Facile Green Chemistry Synthesis of Ag Nanoparticles Using Areca Catechu Extracts for the Antimicrobial Activity and Photocatalytic Degradation of Methylene Blue Dye

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ScienceDirect Materials Today: Proceedings 9 (2019) 499–505

www.materialstoday.com/proceedings

GMSP&NS’18

Facile Green Chemistry Synthesis of Ag Nanoparticles Using Areca Catechu Extracts for the Antimicrobial Activity and Photocatalytic Degradation of Methylene Blue Dye Vinay S. P.*, Chandrasekhar N Research and Development Center, Department of Chemistry, Shridevi Institute of Engineering and Technology, Sira Road, Tumakuru - 572106, Karnataka, India.

Abstract Green synthesis of silver nanoparticles (Ag NPs) using Areca catechu extract as reducing agent was investigated. The bioreduction of silver ions into silver nanoparticles was monitored using UV-visible spectrophotometry. The synthesized Ag NPs were studied by Fourier transform infra-red spectroscopy (FTIR), UV–visible spectroscopy, Transmission electron microscopy (TEM) analysis, Energy dispersive X-ray analysis (EDX) and X-ray diffraction (XRD) techniques. The photocatalytic degradation of methylene blue was estimated spectrophotometrically using the green synthesized silver (Ag) as nanocatalyst. Disc diffusion method was used to investigate the antimicrobial properties of the Ag NPs against the tested bacterial strains. © 2018 Elsevier Ltd. All rights reserved. Selection and peer-review under responsibility of the scientific committee of the International Conference on Green Methods for Separation, Purification and Nanomaterial Synthesis, GMSP&NS’18, 24–25th April 2018, Centre for Nano and Material Sciences, Jain University, Bangalore 562112, Karnataka, India. Keywords: Green synthesis; Areca catechu (Areca nut); Ag NPs; TEM; Photocatalytic degradation; Antimicrobial activity.

1. Introduction Nanoparticles are the fundamental building blocks of nanotechnology. The most important and distinct property of nanoparticles (NPs) is their larger surface area to volume ratio [1,2]. Now-a-days, nanotechnology owes to the tremendous improvement in human life and it has emerged as a multidisciplinary research field. Among the various fields of nanotechnology, green nanotechnology provides more effective nanoparticles synthesis with expected products and economical manner. The eco-friendly green mediated synthesis of inorganic nanoparticles is a fast *E-mail address: [email protected] 2214-7853 © 2018 Elsevier Ltd. All rights reserved. Selection and peer-review under responsibility of the scientific committee of the International Conference on Green Methods for Separation, Purification and Nanomaterial Synthesis, GMSP&NS’18, 24–25th April 2018, Centre for Nano and Material Sciences, Jain University, Bangalore 562112, Karnataka, India.

