Accepted Manuscript Biogenic Synthesis of Multi-applicative Silver Nanoparticles by using Ziziphus Jujuba Leaf Extract N.L. Gavade, A.N. Kadam, M.B. Suwarnkar, V.P. Ghodake, K.M. Garadkar PII: DOI: Reference:

S1386-1425(14)01476-0 http://dx.doi.org/10.1016/j.saa.2014.09.118 SAA 12794

To appear in:

Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy

Received Date: Revised Date: Accepted Date:

11 June 2014 27 July 2014 28 September 2014

Please cite this article as: N.L. Gavade, A.N. Kadam, M.B. Suwarnkar, V.P. Ghodake, K.M. Garadkar, Biogenic Synthesis of Multi-applicative Silver Nanoparticles by using Ziziphus Jujuba Leaf Extract, Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy (2014), doi: http://dx.doi.org/10.1016/j.saa.2014.09.118

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Biogenic Synthesis of Multi-applicative Silver Nanoparticles by using Ziziphus Jujuba Leaf Extract N. L. Gavadea, A. N. Kadama, M. B. Suwarnkara, V. P. Ghodakeb, K. M. Garadkara* a Nanomaterials Research Laboratory, Department of Chemistry, Shivaji University, Kolhapur - 416004, M. S. India. b

Deparetment of Biotechnology Engineering, KIT’s College of Engineering, Kolhapur - 416004, M. S. India.

______________________________________________________________________________ *Corresponding author: K. M. Garadkar, Email: [email protected] Tel: +91-0231-260916; Fax: +91-0231-2692333.

Abstract Herein, we are reporting for the first time one step biogenic synthesis of silver

nanoparticles (AgNPs) at room temperature by using Ziziphus Jujuba leaf extract as a reducing and stabilizing agent. The process of nanoparticles preparation is green, rapid, environmentally benign and cost effective. The synthesized AgNPs were characterized by means of UV-Vis., XRD, FT-IR, TEM, DLS and Zeta potential. The absorption band centered at λ

max

434 nm in

UV-Vis. reflects surface plasmon resonance (SPR) of AgNPs. XRD analysis revealed, that biosynthesized AgNPs are crystalline in nature with the face centered cubic structure. FT-IR analysis indicates that nanoparticles were capped with the leaf extract. TEM images shows the synthesized nanoparticles are having different shapes with 20 to 30 nm size. The data obtained from DLS that support the hydrodynamic size of 28 nm. Zeta potential of –26.4 mV indicates that the nanoparticles were highly stable in colloidal state. The effect of pH, quantity of leaf extract and concentrations of AgNO3 were also studied to attend control over the particle size 1

and stability. The synthesized AgNPs shows highly efficient catalytic activity towards the reduction of anthropogenic pollutant 4-nitrophenol (4-NP) and Methylene Blue (MB) for environmental protection. Synthesized AgNPs also exhibited good antimicrobial activity against Escherichia coli. Keywords: Ag Nanoparticles; Biogenic synthesis; Ziziphus Jujuba leaf extract; Antimicrobial activity; Reduction of 4-nitrophenol; XRD analysis. 1. Introduction The demands of nanomaterials are increasing day by day due to their distinctive physicochemical properties as compared to bulk counterparts [1]. In particular noble metal nanoparticles have wide applications in various fields such as medicine, electronics, energy and catalysis [2-4]. The synthetic route for the preparation of nanostructure materials are more important in the current research due to their physical properties which can be tailored for a specific application by controlling their size and morphology [5, 6]. Generally metal nanoparticles have been prepared by physical and chemical methods [7]. However, due to their negative impact, there is an urgent need to replace the chemical method with clean, non toxic and environmentally acceptable biological method. From the literature survey, plenty of reports are available on the synthesis of silver nanoparticles by bio-reduction method based on bacteria, fungi, microorganisms and plant extract [8-12]. The use of plant extract for the synthesis of nanomaterials could be more advantageous as it does not involve

elaborative process and recoil from maintenance of

microbial cell culture [13]. From the literature survey, it is found that the rate of metal ion reduction by plant extract is much more faster than the microorganism [14]. The biomolecules in 2

