Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 121 (2014) 596–604

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Spectrochimica Acta Part A: Molecular andBiomolecular Spectroscopy

journal homepage: www.elsevier .com/locate /saa

Surface plasmon resonance optical sensor and antibacterial activitiesof biosynthesized silver nanoparticles

1386-1425/$ - see front matter � 2013 Elsevier B.V. All rights reserved.http://dx.doi.org/10.1016/j.saa.2013.11.019

⇑ Corresponding author. Tel.: +91 04542241685; fax: +91 04542 241122.E-mail address: ums10@yahoo.com (M. Umadevi).

M.R. Bindhu, M. Umadevi ⇑Department of Physics, Mother Teresa Women’s University, Kodaikanal 624101, Tamil Nadu, India

h i g h l i g h t s

� Green synthesis of Ag nanoparticlesusing Ananas comosus fruit extract asreducing agent.� Stable and spherical Ag nanoparticles

were prepared.� Shows size dependent antimicrobial

activity.� Act as very good copper and zinc

sensors.

g r a p h i c a l a b s t r a c t

a r t i c l e i n f o

Article history:Received 18 September 2013Received in revised form 3 November 2013Accepted 5 November 2013Available online 13 November 2013

Keywords:Silver nanoparticlesAnanas comosusSurface plasmon resonanceAntibacterial activityZinc sensorCopper sensor

a b s t r a c t

Silver nanoparticles were prepared using aqueous fruit extract of Ananas comosus as reducing agent.These silver nanoparticles showed surface plasmon peak at 439 nm. They were monodispersed andspherical in shape with an average particle size of 10 nm. The crystallinity of these nanoparticles was evi-dent from clear lattice fringes in the HRTEM images and bright circular spots in the SAED pattern. Theantibacterial activities of prepared nanoparticles were found to be size-dependent, the smaller nanopar-ticles showing more bactericidal effect. Aqueous Zn2+ and Cu4+ selectivity and sensitivity study of thisgreen synthesized nanoparticle was performed by optical sensor based surface plasmon resonance(SPR) at room temperature.

� 2013 Elsevier B.V. All rights reserved.

Introduction

Noble metal nanoparticles are used for the purification of waterwhich is one of the essential enablers of life on earth [1]. Water isone of the purest symbols of wealth, health, tranquility, beauty andoriginality. Pure water, which is free of toxic chemicals and patho-genic bacteria, is necessary for human health. Water and environ-ment get contaminated by the heavy metals due to industrial and

agricultural pollution. Heavy metals such as cadmium (Cd), Zinc(Zn), mercury (Hg), arsenic (As), silver (Ag), chromium (Cr), copper(Cu), iron (Fe), and the platinum group elements [2] in watercauses dangerous toxic effects on human beings. In particular thedetection of Cu4+ and Zn2+ in water, air and soil has been of greatresearch interest due to the important role of Zn and Cu plays inthe biological processes within the human body [3–5]. Zinc andcopper are essential at trace concentrations as heavy metal ionsor nutrients to maintain the metabolism of human body [5,6].Drinking water can be a source of Zn and Cu to humans as the re-sult of water treatment, usage of galvanized pipes, copper pipesand tanks in distribution systems. The beverages stored in metal

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containers, which are coated with zinc in order to resist rust, con-tain high levels of Zn. So, detection and control of Cu and Zn ions invarious media such as water, biological, environmental, medicaland industrial samples is very important. Keeping this in mind,detection of Zn and Cu concentration in water is explained in thepresent study.

A number of techniques have been developed for heavy metalions analysis, including Atomic absorption spectroscopy (AAS),Inductively coupled plasma mass spectrometry (ICPMS), Anodicstripping voltammetry, X-ray fluorescence spectrometry andmicroprobes. However, these techniques generally require expen-sive equipment, sample pretreatment, and/or a long measuringperiod. Thus, a simple, rapid, inexpensive, sensitive and selectivemethod is strongly needed. Optical sensor based on surface plas-mon resonance for detection of heavy metals in water is one ofthe most sensitive methods which will be advantageous over otherearlier techniques as it will be a simple, inexpensive and fast. In thepast years a number of nanoparticles based sensor have been re-ported [7–10]. Recently green synthesized silver nanoparticlesused in an optical sensor based on localized SPR for ammoniaand mercury detection was studied [11,12]. In this study, we havedesigned a silver nanoparticles based optical sensor for detectionof concentration of Cu4+ and Zn2+ ions in water.