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growing research in the limb of nanotechnology [2]. The biosynthesis method employing plant extracts have drawn attention as a simple and viable alternative to the conventional chemical and physical methods. Silver nanoparticles have given more research interest due to their wide range of applications in photography, catalysis, biosensor, biomolecular detection, diagnostics, and particularly antimicrobial activities antimicrobial paint coatings, water treatment, textiles, medical devices and HIV prevention as well as treatment [3-29]. It is very much required to develop the eco-friendly, reliable, cost effective and simple methods for the production of NPs. The development of green synthesis of nanoparticles is an important aspect of nanoscience and nanotechnology. Researchers have used various parts of the plants such as leaf [30], tuber [25], bark [26] and buds [24] for the synthesis of nanoparticles. Microorganisms such as bacteria [7], fungi [17, 22] and algae [31-34] are also used for the synthesis of nano particles. Among many methods of synthesizing the nanoparticles, plant mediated synthesis process is more advantages over the chemical and microbe mediated synthesis method because it eliminate the culture maintaining process [13, 23]. Plants have alkaloids, flavonoids and polyphenolic mixes which may decrease the Ag ions to Ag NPs and goes about as topping and balancing out specialist [12]. Silver nanoparticles are promising, exceptionally efficient and stable photocatalysts under room temperature with visible light illumination for degrading organic dyes molecules. In this study, a restorative important plant Areca catechu nut extracts was utilized for the biosynthesis of Ag NPs. 2. Experiment 2.1. Materials Healthy nuts of Areca catechu were collected from from Kora village, India amid the long stretch of November 2016. Lyophilized culture of microorganisms E-coli, Pseudomonas aeruginosa, Klebsiella aerogenes and Staphylococus aureus were procured from the department of microbiology, Shridevi Institute of Medical Sciences and Research Hospital, India. The nutrient broth (N.B) media was obtained by Hi-Media Laboratories; AgNO3 solution was purchased from Merck Pvt. Ltd. India. 2.2. Preparation of extracts The crisply gathered nuts of Areca catechu were washed altogether with tap water to evacuate the clean and earth particles and after that it washed with twofold refined water. 20 g of slashed Areca catechu nuts were added to 250 mL conical flask containing 100 mL twofold refined water and mixed at 60 °C for 20 min on warming mantle. At that point the blend was cooled for 5 min and the filtrate was separated utilizing Whatman filter paper No.1. The gathered filtrate was stocked at 4 °C for further analysis. The extract (dark orange color) was utilized as a reducing agent for the biosynthesis of silver nanoparticles. 2.3. Synthesis of silver nanoparticles 10 mL of Areca catechu nuts concentrate was added to the 90 mL of 5mM AgNO3 solution and mixed persistently for 20 min using magnetic stirrer. The blend was further taken for bioreduction process. After 24 h, a dark orange color of the blend was changed to dark brown color due the formation of silver nanoparticles. 2.4. Characterizations of Ag nanoparticles The reduced silver nanoparticles were characterized using UV-visible spectrophotometer. The UV-visible spectra were recorded on a Shimadzu UV-visible sepectrophotometer in the wavelength range of 300-700 nm. The structural studies of the synthesized silver nanoparticles were studies using TEM (Tecnai T20, FEI model) instrument at 200 KeV. Before taken the Ag NPs to TEM instrument, the Ag NPs were dispersed with iso-propanol and exposed in ultra sonication for 10 min and dried in hot air oven. EDX analysis (TESCAN model) carried out to analyse the presence of Ag element as well as to observe the other elementary composition. The bio-synthesized Ag NPs were refined by continuous centrifugation at 10000 rpm for 15 minutes, using Remi cooling centrifuge C-24. The synthesized silver nanoparticles were further analyzed by Rigaku-Smart Lab model (40 kV, 30 mA). Obtained

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data was registered in the graph with 2θ v/s intensity. The FT-IR analysis was carried by using Nicolet IS10 using KBr pellet in the range of 400-4000 cm-1.. 2.5. Photocatalytic degradation The photocatalytic degradation properties of green synthesized silver nanoparticles were assessed using methylene blue (10 mg/L) in water solution under the direct sunlight. The measured amount of Ag NPs and dye were taken in the water and kept under dark to obtain the equilibrium. Then this solution mixture was exposed to sunlight for six hours. The degradation of the dye solution was estimated through recording the optical absorption of the dye molecules in a periodical intervals. 2.6. Antimicrobial assay The antimicrobial activity of blended Ag NPs was assessed by the disc diffusion strategy for their potential biomedicinal applications. E-coli, Pseudomonas aeruginosa, Klebsiella aerogenes and Staphylococus aureus bacterial strains were produced in nutrient broth (NB) media for 24 h at 37°c and 1 mL of every broth culture was conveyed on nutrient agar media. 5 mm sterilized filter paper discs were dipped in synthesized Ag NPs suspension (10µL), twofold refined water as negative control, Taxim (1µg/mL) as standard drug likewise utilized as a positive control and nuts concentrate was set over the agar plates and incubated for 24 h at encompassing (room) temperature. The diameter of zone of inhibition was measured. 3. Results and discussions Silver nanoparticles were synthesized utilizing Areca catechu nut extract from aqueous AgNO3 solution (5 mM) at ambient temperature. Initially, the Areca catechu extract and AgNO3 solution reaction mixture slowly turns into dark brown in colour as shown in Fig. 1 [6].