plant extract like proteins, phenols and flavonoids plays crucial role for the reduction of metal ions [15]. Silver nanoparticles have gained much popularity on account of their broad spectrum of antimicrobial and surface plasmon resonance (SPR) effect [16-20]. AgNPs have been synthesized by variety of plants extract that includes Aloevera [21], Cassia auriculata, Mentha piperita, Ocimum tenuif lorum, Mentha piperita [22]. A very few reports are available on the use of biogenically synthesized AgNPs without supporting on other materials for the reduction reactions. To the best of our knowledge, we are reporting for the first time, one pot environmentally benign and rapid synthesis of AgNPs using easily available Z. Jujuba leaf extract as a green reducing and stabilizing agent. The synthesized AgNPs were thoroughly characterized by UV-Vis., XRD, FT-IR, TEM, DLS and Zeta potential. Moreover, the synthesized AgNPs were tested for the reduction of 4-nitrophenol to 4-aminophenol (4-AP) and Methylene Blue. Further AgNPs were also tested for antimicrobial activity against Escherichia coli (E. coli). 2. Materials and methods Z. Jujuba leaves were collected from Kolhapur India. Silver nitrate was obtained from Sigma Aldrich chemicals Pvt. Ltd., Mumbai. NaOH, H2SO4, NaBH4, 4-nitrophenol and Methylene Blue were purchased from Spectrochem Pvt. Ltd., Mumbai (India). All the chemicals were of analytical grade and used as supplied. All solutions were prepared in millipore water obtained from millipore water system (Millipore Corp. Bangalore, India). All glassware used for preparation of AgNPs were properly washed with distilled water and dried in oven.

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2.1. Synthesis of AgNPs Fresh leaves of Z. Jujuba were collected and washed several times with distilled water then shade dried to remove residual moisture. 10 g of dried leaves were crushed, boiled in 200 mL of distilled water for 10 min then leaf extract was filtered through Whatman No. 1 filter paper and centrifuged at 1500 rpm for 5 min to remove heavy biomaterials. Further, for the formation of AgNPs 1.5 mL of leaf extract was added into 100 mL (0.001M) aqueous solution of AgNO3 with constant stirring at room temperature. 2.2. Effect of quantity of leaf extract on AgNPs synthesis In order to optimize the quantity of leaf extract for the synthesis of AgNPs the leaf extract was varied from 0.5 to 2.5 mL in 100 mL of AgNO3 solution. The formation of AgNPs were tested by using UV-Vis. spectrophotometer. 2.3. Effect of concentration of AgNO3 on synthesis of AgNPs The concentration of precursor is also significant parameter for the synthesis of nanoparticles. Therefore, the concentration of AgNO3 solution was optimized by varying the concentration of AgNO3 from 0.0001 to 0.01M. 2.4. Effect of pH on synthesis of AgNPs pH is a significant factor for biosynthesis of nanoparticles. The effect of pH on the synthesis of AgNPs was studied in the range of 4 to 9. To control the pH, H2SO4 and NaOH (0.01M) solutions were used. The other factors like volume of leaf extract, concentration of

4

AgNO3 and temperature were kept constant. The effect of pH on the synthesis of AgNPs was monitored by UV-Vis. spectroscopy. 2.5. Characterization of AgNPs The formation of AgNPs was monitored by UV-Vis. NIR spectrophotometer (Shimadzu, UV-3600). XRD analysis describes the crystallinity and an average crystalline size of synthesized material. FT-IR (Cary 630, Agilent Technology) spectral analysis in the range of 4000-650 cm-1 has been carried out to identify the possible biomolecules present in the Z. Jujuba leaf extract which are responsible for the formation and stability of AgNPs. The shape and size of the materials were obtained by using transmission electron microscopy (TEM). TEM images were scanned with JEOL JEM 2100 TEM equipped with high resolution Gatan CCD camera. The particle size distribution and stability of nanoparticles were evaluated by DLS and Zeta potential measurements using Malvern Zetasizer (nano ZS-90) equipped with 4 mW, 633 nm HeNe Laser (U. K.) at 25°C under a fixed angle of 90° in disposable polystyrene cuvettes. 2.6. Catalytic activity of AgNPs The catalytic activity of synthesized AgNPs was tested for the reduction of 4-NP and MB. The reduction reactions were monitored by UV-Vis. spectrophotometer. 2.6.1. Catalytic activity of AgNPs for reduction of 4-NP In order to check the catalytic activity of synthesized AgNPs for the reduction of 4-NP in a 3.5 mL quartz cuvette 1.5 mL of water, 0.5 mL of 4-NP (0.5 mM) and 1 mL NaBH4 (0.02 M) were taken. In the same solution 15 µL colloidal solution of AgNPs were added and time dependant absorption spectra were recorded for the reduction reaction at room temperature.

