Optimization and Antifungal Activity of Silver Nanoparticles Synthesized Using the Leaf Extract of Corylus colurna L. (Turkish hazelnut)

Abstract

Objective: This study aims to optimize the production of microwave-synthesized silver nanoparticles (AgNPs) using Corylus colurna leaf extracts with response surface methodology based on the face-centered central composite design (FCCCD), characterize the synthesized nanoparticles by various spectroscopic and microscopic methods, and evaluate their antifungal effects on some Phytophthora species. Materials and Methods: In the study, the FCCCD of the response surface methodology was used to investigate the combined effect of three different synthesis variables (AgNO3 concentration, hazelnut leaf extract/AgNO3 ratio and reaction time) to achieve the maximum amount of AgNP at the spectral wavelength of 350-420 nm. The estimated area under the spectral curve was calculated with the help of Microsoft Office Excel program using a simple midpoint rule. Ultraviolet Visible (UV-Vis) Spectroscopy, Fourier Transform Infrared (FT-IR) Spectroscopy, and Transmission Electron Microscope (TEM) were used to characterize hazelnut leaf extract-AgNPs synthesized under optimum conditions. The antifungal activity of AgNPs was tested against six Phytophthora species (P. cactorum, P. capsici, P. cinnamomi, P. citrophthora, P. nicotianae and P. palmivora) under in vitro conditions, and the experiment was carried out with 6 replications. Results: UV-Vis spectroscopy revealed that typical surface plasmon resonance values of AgNPs synthesized under different conditions were in the wavelength range of 396 to 411 nm. Optimal AgNP production was achieved within the investigated range when the AgNO3 concentration, plant leaf extract/AgNO3 ratio, and reaction time were 5 mM, 0.1, and 90 seconds, respectively. FT-IR spectrum showed that AgNPs contain O–H, N–H, C=C, C–N, and C–O groups, and various compounds in hazelnut leaf extract play an important role in the synthesis of AgNPs. TEM analysis results revealed that AgNPs were in spherical form with an average size of 17.48 nm (Gaussian fit). Green synthesized AgNPs decreased mycelial growth of P. cactorum, P. capsici, P. cinnamomi, P. citrophthora, P. palmivora and P. nicotianae by 81.67%, 74.80%, 73.54%, 81.01%, 74.50%, and 62.39%, respectively. In addition, it was determined that the EC50 values of AgNPs varied between 118.58-292.56 µg ml-1, and the MIC values were above 340 µg ml-1. Conclusion: This study suggests that AgNPs synthesized by hazelnut leaf extract should be further investigated for use in combating diseases caused by Phytophthora species.

Keywords

• Corylus colurna
• silver nanoparticles
• face-centered central composite design
• Phytophthora
• toxicity

Corylus colurna L. (Türk Fındığı)’nin yaprak ekstraktı kullanılarak sentezlenen gümüş nanopartiküllerin optimizasyonu ve antifungal aktivitesi

Öz

Amaç: Bu çalışma, yüz merkezli merkezi kompozit tasarım (FCCCD)’a dayalı yanıt yüzey yöntemi (RSM) ile Corylus colurna yaprak ekstraktı kullanılarak mikrodalgada sentez edilen gümüş nanopartiküllerin (AgNP’lerin) üretimini optimize etmeyi, sentezlenen nanopartikülleri çeşitli spektroskopik ve mikroskobik yöntemlerle karakterize etmeyi ve bazı Phytophthora türleri üzerindeki antifungal etkilerini değerlendirmeyi amaçlamaktadır. Materyal ve Yöntem: Çalışmada, yanıt yüzey yönteminin FCCCD’i, 350-420 nm spektral dalga aralığında maksimum AgNP miktarını elde etmek için üç farklı sentez değişkeni (AgNO3 konsantrasyonu, fındık yaprak ekstraktı/AgNO3 oranı ve reaksiyon süresi)’nin birleşik etkisini araştırmak için kullanılmıştır. Spektral eğri altındaki tahmini alan basit bir orta nokta kuralı kullanılarak Microsoft Office Excel programı yardımı ile hesaplanmıştır. Optimum koşullar altında sentezlenen fındık yaprak ekstraktı-AgNP’leri karakterize etmek için Ultraviyole Görünür (UV-Vis) Spektroskopisi, Fourier Dönüşümlü Kızılötesi (FT-IR) Spektroskopisi ve Transmisyon Elektron Mikroskobu (TEM) kullanılmıştır. AgNP’lerin antifungal etkinliği, altı Phytophthora türü (P. cactorum, P. capsici, P. cinnamomi, P. citrophthora, P. nicotianae ve P. palmivora)’ne karşı in vitro koşullarda denenmiş olup, deneme 6 tekerrürlü olarak yürütülmüştür. Araştırma Bulguları: UV-Vis spektroskopisi, farklı koşullar altında sentezlenen AgNP’lerin tipik yüzey plazmon rezonans değerlerinin 396 ile 411 nm dalga boyu aralığında değiştiğini ortaya koymuştur. AgNO3 konsantrasyonu, bitki yaprak ekstraktı/AgNO3 oranı ve reaksiyon süresi sırasıyla 5 mM, 0.1 ve 90 saniye olduğunda, araştırılan aralıkta optimum AgNP üretimi elde edilmiştir. FT-IR spektrumu, AgNP’lerin O–H, N–H, C=C, C–N ve C–O gruplarını içerdiğini ve fındık yaprak ekstraktındaki çeşitli bileşiklerin AgNP’lerin sentezinde önemli bir rol oynadığını göstermiştir. TEM analiz sonuçları, AgNP’lerin ortalama 17.48 nm (Gauss uyumu) büyüklüğe sahip küresel formda olduğunu ortaya koymuştur. Yeşil sentezlenen AgNP’lerin P. cactorum, P. capsici, P. cinnamomi, P. citrophthora, P. palmivora ve P. nicotianae’nın misel gelişimini sırasıyla %81.67, %74.80, %73.54, %81.01, %74.50 ve %62.39’a kadar azaltmıştır. Ayrıca AgNP’lerin EC50 değerlerinin 118.58-292.56 µg ml-1 arasında değiştiği ve MIC değerlerinin ise 340 µg ml-1’in üzerinde olduğu belirlenmiştir. Sonuç: Bu çalışma, fındık yaprak ekstraktı ile sentezlenen AgNP’lerin, Phytophthora türlerinin neden olduğu hastalıkların mücadelesinde kullanılmak üzere daha fazla araştırılması gerektiğini önermektedir.