Fig. 1. Green synthesis of AgNPs.

3.1. Phytochemical analysis The results of phytochemical screening of Areca catechu extracts demonstrated the presence of cardiac glycosides, alkaloids, tannins, phenols, amino acids, terpenoids, saponins and flavonoids (Table 1). Table 1. Phytochemical analysis S. No. 1 2 3 4 5 6

Phytochemicals Areca catechu extracts Flavonoids + Alkaloids + Phenols + Tannins － Cardiac glycosides － Starch + +: present; －: absent

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3.2. UV-Vis spectroscopy analysis The formation of Ag NPs was confirmed by UV-Visible spectrum by observing the surface plasmon resonance band at 430 nm as shown in Fig. 2 [15].

Fig. 2. UV-vis absorption spectra of AgNPs.

3.3. TEM analysis The shape and particle size of Ag NPs were assessed with Transmission Electron Microscope as shown in Fig. 3. The TEM image shows that the synthesized Ag NPs using the Areca catechu possess an average size of 12 nm with spherical and irregular in shapes [11, 20].

Fig. 3. TEM image of AgNPs.

3.4. FTIR analysis Further, the synthesized Ag NPs were characterized by FT-IR spectroscopy (Fig. 4). The FT-IR spectral result showed a broad peak at 3370 cm-1 that assigned to O-H stretching vibration of the alcohol group. The other peaks at 2921 and 1628 cm-1 were assigned to C-H and C=C stretching vibration of alkane and ketone groups, respectively. The sharp peaks at 1374 cm-1 represented the N-O nitro compound and 1025 cm-1 was assigned to C-O stretching vibration, represented the anhydride group and broad peak at 529 cm-1 could be due to the C-Br halocomound group. The spectral result of FT-IR formerly reported indicated that the Areca catechu extract phytochemicals such as phenol, starch, alkaloids, flavonoids and alkaloids (Table.1) [29] might be active in the Ag NPs synthesis process [28, 5].

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Transmittance %

0.95 0.90 0.85 0.80

529 2921

0.75

1025 1374

3370

0.70

1628 0.65 500

1000

1500

2000

2500

3000

3500

4000

-1

Wavenumber (cm )

Fig. 4. FT-IR spectrum of AgNPs.

3.5. XRD analysis XRD analysis of Ag NPs confirmed that the structure of Ag NPs is face centered cubic and are crystalline in nature (Fig. 5). The diffraction peak at 2θ values at 38.21, 44.52, 64.15 and 77.73° could be attributed to lattice planes at (111), (200), (220) and (311) respectively. The obtained XRD pattern was matched with the JCPDS file No. 4-783 [15]. It predicted that the unassigned peaks are due to bio-organic phases that take place on the overhead of the Ag NPs [14, 31].

Fig. 5. XRD pattern of the synthesized Ag NPs.

3.6. EDX analysis The formation of Ag metal was confirmed by EDX examination (Fig. 6) [21] and other elements appeared may be due to the extract.

Fig. 6. EDX spectrum of AgNPs.

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3.7. Photocatalytic degradation It is clear from Fig. 7 that the synthesized Ag NPs have exhibited the photocatalytic degradation of methylene blue dye. The characteristic absorption peak of methylene blue solution was observed to be 664 nm. Degradation of methylene blue was analyzed by decreasing peak intensity of the dye molecules with time. The degradation of dye could be due to the production of reactive oxygen species by the silver nanoparticles upon the light irradiation. 6 hr 1 hr 40 min 20 min 0 min

2.0

Absorbance (a.u)

1.5

1.0

0.5

0.0 500

600

700

Wavelength (nm)

Fig. 7. Photocatalytic degradation of methylene blue using synthesized silver nanoparticles

3.8. Antimicrobial assay From the results, it was observed that the synthesized Ag NPs exhibit promising antimicrobial activity when compared to the standard i.e. Taxim (Fig. 8, Table. 2) The zone of inhibition of Ag NPs was found to be in the range of 18 - 23mm against all tested microbes [35, 10].