Waterborne pathogens including helminthes, protozoa, fungi,bacteria, rickettsiae, viruses and prions, can cause many diseases[13]. In India 80% of the diseases are due to bacterial contaminationof drinking water. To protect the water purity, the removal or deac-tivation of pathogenic bacteria in water is very important. Silver isbeing used as a bactericide for water purification and also to pre-vent the buildup of bacteria and algae in water filters since morethan a decade. Antibacterial activities of silver nanoparticlesagainst various pathogens have also been established [14–16]. Inthe present study, the antibacterial assay was done on variouspathogenic bacteria like Escherichia coli and Proteus Mirabilis, whichare commonly found in water.

The physicochemical and optoelectronic properties of metalnanoparticles are based on specific characteristics such as size, dis-tribution and morphology [17]. Among the known nanoparticles,silver has been widely studied for its optical, spectroscopic, cata-lytic, antimicrobial and SERS properties. Due to these properties,silver nanoparticles have been broadly applied in consumer prod-ucts and industrial fields. In recent years, green synthesis ap-proaches of metal nanoparticles, using microorganisms andplants have received great attention to chemical and physicalmethods. Generally the reduction of silver nitrate using plant ex-tract was slow, but it had some advantage of producing stableand uniform size nanoparticles without using any additionalchemical stabilizers [18]. Recently, green synthesis of silver nano-particles using Hibiscus cannabinus, Solanum lycopersicums, Moringaoleifera, Murraya koenigii leaf, Citrus limon and Daucus carota havebeen reported [19–24]. Ananas comosus is a readily available fruitand it is a good source of water, carbohydrates, sugars, vitaminsA, C and carotene, beta [25]. It is one of the fruits with highest inthe flavonoid antioxidant Vitamin C. This antioxidant reduces theoxidative damage such as that caused by free radicals and chelat-ing metals [26]. It contains low amount of protein, fat, ash and fi-ber. There are three types of amino acids in A. comosus thatpromote exceptional health benefits through essential, semi-essential, and non-essential amino acids. Along with this, A. como-sus also contain bromelain, a protein-digesting enzyme that re-duces inflammation. Modified pineapple peel fiber was used toremove heavy metal ions in water through the reaction with succi-nic acid anhydride [27,28]. Bhosale et al. has reported the synthesisof silver and gold nanoparticles using A. comosus extract with kana-mycin A, and neomycin as stabilizing agents [29]. They preparedlarger nanoparticles with agglomeration. In the present study,

the synthesis and characterization of monodispersed smaller silvernanoparticles using fruit extract of A. comosus has been described.Here the size and aggregation of the nanoparticles were controlledwithout additional stabilizing agents. In the present study, the syn-thesis and characterization of biosynthesized silver nanoparticlesfor sensor and antibacterial activities have been described.

Experimental details

Material and methods

A. comosus fruit was collected from the local supermarket inKodaikanal, Tamilnadu, India. Silver nitrate, copper sulphate, ferricchloride, nickel nitrate, potassium chloride, cadmium acetate,manganous acetate, mercuric iodide, lithium hydroxide and zincacetate were obtained from Sigma Aldrich Chemicals. All glass-wares were properly washed with distilled water and dried inhot air oven before use.

Preparation of A. comosus extract

Fully riped A. comosus fruit, weighing 50 g was taken and cutinto fine pieces and were crushed into 100 ml distilled water in amixer grinder for extraction. The extract was then separated bycentrifugation at 1000 rpm for 10 min to remove insoluble frac-tions and macromolecules. The extract thus obtained was filteredand finally a light yellow extract was collected for furtherexperiments.

