesusingpapayapeelextract.

(311)

(200)

(111)

5 1/nm

(222) (220)

Fig.12.SAEDpatternsofsilvernanoparticlesusingpapayapeelextract.

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5 ml 10 ml 5 ml 10 ml

20 ml

25 ml 15 ml

15 ml

25 ml

a b

Fig.13.Antibacterialactivitiesofsilvernanoparticlesagainst(a)Escherichiacoli(b)Staphylococcusaureus.

Table2

Zoneofinhibitionofsilvernanoparticlesusingdifferentconcentrationsofpapayapeelextract.

Nameofhumanpathogens Standard(ampicillin)(30␮l) ZoneofinhibitionofAgNPswithdifferentconcentrationofextract

5ml 10ml 15ml 20ml 25ml

50␮l 100␮l 50␮l 100␮l 50␮l 100␮l 50␮l 100␮l 50␮l 100␮l

E.coli 14 0.4 0.45 0.5 0.55 0.65 0.75 0.55 0.65 0.45 0.5

S.aureus 15 0.3 0.45 0.5 0.55 0.55 0.65 0.5 0.55 0.45 0.5

thanthatreportedbyKokilaetal.as15mmforE.coliand10mm forS.aureusintheirstudy.

4. Conclusion

The present study describes the green synthesis of silver nanoparticlesusingvariousconcentrations(5ml,10ml,15ml, 20ml and25ml)of caricapapaya peel extract as the reduc- ingagent.Thesenanoparticlesareenvironmentalfriendlyand completelysafe.TheUVvisiblespectraofsilvernanoparticles showabsorptionpeaksaround400–435nmforthedifferentcon- centrationsof the papaya peel extract. It isobserved that on increasing the concentration of the peel extract from 5ml to 15ml,theabsorptionpeaksarebeingredshiftedandtheopti- calbandgapisfoundtodecrease,whichimpliesthat thereis anincreaseintheparticlesize.Itisfurtherobservedthat,with increaseinconcentrationofpeeltheextractto20mland25ml, theabsorptionpeaksarebeingblueshiftedandtheopticalband gapis increased,which impliesthat there will be adecrease intheparticlesize.Thesevariationsinparticlesizecouldalso beconfirmedfromXRDanalysis.TheFT-IRspectra confirm thepresenceofcarboxylgroupandaminegroupinthereaction

mixture byshowingtheappropriatebands,whicharerespon- sible forthe reductionof silverionsintosilvernanoparticles.

It is also observedthat withincrease inconcentration of the papaya peelextract from5mlto25ml,theFT-IR absorption bandsareshiftedtohigherwavelengthsbecauseofmoreinter- actionofsilverionswithreducingagents.XRDspectrareveal that the silver nanoparticlesare crystalline in nature and the averagecrystallitesizesarefoundtobefrom16nmto18nm whichisingoodagreementwithTEManalysis.SEMmicro- graphsshowthesphericalshapeof thenanoparticlesandalso theagglomerationforconcentrationsofextractabove15ml.The antibacterialactivityofsilvernanoparticleswastestedagainst twobacterialstrains,E.coliandS.aureus.Thezoneofinhibi- tionisfoundtobegreaterforgramnegativebacteriathangram positivebacteria.Themaximumzoneofinhibitionisobtained forthe15mlconcentrationofpeelextract.Thismayduetothe presenceofmorenumberofsilvernanoparticlesinthereaction mixturewhichcouldalsobeevidencedfromUV–visanalysis.

Itmaybeconcludedfromthedifferentcharacteristicanalyses thatthenanoparticlessynthesizedusing15mlconcentrationof extractaremoreeffectivecomparedtotheotherconcentrations ofpapayapeelextract.

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Conﬂictofinterest

Theauthorshavenoconflictsofinteresttodeclare.

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