Fig. 8. Antibacterial activity of AgNPs. Table 2. Antibacterial Zone of Iinhibition. S. No. 1 2 3 4

Strains E-coli Pseudomonas aeruginosa Staphylococcus aureus Klebsiella aerogenes

Control Double distilled water

Standard 16 19 16 14

Ag Nps 18 23 20 20

Plant extract Areca catechu

4. Conclusion In conclusion, the green synthesis of silver nanoparticles was successfully carried out using the Areca catechu extracts. From the XRD analysis, it was confirmed that the synthesized silver nanoparticles possess f.c.c structure along with good crystalline nature. The TEM analysis revealed that the average size of silver nanoparticles was 12

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nm with special and irregular shapes. The Ag NPs were found to be promising in degrading methylene blue dye under sunlight light irradiation. Also, this green synthesized Ag NPs have exhibited enhanced antimicrobial activity against various microorganisms such as E-coli, Pseudomonas aeruginosa, Klebsiella aerogenes and Staphylococus aureus. These observed results propose that the green synthesized Ag NPs can be the promising materials for the visible light degradation of dyes as well as the antimicrobial agents. Acknowledgements We thank Dr. M R Hulinaykar, Managing Trustee, Sri Shridevi Charitable Trust, Dr. H. B. Phani Raju, Principal, SIET, Tumakuru, India for their encouragement during the research work. We thank the staff, Department of Chemistry, Government Arts and Science College, Bangalore, Karnataka, India, for their service in assisting with FTIR analysis facility during this research work References [1] M.A. Albrecht, C.W. Evans, C.L. Raston, Green Chem. 8 (2006) 417–432. [2] Arangasamy Leela, Munusamy Vivekanandan, African.J.Biotechnol. 7(17) (2008) 3162-3165. [3] S. Basu, S. Jana, S. Pande, T. Pal, J. Colloid and Interface Sci. 321 (2008) 288-93. [4] I. Brigger, C. Dubernet, P. Couvreur, Adv. Drug. Deliv. Rev. 54 (2002) 631–651. [5] P. Daizy, Spectrochimica Acta Part A. 78 (2011) 327–331. [6] Diana Cruz, L. Fale, Mourato Ana, D. Vaz, M. Lusia Serralheiro, Ana Rosa LLino, Colloids Surf B: Biointerfaces. 810 (2010) 67-73. [7] D. Mandal, M.E. Bolander, D. Mukhopadhyay, S. Sankar, P. Mukherjee, Appl. Microbiol. Biotechnol. 69 (2006) 485–492. [8] N. Duran, P.D. Marcato, G.I. De Souza et al., J. Biomed Nanotechnol. 3 (2007) 203–8. [9] M.G. Guzman, J. Dille, S. Godet, World Academy of Science, Engineering and Technology. 43 (2008) 357-364. [10] Hemali Pandalia, Pooja Moteriya, Sumitra Chanda, Arabian Journal of Chemistry. 8 (2015) 732-741. [11] U.B. Jagtap, V.A. Bapat, Industrial Crops and Products. 