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2.6.2. Catalytic activity of AgNPs for reduction of MB The catalytic activity of AgNPs for the reduction of MB was investigated by comparing the absorbance values of three different reaction mixtures which is filled in three test tubes. In the first test tube, 1 mL MB (5 ppm) and 2 mL water were taken and the reaction was monitored by using UV-Vis. spectrophotometer. In the second test tube, 1 mL MB (5 ppm) and 0.2 mL Z. Jujuba leaf extract were mixed into 1.8 mL water and the absorbance was monitored after 30 min. In the third test tube, 1 mL MB (5 ppm) and 0.2 mL Z. Jujuba leaf extract were mixed into 1.8 mL AgNPs and absorbance was monitored after 30, 45, 60 and 75 min. After that absorbance values of first and second reaction mixture were compared with the absorbance values of third one. 2.7. Antimicrobial activity The antimicrobial activity of AgNPs against E. coli was studied by agar well diffusion method. Stock culture was maintained at 4°C on agar slant of nutrient media. Before experiment, pure culture was sub cultured onto nutrient agar slant and it was incubated overnight at 37°C. Later nutrient agar plate was prepared and punctured for wells and located for 50 µL of AgNPs. After 24 hrs of incubation, the zones of growth inhibition were observed. We also studied controlled activity by Z. Jujuba leaf extract. 3. Results and discussion 3.1. Visual observation and UV-Vis. studies It is well known that AgNPs exhibit strong absorption band in visible region and generate specific color to the solution. The qualitative analysis of AgNPs was carried out by visual

6

observations. When the addition of Z. Jujuba leaf extract into the aqueous solution of AgNO3 began to change from clear solution to yellowish brown and finally it becomes brick red that indicates the formation of AgNPs [23]. UV-Vis. spectroscopy is a foremost technique to authenticate the formation and stability of AgNPs in an aqueous solution. The spectrum band at 434 nm (Fig.1.) corresponding to the surface plasmon resonance of the metallic AgNPs [24]. 3.1.1. Effect of quantity of leaf extract on AgNPs synthesis Effect of quantity of leaf extract on the synthesis of AgNPs was evaluated by UV-Vis. spectroscopy. As the quantity of leaf extract increases the intensity of the band (434 nm) of AgNPs also increases which is shown in Fig. 2. From the figure it is clear that for 0.5 and 1.0 mL of leaf extract, the absorption bands were broad which indicates larger particle size, at 1.5 mL of leaf extract absorbance band was found to be narrow that indicates narrow size distribution [25]. Further at 2.0 and 2.5 mL of leaf extract absorbance bands were red shifted. From above discussion, it is clear that optimized quantity of leaf extract for the preparation of AgNPs was found to be 1.5 mL for

100 mL of AgNO3 (0.001M).

3.1.2. Effect of concentration of AgNO3 on synthesis of AgNPs The effect of concentrations of AgNO3 on the formation of AgNPs were monitored by using UV-Vis. spectroscopy (Fig. 3.). It is seen from the figure, at 0.0001 M AgNO3, wider SPR band

occurred at 425 nm that is a characteristic band of AgNPs with large size. At 0.001 M

AgNO3, the SPR narrow band at 434 nm indicates smaller size of AgNPs. For 0.01 M AgNO3, wider SPR band obtained at 446 nm which is red shift and that indicates large size distribution of AgNPs. From the above discussion it is clear that in the present study, the

an optimum

concentration of precursor for the synthesis of AgNPs by using using Z. Jujuba leaf extract (1.5 7