Synthesis of silver nanoparticles

5 ml of A. comosus extract was added to aqueous solution ofAgNO3 (3 mM) and stirred continuously for 5 min. The reactioncompleted slowly and it showed stable reddish brown color ofthe silver colloid (S1). Similarly by adding 10 and 15 ml of fruit ex-tract two more set of samples, henceforth called (S2) and (S3)respectively, were prepared. UV–vis spectra of these solutionswere recorded. Here the formation of silver nanoparticles startedwithin 20 min, and increased up to for 2 h. After 2 h, no color var-iation was observed up to 1 month, showing that the silver nano-particles prepared by this green synthesis method were verystable. Then the solutions were dried. The dried powders werecharacterized by X-ray diffraction (XRD), Fourier Transform Infra-red Radiation (FTIR), Transmission Electron Microscope (TEM)and Energy Dispersive X-ray Spectroscopy (EDX).

Characterization methods and instruments

The absorption spectra of the prepared nanoparticles were mea-sured using a Shimadzu spectrophotometer (UV 1700) in 300–800 nm range. X-Ray Diffraction analysis of the prepared nanopar-ticles was done using PANalytical X’pert – PRO diffractometer withCu Ka radiation operated at 40 kV/30 mA. FTIR measurementswere obtained on a Nexus 670 FTIR instrument with the sampleas KBr pellets. Transmission Electron Microscopic (TEM) analysiswas done using a JEOL JEM 2100 High Resolution TransmissionElectron Microscope equipped with an EDX attachment, operatingat 200 kV. For the detection of concentration of zinc in water usingoptical characteristics of silver nanoparticles, different concentra-tion of aqueous solution of zinc acetate dehydrate salt were pre-pared. This analyte solution was added to the prepared silvernanoparticle solution from 3 � 10�4 M to 12 � 10�4 M and stirredcontinuously for 1 min at room temperature. For the detection ofconcentration of copper in water, different concentration of aque-ous solution copper sulphate (2 mM to 7 mM) and of the same con-ditions were added into the silver nanoparticles. The antibacterial

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assay was done on various pathogenic bacteria like E. coli and P.Mirabilis. Nutrient agar medium was used to cultivate bacteria.Sterile paper disc of 3 mm diameter containing freshly prepared50 lg/ml silver nanoparticles were prepared. These plates wereincubated at 37 �C for 24 h. After 24 h the diameter of the growthinhibition zones were measured.

Results and discussion

Optical studies

The optical properties of prepared silver nanoparticles werecharacterized by UV–vis spectroscopy. Fig. 1 represents the opticalabsorption spectra of silver colloids S1, S2 and S3 obtained at dif-ferent concentration of fruit extract. Silver nanoparticles exhibitan intense absorption peak in the visible region due to the surfaceplasmon excitation. Surface plasmon resonance (SPR) absorptionband is observed due to the combined oscillation of free conduc-tion electrons of metal nanoparticles in resonance with light wave.The absorption spectrum of isolated spherical particles is charac-terized by the Mie resonance occurring at a frequency x0 suchthat: es (x0) = �2em, where es(x0) is the dielectric function of thesilver spherical particles and em is the dielectric function of thesurrounding medium [30]. The color variation of the S1, S2 andS3 has been shown in Fig. 1 (inset). When the frequency of theelectromagnetic field becomes resonant with the coherent electronmotion, a strong absorption takes place, which is the origin of theobserved color. The observed characteristic color variations of theprepared silver nanoparticles is changed from light orange to darkreddish brown as the concentration of fruit extract increases. Thisabsorption strongly depends on the particle size, dielectricmedium and chemical surroundings [31]. S1 was showing theformation of SPR band at 446 nm with broad band confirmingthe formation of varied size and shape nanoparticles and silver col-loid S3 showed SPR band at 439 nm with narrow peak indicatingthe formation of spherical nanoparticles. As the concentration offruit extract increased, the SPR bands of the prepared colloidsexhibited blue shift in the reaction medium. This result representsthat the diameter of the prepared silver nanoparticles decreasewith increasing concentration of the fruit extract, when electronsare donated to the particles [32]. The position of the plasmonabsorption peak depends on the particle size and shape and the