46 (2013) 132-137. [12] J.L. GardeaTorresdey, E. Gomez, J. PeraltaVidea, J.G. Parsons, H.E. Troiani, Nano. Lett. 2 (2002) 397–401. [13] J. Anarkali, D. Vijaya Raj, K. Rajathi, S. Sridhar, Archives of Applied Science Research. 4(3) (2012) 1436-1441 [14] K. Kalimuthu, R.S. Babu, D. Venkataraman, M. Bilal, S. Gurunathan, Colloids Surf. B: Biointerfaces. 65 (2008) 150-153. [15] Kaushik Roy, C.K. Sarkar, C.K. Ghosh, Appl Nanosci. 5 (2015) 945–951. [16] R.A. Khaydarov, R.R. Khaydarov, O. Gapurova, Y. Estrin, T. Scheper, J. Nanopart. Res. 11 (2008) 1193. [17] K.S. Hemath Naveen, Gaurau Kumar, Karthik, K.V. Bhaskara Rao, Archives of Applied Science Research. 2(6) (2010) 161-167. [18] A. Kumar, P.K. Vemula, P.M. Ajayan, G. John, Nature. 7(3) 2008 236-41. [19] C.J. Murphy, A.M. Gole, S.E. Hunyadi, J.W. Stone, P.N. Sisco, A. Alkilany, B.E. Kinard, P. Hankins, Chem. Commun. 7(5) (2008) 544-557. [20] Naheed Ahmad, Seema Sharma, Green and Sustainable Chemistry. 2 (2012) 141-147. [21] K.L. Niraimathi, V. Sudha, R. Lavanya, P. Brindha, Colloids Surf. B, Biointerfaces. 102 (2013) 288-291. [22] N. Ahmad, S. Sharma, MdK Alama, V.N. Singh, S.F. Shamsid, B.R. Mehta, A. Fatma, Colloids Surf B: Biointerf. 81 (2010) 81–86. [23] N.C.J. Packia Lekshmi, S. Benarcin Sumi, S. Viveka, S. Jeeva, J. Raja Brindha, J. Microbiol. Biotech. Res. 2(1) 2012 115-119. [24] D. Raghunandan, M.D. Bedre, S. Basavaraja, B. Sawle, S.Y. Manjunath, A. Venkataraman, Colloids Surf. B. Biointerfaces. 79 (2010) 235– 40. [25] M. Sathishkumar, K. Sneha, Y.S. Yun, Bioresour.Technol. 101 (2010) 7958–7965. [26] M. Sathishkumar, K. Sneha, S.W. Won, C.W. Cho, S. Kim, Y.S. Yun, Colloids Surf. B. Biointerfaces. 73 (2009) 332–8. [27] A. Sathya, V. Ambikapathy, Drug invent. Today. 4(8) (2012) 408- 410. [28] R. Sathyavathi, M.B. Krishna, S.V. Rao, R. Saritha, D.N. Rao, Adv Sci Lett. 3 (2010) 1-6. [29] Senthil Amudhan, V. Hazeena Begum, K.B. Hebba, IJPSR. 3(11) 2012 4151-4157. [30] S.S. Shankar, A. Rai, A. Ahmad, M. Sastry, J. Colloid InterfaceSci. 275 (2004) 496–502. [31] G. Singhal, R. Bhavesh, K. Kasariya, A.R. Sharma, R.P. Singh, J Nanopart Res. 13 (2011) 2981-2988. [32] T. Sun, K. Seff, Chem. Rev. 94(4) 1994 857. [33] S. Rajeshkumar, C. Kannan, G. Annadurai, Int J Pharm Bio Sci. 3(4) (2012) 502 – 510. [34] S. Rajeshkumar, C. Kannan, G. Annadurai, Drug Invention Today. 4(10) 2012 511-513. [35] M. Thakur, S. Pandey, A. Mewada, R. Shah, G. Oza, M. Sharon, Acta, Part A, Mol. Biomol. Spectrosc. 109 (2013) 344-347. [36] D.J. Xiong, M.L. Chen, H. Li, Chem. Commun.(2008) 880-2. [37] Z. Zhu, L. Kai, Y. Wang, Materials Chemistry and Physics. 96 (2006) 447-453. [38] G. Ganapathy Selvam, K. Sivakumar, Appl Nanosci 5 (2015) 617–622 .