mL) is found to be 0.001M AgNO3. Similar observations were also reported by Tripathi et al. [25] 3.1.3. Effect of pH on synthesis of AgNPs The effect of pH on the formation of AgNPs was evaluated by using UV-Vis. spectroscopic studies. Formation of AgNPs mainly depends on the pH of the reaction medium and the results are shown in Fig. 4. From figure it is clear that as the pH increases from 4 to 9, the absorbance value increases steadily which indicates the rate of formation of AgNPs increases from acidic to basic medium. At acidic pH 4 to 6, bands were wider and display red shift owing to increase in particle size. In basic condition at pH 8 and 9, bands were narrow and display blue shift due to decrease in particle size [26]. The formation of AgNPs occurs rapidly, in neutral and basic pH this may be due to the ionization of the phenolic groups present in the leaf extract [27]. The slow rate of formation and aggregation of AgNPs in acidic pH could be related to electrostatic repulsion of anions present in the solution [28]. At basic pH there is a possibility of AgOH precipitation [29]. On the basis of these results, it could be concluded that the optimum condition for the preparation of AgNPs using Z. Jujuba leaf extract was neutral medium. The effect of reaction time is shown in Fig. 5. from figure and visual observations it is clear that the color of solution changes after addition of extract into the AgNO3 solution and the aqueous solution of AgNO3 turned colorless to brown within 10 min. SPR intensity of AgNPs increases with time and saturates within 40 min, this result suggest the formation of anisotropic molecules that are stabilized later in the medium [30]. Many researchers have reported on the synthesis of AgNPs by using various plant extract and time duration from several days to hours ,typically time duration was 7 days, 4 and 2 hrs [16, 31, 32] respectively. In the present

8

investigation the rate of formation of AgNPs by using Z. Jujuba leaf extract was found to be 10 min which is very fast as compared to above reported plant extract, it might be due to the presence of alkaloid, terpenoid and poly phenolic compounds like rutin and apigenin-7-glucoside in Z. Jujuba leaf extract which acts as strong reducing agent. 3.2. XRD analysis The XRD pattern of biosynthesized AgNPs (Fig. 6.) clearly indicates the intense bands in the whole pattern of 2θ values ranging from 20 to 90°. The distinct diffraction bands at 38.04°, 43.69°, 64.39°, 77.07° and 81.49° are indexed to the [111], [200], [220], [311] and [222] crystallographic planes of AgNPs, respectively which plainly indicates crystalline nature of AgNPs. The observed set of lattice planes were indexed based on the face centered cubic (FCC) structure of silver by comparing the XRD data with JCPDS Card No. 89-3722. The unpredicted crystalline band (31.84°) also present this band could be due to the organic compound belongs in the leaf extract [33, 34]. The average crystallite size of 6 nm has been estimated by using well known Scherrer’s formula. 3.3. FT-IR analysis The FT-IR spectrum of Z. Jujuba leaf extract (Fig. 7. a.) shows a number of bands that shows a complex nature of leaf extract. Absorption band at 3305 cm–1, which is characteristic of OH stretching of a phenolic group and the small sharp band appearing at 2936 cm–1 may be due to C–H stretching of methylene group. The band at 1365 cm–1 corresponding to C‒N stretching and absorption band at 1449 cm–1, corresponds to aromatic C‒H stretching. Band at 1027 cm–1, indicates C‒OH stretching is shifted (from 1027 cm–1 to 1175 cm–1) after reaction and the band arising at 846 cm–1, is due to out of plane N‒H bending of amides. The absorption band at 1597 9

cm–1 corresponds to C═O stretching (carboxylic or amide) which is shifted (from 1597 cm–1 to 1640 cm–1) after reaction. bands at 3835, 3366, 1643, 1495 and 1175 cm–1 in the spectrum of synthesized AgNPs (Fig. 7. b.) and shifting of bands after reaction that indicates the possible involvements of carboxyl groups, intermediate form of phenolic groups, proteins and carbohydrates of Z. Jujuba leaf extract in the reduction processes for the synthesis of nanoparticles. 3.4. TEM analysis TEM images were used to observe the surface morphology of as synthesized AgNPs by using Z. Jujuba leaf extract. Fig. 8(a–d.) shows various TEM images with different magnifications which reveals the formation of better silver nanoparticles with various shapes (in the range 20-30 nm). This better particle size with various shapes of AgNPs may be due to complex constituents of plant extract [35-37]. HRTEM image of AgNPs is shown in (Fig. 8. e.) clearly indicates the interplanar distance closely matches with XRD pattern. Selected Area Electron Diffraction (SAED) (Fig. 8. f.) shows the dotted concentric ring which indicates the obtained AgNPs are nanocrystalline form with the different shapes [25]. The planes obtained from XRD were also matched with the SAED pattern. 3.5. DLS and Zeta potential measurement The size distribution and an average particle size of synthesized AgNPs were obtained using particle size analyzer (Fig. 9 a). Dynamic Light Scattering (DLS) histogram analysis shows the size distribution of particles with maximum intensity was found to be at 28 nm. The zeta potential (Fig. 9. b.) of about –26.4 mV was observed which indicates AgNPs are very stable in colloidal state [38]. 10