Fig. 1. Optical absorption spectra of silver nanoparticles at different concentrationof Ananas comosus fruit extract (inset: the Colour changes of the solution atdifferent concentration of Ananas comosus fruit extract in reaction system) (a, b, andc versus S1, S2 and S3 respectively).

adsorption of nucleophile or electrophile to the particle surface.In the case of aqueous silver nanoparticles, the Fermi level can floatupon chemisorption, depending on whether the substrate is nucle-ophilic and donates electron density into the particles or is electro-philic and withdraws electron density. Usually, a blue shift isassociated with a decrease in particle size or with the donationof electron density from the surface. It is well-known that adsorp-tion of the nucleophile to the particle surface bind the silver parti-cles and increases the Fermi level of the silver particle due to itsdonation of electron density to the particles [32].This directly cor-responds to a shift towards the blue end or red end, whereby smallsilver nanoparticle sizes would cause an absorption peak shift tosmaller wavelengths, higher frequency and energies [30]. At thesame time, the observed decrease of full-width at half-maximum(FWHM) value from 234 nm to 220 nm with increasing concentra-tion of fruit extract indicates that the size of the particle decreases[33]. The FWHM is reported to be helpful in understanding the par-ticle size and their distribution within the medium. The plasmonpeak and full width at half maxima depends on the extent of col-loid aggregation [34]. In the present case, the particle size of silvernanoparticles decreases with decreasing FWHM value. Thus fromthe results it can be concluded that the concentration of fruit extractplays an important role in the formation of silver nanoparticles. Thesymmetric nature of the SPR and the absence of peaks in the longerwavelength region indicated the absence of nanoparticle aggrega-tion. This was also confirmed by the TEM results. These nanoparticlesolutions were observed to be stable for a time period of one month.In the present case, the prepared silver colloidal nanoparticle intro-duced negative charge due to the biomolecules and thus repels theparticles away from each other, preventing them from aggregation.

FTIR studies

FTIR analysis was carried out to identify the possible reducingbiomolecules in the fruit extract responsible for the formation ofsilver nanoparticles and to identify the chemical change of thefunctional groups involved in bioreduction. Fig. 2(a and b) showsthat the FTIR spectrum of A. comosus fruit extract and the silvernanoparticles, respectively.

The peaks at 3417, 1640, 1019 and 801 cm�1 are assigned toOAH stretching, CH stretching, C@C ring stretching and CAC ringstretching of Vitamin C (ascorbic acid), respectively [35]. Thebroad and asymmetric feature of the C@C ring stretching of

Fig. 2. FTIR spectra of (a) Ananas comosus fruit extract and (b) S3.

Fig. 3. TEM micrograph of the S3 (a) the scale bar corresponds to 50 nm, (b) the scale bar corresponds to 20 nm, (c) the scale bar corresponds to 10 nm and (d) SAED pattern.

Fig. 4. TEM micrograph of the S1(a) the scale bar corresponds to 50 nm, (b) the scale bar corresponds to 20 nm, (c) the scale bar corresponds to 10 nm and (d) SAED pattern.

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Vitamin C mode at 1640 cm�1 in the spectrum of extract ap-peared at a well-defined peak of 1621 cm�1 in the spectrum ofsilver nanoparticles (S3). An asymmetric band at 1411 cm�1 inthe extract was observed in S3 at 1394 cm�1 assigned to the

ACAO stretching vibration modes of phytochemicals like watersoluble components such as phenolic compounds includingflavonoids and antioxidant vitamins [22,36]. It was possible thatthe antioxidant vitamin C (ascorbic acid) present in the extract,

Fig. 5. X-ray diffraction pattern of (a) Ananas comosus fruit extract and (b) S1 and(c) S3 (*due to Ananas comosus fruit extract).

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adsorbed on the surface of silver nanoparticles, could have led tothe reduction of Ag+ to Ag0 state.

The peak at 1098 cm�1 corresponds to CAO, CAN or CACstretching of amino acids. This region indicates the presence ofproteins [37]. A peak at 2369 cm�1 in the spectrum of extractwas assigned to NH� stretching of amines. This vibrational modemight be due to the presence of bromelain in the extract. Thisenzyme has the ability to separate all important amino acid bondsin protein. This represents that release of some protein compo-nents into the reaction medium may bind the nanoparticlesthrough cysteine residues in the proteins through hydrogen bondor it may cap the silver nanoparticles through electrostatic attrac-tion preventing from agglomeration and enhance the stability ofthe silver nanoparticles.

In this green synthesis method, silver nanoparticles wereformed by self assembling of the phytochemicals present in the ex-tract. The nucleation of silver nanoparticles might have formed bytransferring charge from ascorbic acid to Ag+ ions. Due to the Cou-lomb force, molecules of other phytochemicals, possibly of pro-teins got adsorbed towards the silver nanoparticles and formelectrostatic double layers and particle size was controlled. Dueto the Van der walls force of attraction, diffuse double layer was

Fig. 6. EDX gr

formed by the adsorption of various layers of phytochemicalsaround this electrostatic double layer. Steric or electrostatic barri-ers take place around the silver nanoparticle surface due to the var-iability of the molecule structures of phytochemicals in the extract.This may have thereby helped to cap the obtained silver nanopar-ticles, restrict the agglomeration and enhance the stability.

Morphological studies

Transmission electron microscopy (TEM) has been used to char-acterize the size, shape and morphologies of formed silver nano-particles. The size dependent morphology of the silvernanoparticles prepared using different fruit extract concentrationwas studied. The TEM images of the silver colloid S3 and S1 areshown in Figs. 3 and 4 respectively. The TEM image of S3 showingthe presence of mono dispersed and isotropic spherical nanoparti-cles with average size of 10 nm ranging from 7 to 13 nm size wereobserved at Fig. 3(a–d). This effect was in agreement with theshape of SPR bands of silver colloid S3. The present case shows thatthe presence of a large quantity of biomolecules in the extract,strong interaction between biomolecules in the fruit extract andsurface of nanoparticles was sufficient to the formation of sphericalnanoparticles preventing them from sintering. The presence oftwining in silver nanoparticles observed in Fig. 3(b). The twinnedparticles were identified by showing brightness in part of the par-ticles as compared to the other parts. Generally, twinning, the pla-nar defect is observed for face-centered cubic (fcc) structuredmetallic nanocrystals. Sharing of a common crystallographic planeby two subgrains give rise to twinning. The twinned silver nano-particles were further evidenced by the presence of (111) planein selected-area electron diffraction (SAED) pattern shown inFig. 3(d) (inset). Face-centered cubic (fcc) structured metallic nano-crystals have a tendency to nucleate and grow into twinned parti-cles with their surfaces bounded by lowest energy facets (111)[38].

TEM image of S1 was shown in Fig. 4(a–d). It shows the forma-tion of anisotropic nanocrystals and broad size distribution in therange of 12–40 nm. This result was also confirmed by UV–vis spec-tra of S1. The anisotropic nanostructures grow by a process involv-ing rapid reduction, assembly and room temperature sintering ofspherical nanoparticles. As the concentration of fruit extract de-creases less number of ascorbate ions are available to reduce silverions and thus forms large nanoparticles. The less specific surfacearea and surface area to volume ratio, and polycrystalline natureof the S1 also tends to larger particles. The formation of nonpoly-hedral shapes such as silver nanotriangles, hexagonal and rods,

aph of S3.

Fig. 7. Zone of inhibition of (a) S3 and (b) S1 for (i) E. coli and (ii) Proteus Mirabilis,respectively and (c) antibacterial activity of S3 and S1.

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which are a result in stable morphologies, might be due to the min-imization of surface energy by the low-index crystal planes [39]. Aface-centered cubic (fcc) lattice of noble metals possesses differentsurface energies for different crystal planes. The excess free energyper unit area for a particular crystallographic face is surface energy.It determines the faceting and crystal growth for nanoparticles.The selected–area electron diffraction (SAED) pattern of S1wasshown in Fig. 4(d). The observed SAED pattern with bright circularrings corresponding to the (111), (200), (220) and (311) planesshow the crystalline (fcc) nature of the nanoparticles.