3.6. Catalytic activity of AgNPs The catalytic activity of synthesized AgNPs was tested for the reduction reactions of 4NP and MB. 3.6.1. Catalytic activity of AgNPs for reduction of 4-NP An aqueous solution of 4-NP shows absorption maxima at 317 nm which is shown in Fig. 10. After addition of NaBH4 into the solution of 4-NP the band of 4-NP was red shifted from 317 to 400 nm and color of 4-NP changed from pale yellow to deep yellow due to formation of 4-nitrophenolate ion in alkaline solution [39, 40]. The band at 400 nm remain unchanged even for two days in absence of AgNPs. After the addition of 15 µL of AgNPs, color fades to colorless and band at 400 nm gradually decreases with simultaneous appearance of band at 298 nm corresponding to the formation of 4-AP [41]. Thus in the process of catalytic reduction of 4NP the AgNPs transfer the electrons from the BH4¯ ions to nitro compound [42] which was qualitatively monitored by UV-Vis. spectrophotometer. The band at 400 nm completely disappears with increasing intensity of the band at 298 nm within 15 minutes indicating complete reduction of 4-NP to 4-AP. Since the concentration of NaBH4 was taken in large excess to that of the 4-NP, the reduction can be considered as pseudo first-order reaction with respect to 4-NP alone. Therefore, the reaction kinetics can be ascribed as: ln(C/C0) = ‒ kt, where k is the apparent first-order rate constant, t is the reaction time; C and C0 are the concentrations of 4-NP at time t and at initially time 0, respectively,

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The absorbance of 4-NP is proportional to concentration in solution, the absorbance at time t (A) and time 0 (A0) are equivalent to concentration at time t (C) and time 0 (C0) respectively. Linear relationship between ln(C/C0 ) and reaction time t in the reduction reaction catalyzed by AgNPs (Fig. 11.) follows pseudo first-order kinetics with a correlation coefficient of 0.945. The rate constant (k) was found to be 0.22 min-1 which is directly obtained from the slope of straight line. 3.6.2. Catalytic activity of AgNPs for reduction of MB In many homogeneous and heterogeneous reactions the electron transfer step is a significant in an electron transfer, there may be a large redox potential difference between the donor and accepter, and this state may restrict the passage of electrons. Easy transfer of electrons takes place when effective catalyst with an intermediate redox potential value of the donor– acceptor partner is used. The catalytic activity of AgNPs for the reduction of MB was monitored by UV-Vis. spectrophotometer and the results are shown in Fig. 12. Pure MB has λmax value of 664 nm. After 30 min. of addition of extract into the dye solution, the absorbance was decreased slowly along with red shift. A decrease in absorbance is an indication of reduction of MB by phytoextract. System containing dye, AgNPs and the extract shows a noticeable decrease in the absorbance of MB and increase of SPR band of AgNPs after 30, 45, 60 and 75 min. In the present study synthesized AgNPs act as an efficient electron transfer mediator between leaf extract and MB by acting as a redox catalyst, which is called electron-relay system [43]. 3.7. Antimicrobial activity The AgNPs shows excellent antimicrobial activity against E. coli. Clear zones of inhibition of bacterial growth on nutrient agar plates around the holes impregnated with test 12