Structural studies

The crystalline nature of the prepared silver nanoparticles wasconfirmed with X-ray diffraction (XRD) analysis. The XRD patternfor the dried powder of A. comosus was shown in Fig. 5(a). The ob-served peaks at 2h values 28.5�, 40.8� and 50.9� indicates the pres-ence of ascorbic acid (JCPDS 22-1560) in the A. comosus extract.

Fig. 5(b and c) shows the XRD patterns of the prepared silvernanoparticles. The diffraction peaks were observed at 38.3�,44.4�. 64.6� and 77.4� in the 2h range 20–80� can be indexed tothe (111), (200), (220) and (311) reflection planes of face cen-tered cubic (fcc) structure of metallic silver, respectively (JCPDS04-0783). No peaks of crystallographic impurities in the samplehave been found. Generally, the breadth of a specific phase ofmaterial is directly proportional to the mean crystallite size of thatmaterial. The sharper peaks indicating the crystallite size was largewhereas broader peaks are indicative of small crystallite materials.Based on our XRD data, broadening of diffraction peaks was ob-tained with increasing fruit extract concentration. This reveals thatas the concentration of fruit extract increases, the average crystal-lite size decreases. The average size of the prepared silver nanopar-ticles was determined from the Debye–Scherrer Eq. (1) by usingthe width of the (111) Bragg’s reflection.

D ¼ kkb cos h

ð1Þ

k is the Scherrer constant (k = 0.94), k is the wavelength of the X-ray, b is the FWHM of the peak and h is the half of the Bragg angle.

The calculated average particle size was found to be 16 nm forS1 and 9 nm for S3.This indicates that the particle size decreasedwith the increase in concentration of the fruit extract. The calcu-lated cell volume for S1 and S3 was 68.02 Å3 and 67.69 Å3, respec-tively. The lattice parameter for S1 and S3 has been found out to be4.082 Å and 4.075 Å, respectively, It was in very good agreementwith the standard value 4.086 Å (JCPDS 04-0783). The ratio be-tween the intensity of the (200) and (111) diffraction peaks was0.28 for S1 and 0.27 for S3, which was lower than the conventionalbulk intensity ratio 0.52, suggesting that the (111) plane was thepredominant orientation as confirmed by high-resolution TEMmeasurements.

In order to determine the type and property of a material, sur-face area to volume ratio (SA:V) or Specific Surface Area (SSA) canbe calculated. Each material has its own SSA. The SSA is of partic-ular importance in reactivity. It gives the rate at which the reactionwill proceed. SSA can be calculated using the following equation:

SSA ¼ SApartVpart� density

ð2Þ

where SApart is particle surface area, Vpart is particle volume anddensity of silver is 10.5 g/cm3. The values of SA: V ratio and SSAof S1 was 0.38 and 37 m2/g respectively. For S3, the values of SA:V ratio and SSA was 0.66 and 64 m2/g respectively. The SA: V ratioand SSA was high for the silver nanoparticles synthesized usinghigher concentration of fruit extract.

Crystallinity was evaluated by comparing the crystalline sizeobtained by XRD to TEM particle size determination. Crystallinityindex of the prepared silver nanoparticles was evaluated by the fol-lowing equation:

Icry ¼ DpðSEM;TEMÞDcryðXRDÞ ðIcry P 1:00Þ ð3Þ

where Icry is the crystallinity index, Dp is the obtained particle sizefrom either TEM or SEM analysis and Dcry is the particle size ob-tained from Scherrer equation. If Icry is close to 1, then it is mono-crystalline whereas if it is greater than 1, it is a polycrystalline innature [40]. The calculated values of crystallinity index of S1 andS3 were �2.25 and �1.11 respectively. This indicates that S3 showsmonocrystalline whereas S1 shows polycrystalline nature.

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Fig. 6 shows the energy dispersive X-ray analysis (EDX) of S3. Itconfirms the formation of silver nanoparticles and reveals highercounts at 3 keV due to silver nanoparticles. It was also due to sur-face plasmon resonance of metallic silver nanocrystals as shown inoptical absorption spectra.