AgNPs are shown in Fig.13. The radial diameter of inhibition zone is 14 mm at the same time the controlled cavities containing Z. Jujuba leaf extract does not shown any inhibition zone. 4. Conclusions In summary, a green novel route for the rapid, ecofriendly and non-toxic synthesis of AgNPs at room temperature using easily available Z. Jujuba leaf extract which act as a green reducing as well as stabilizing agent. The synthesized AgNPs are found in various shapes with size of 20 to 30 nm and also found to be very stable. As-synthesized AgNPs was found to be highly efficient catalytic activity towards the reduction of 4-NP to 4-AP and MB within shorter reaction time. The biosynthesized AgNPs also have significant antimicrobial activity against E. coli. The use of the leaf extract for the rapid synthesis of AgNPs is an alternative green route to chemical as well as physical methods. The reaction time for the synthesis of AgNPs in the present study was found to be significantly lower than that of earlier reports. This biogenic synthesis may open door for the other noble metals also. Acknowledgements One of the authors (NLG) is thankful to DST-PURSE for providing fellowship and (KMG) acknowledges to DST for providing financial assistance under the Major Research Project (SR/S1/PC/0041/2010). We also acknowledge To Director, SAIF North-Eastern Hill University Shillong for providing TEM facility.

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Figure Captions 1) Fig. 1. UV-Vis. spectrum of synthesized AgNPs from Z. Jujuba leaf extract. 2) Fig. 2. UV-Vis. absorption spectra of AgNPs at various amount of Z. Jujuba leaf extract. 3) Fig. 3. UV-Vis. absorption spectra of AgNPs at different concentrations of AgNO3. 4) Fig. 4. pH dependence UV-Vis. absorption spectra of AgNPs. 5) Fig. 5. Time dependant UV-Vis. absorption spectra of formation of AgNPs. 6) Fig. 6. XRD pattern of Biogenically synthesized AgNPs. 7) Fig. 7. FT-IR spectra, a) Z. Jujuba leaf extract, b) synthesized AgNPs. 8) Fig. 8. a–d shows TEM images of synthesized AgNPs. e) HRTEM image of synthesized AgNPs, f) SAED pattern of AgNPs. 9) Fig. 9. Particle size distribution analysis, a) Dynamic light scattering measurements, b) Zeta potential. 10) Fig. 10. UV-Vis. spectra for 4-NP and successive reduction of 4-NP with NaBH4 catalyzed by AgNPs. 11) Fig. 11. Plot of ln(C/C0) against the time for reduction of 4-NP by NaBH4 using AgNPs as a catalyst. 12) Fig. 12. UV-Vis. spectra of reduction of MB by Z. Jujuba in the presence of AgNPS, a) MB, b) MB+Z. Jujuba leaf extract (after 30 min), c) MB +Z. Jujuba leaf extract +AgNPs (after 30min), d) MB + Z. Jujuba leaf extract +AgNPs (after 45 min.), e) MB + Z. 18

Jujuba leaf extract +AgNPs (after 60 min.), f) MB + Z. Jujuba leaf extract +AgNPs (after 75 min.), 13) Fig. 13. Representative results of antimicrobial activity of AgNPs against E. Coli.

Figures

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0.2

0.0 300

400

500

Wavelength(nm)

Fig. 5.

21

600

700

240

[111]

220 200

Intensity

180

[200]

160 140 120 100

[311]

[220]

80

[222]

60 40 20

30

40

50 60 2θ (degree)

Fig. 6.

22

70

80

90

Fig. 7. a, b.

23

Fig. 8. a, b, c, d, e, f.

24

Fig. 9. a, b.

25

3.0

0 Min.

2.5

4-NP

Absorbance

2.0

1.5

1.0

4-AP

0.5

15 Min.

0.0 300

400

500

Wavelength (nm)

Fig. 10.

0.0 -0.5

ln(C/C0)

-1.0 -1.5 -2.0 -2.5 -3.0 -3.5 0

2

4

6

8

10

Time (min)

Fig. 11.

26

12

14

16

0.25

Absorbance

0.20

a b c d e f

0.15

0.10

0.05

0.00 500

600

Wavelength (nm)

Fig. 12.

27

700

Fig. 13.

28

9) Fig. 9. Particle size distribution analysis a) Dynamic light scattering measurement, b) Zeta Potential

Fig. 9.a.

Fig. 9.b.

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Graphical Abstract

19

Highlights: •

A novel synthesis of AgNPs at room temperature using Ziziphus Jujuba leaf extract.

•

A green, rapid, one step, cost effective, environmentally friendly synthesis.

•

Biogenically synthesized AgNPs were found to be stable for more than six months.

•

It does not require external energy, stabilizing agent and hazardous chemicals.

•

Reduction of 4-nitrophenol and methylene blue for environmental protection. Also the synthesized AgNPs shows good antimicrobial activity against Escherichia coli

20