Antibacterial activity

In this present case, the antibacterial activity of prepared silvernanoparticles was deliberated against Gram negative pathogensE. coli and P. Mirabilis. The antibacterial effect of silver nanoparti-cles on microorganisms may be held through the electrostaticattraction of positive charged silver and negative charged cell sur-face of microorganism. Fig. 7(a and b) shows the zone of inhibitionof silver nanoparticles S3 and S1 against E. coli and P. Mirabilis.Fig. 7(c) shows the antibacterial activity of silver nanoparticlesproduced from (S3) and (S1) against pathogens.

The antibacterial activity of S3 was more than S1 because of itslarger specific surface area, smaller size and spherical shape.Depending on the surface area available for interaction, silvernanoparticles can bind bacteria and disturb its permeability andrespiration function. Smaller particles having larger surface areacan increase the ability to penetrate cell membrane and give morebactericidal effect than the larger particles [41]. Thus, the antibac-terial activity of prepared silver nanoparticles are influenced by thesize and morphology of the particles, and the effect of antibacterialactivity increases with decreasing size of silver nanoparticles. Theprepared silver nanoparticles function as a good antibacterial agentwas significant in biomedical applications as well as in the removalor inactivation of pathogenic bacteria in water.

Detection of zinc and copper ions in water

To investigate the interaction of prepared AgNPs (S3) with var-ious alkali metal (Li+, K+, Fe3+) and transition metal ions (Ni2+, Mn2+,Cu4+, Zn2+, Hg2+, Cd2+), 0.3 ml of (3 mM) salts of these metals wereadded into 3 ml of AgNPs by drop by drop and stirred for 2 min.The photographs (Fig. 8 (inset)) and UV–vis spectra (Fig. 8) ofAgNPs were taken immediately after addition of metal ions, after2 min of interaction. Upon addition of Zn2+, the reddish brown col-or of the solution changed to colorless. Similarly the reddish browncolor of the AgNPs solution changed to light gray. Based on Fig. 8, it

Fig. 8. UV–vis absorption spectrum and photographs (inset) of S3 with variousheavy metal ions.

was observed that the intensity of the SPR bands got reduced andred shifted for all metal ions as compared to that of the AgNPs. Itwas observed that there was no prominent SPR peak for Zn2+ anda secondary peak for Cu4+, indicating the prepared AgNPs weresensitive and selective towards Zn2+ and Cu4+.

The activity of S3 in the presence of Zn2+ was studied by addingvarious concentration of heavy metal ion (Zn2+) from 3 � 10�4 to12 � 10�4 M in water. Fig. 9(a) shows the UV–vis absorption spec-trum for activity of S3 as zinc sensor. The prepared AgNPs (S3)solution did not exhibit color change from reddish brown as wellas change in absorption spectra up to the addition of 2 � 10�4 MZn2+. This indicates that no aggregation effect has taken place.The addition of 3 � 10�4 to 12 � 10�4 M Zn2+ to the silver nanopar-ticles solution causes color changes from reddish brown to color-less, were observed as shown in Fig. 9(a) inset. The addition of3 � 10�4 M Zn2+ causes immediate reduction in the intensity ofsurface plasmon peak at 444 nm. When zinc acetate added to theprepared nanoparticles, newly produced zinc atoms stronglybonded on the silver surface and moved the capping biomoleculesin the extract, away from the surface, which could be accounted forthe broadening and slight blue shift of the SPR band of silvernanoparticles. The optical absorbance intensity reduces and band

Fig. 9. (a) UV–vis spectra and color changes (inset) of S3 as a function of variousconcentrations of Zn2+ ions (3 � 10�4 to 12 � 10�4 M) and (b) plot of absorbance at444 nm versus Zn2+ concentration (3 � 10�4 to 12 � 10�4 M) with a correlationfactor R2 = 0.959.

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broadens gradually with the increase of concentration of Zn2+, sug-gesting that aggregation had taken place due to the reduction ofinterparticle distance in the aggregates less than about the averageparticle diameter. Upon addition of the 3 � 10�4 M to 12 � 10�4 MZn2+ to S3, color changes from reddish brown to colorless were ob-served, and after the addition of 12 � 10�4 M Zn2+ absorbancechanges were negligible. This shows that the formation of stableaggregates. So the concentration of zinc was limited to 10 �10�4 M with notable color and surface plasmon resonance band.Fig. 9(b) shows a good linear correlation (y = 0.036x + 0.965,R2 = 0.959) between the absorbance (DA) versus concentration ofZn2+, and the sensitivity of the system towards analyte concentra-tion was found to be 0.036/�10�4 M as measured from the plot ofabsorbance (DA) versus concentration of Zn2+.

Fig. 10(a) shows the UV–vis absorption spectrum for activity ofsilver nanoparticles as copper sensor. When 1 mM copper sulphateadded to S3, the color of the nanoparticle solution did notchange, only the addition of 2 mM copper sulphate causes colorchanges. This indicates that the analyte directed aggregation ofnanoparticles taken place only after the addition of 2 mM Cu4+.With increasing analyte concentration from 2 mM to 7 mM, the

Fig. 10. (a) UV–vis spectra and color changes (inset) of S3 with Cu4+ ions (2–7 mM)and (b) plot of absorbance changes (A780 nm/A478 nm) versus Cu4+ concentration (2–7 mM) with a correlation factor R2 = 0.967.

intensity of the SPR peak centered at 478 nm decreases graduallywith band broadening and a new SPR band appears at 780 nm withincreasing intensity. The main reason for SPR broadening is elec-tron surface scattering which may be enhanced for very small clus-ters [42]. The observed band broadening of the surface plasmonband reveals that the copper atoms may bind on the silver surface.The presence of new SPR peak was due to the adsorption of mole-cules causing changes in dielectric environment around a nanopar-ticle or agglomerated particles. When copper sulphate is added toS3, Cu4+ ions interact with the biomolecules in the extract on thesurface of the nanoparticles, forming bonds among nanoparticleswith Cu4+ ions performing as link for binding sites of biomoleculesand eliminating it away from the surface of the nanoparticle sur-face, in that way aggregation of nanoparticles has taken place.The addition of 2 mM to 7 mM Cu4+ to S3 causes color changesfrom reddish brown to light blue shown in Fig. 10(a) inset. The ob-served color changes and absorbance changes after the addition of7 mM Cu4+ were negligible, suggesting the formation of stableaggregates. So the concentration of copper was limited to 7 mMwith notable color and surface plasmon resonance band. Thelinear variation of absorbance (A780 nm/A478 nm) changes and theconcentration of Cu4+ over the range from 2 mM to 7 mM areshown in Fig. 10(b). This plot can be fit by a linear equationy = 0.605x�0.202, R2 = 0.967 and the sensitivity of the system to-wards analyte concentration was found to be 0.605/mM. Applica-tions of nanoparticle sensors by the aggregation of smallparticles were useful because aggregates with multiple particlesyield large enhancements due to the enormous electromagneticfield that coherently interfere at the junction site between the par-ticles. This zinc and copper sensor based on surface plasmon opti-cal sensor can be used in environmental monitoring especially inwater purification.

Conclusion

An ecofriendly method of obtaining spherical silver nanoparti-cles with average size of 10 nm has been synthesized, using A. com-osus as reducing agent. The prepared silver nanoparticles are stablefor one month without aggregation. Crystalline nature of the nano-particles was evident from bright spots in the SAED pattern, andpeaks in the XRD pattern. The prepared silver nanoparticles revealgood antimicrobial activity against Gram negative pathogens E. coliand P. Mirabilis, which are found in water. So the prepared nano-particles have found applications in biomedical and water purifica-tion processes for inhibiting the growth of bacteria. The preparednanoparticles were used for sensing ions of heavy metals likeZn2+ and Cu4+ in water using a SPR optical sensor. A strategy ex-plained in the present study is detectable by using naked eye orUV–vis spectrophotometer. The optical absorption spectrum ofthe colloidal suspension before and after addition of metal ions isa good pointer to detect the concentration of the heavy metal ions.For continuous monitoring and real time information, this Zn andCu sensor based on surface plasmon optical sensor is preferred,and it can be used in environmental monitoring, especially inwater purification.

Acknowledgements

The authors are thankful to DST-CURIE New Delhi, UGC-DAECSR Indore for financial assistance.

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