Anahtar Kelimeler

• Corylus colurna
• Gümüş nanopartikül
• Yüz Merkezli Merkezi Kompozit Tasarım
• Phytophthora
• toksisite

References

1. Agrios, G.N. (2005). Plant pathology (5nd ed.). Burlington, USA. Elsevier academic press.
2. Ahmed, S., Ahmad, M., Swami, B. L., & Ikram, S., (2016). Green synthesis of silver nanoparticles using Azadirachta indica aqueous leaf extract. Journal of Radiation Research and Applied Sciences, 9(1), 1-7.
3. Alamri, S.A., & Moustafa, M.F. (2012). Antimicrobial properties of 3 medicinal plants from Saudi Arabia against some clinical isolates of bacteria. Saudi Medical Journal, 33(3), 272-277.
4. Ali, K., Ahmed, B., Dwivedi, S., Saquib, Q., Al-Khedhairy, A. A., & Musarrat, J. (2015). Microwave accelerated green synthesis of stable silver nanoparticles with Eucalyptus globulus leaf extract and their antibacterial and antibiofilm activity on clinical isolates. PloS One, 10(7), 1-20.
5. Al-Zahrani, S.S., & Al-Garni, S.M. (2019). Biosynthesis of silver nanoparticles from Allium ampeloprasum leaves extract and its antifungal activity. Journal of Biomaterials and Nanobiotechnology, 10(1), 11.
6. Arslan, M. (2005). Batı Karadeniz Bölgesindeki Türk fındığı (Corylus colurna L.) populasyonlarının ekolojik ve silvikültürel yönden incelenmesi. Bolu, Türkiye.
7. Ayan, S., Ünalan, E., Yer, E.N., Sakıcı, O.E., & İslam, A. 2016. Population diversity in Northwest Anatolia Forests in terms of nut characteristics of Turkish hazelnut (Corylus colurna L.) (Kastamonu province), International Multidisciplinary Congress of Eurosia, Odesa, Ukraine.
8. Benov, L., & Georgiev, N. (1994). The antioxidant activity of flavonoids isolated from Corylus colurna. Phytotherapy Research, 8(2), 92-94.

9. Bhushan, B. (Ed.). (2017). Introduction to nanotechnology. In Springer handbook of nanotechnology. Berlin: Springer.
10. Botta, R., Molnar, T. J., Erdogan, V., Valentini, N., Torello Marinoni, D., & Mehlenbacher, S. A. (2019). Hazelnut (Corylus spp.) breeding. Advances in Plant Breeding Strategies: Nut and Beverage Crops, 4, 157-219.
11. Buzea, C., Pacheco, I. I., & Robbie, K. (2007). Nanomaterials and nanoparticles: sources and toxicity. Biointerphases, 2(4), 17-71.
12. Cai, Y., Piao, X., Gao, W., Zhang, Z., Nie, E., & Sun, Z. (2017). Large-scale and facile synthesis of silver nanoparticles via a microwave method for a conductive pen. The Royal Society of Chemistry advances, 7(54), 34041-34048.
13. Ceylan, O., Sahin, M. D., & Avaz, S. (2013). Antibacterial Activity of Corylus colurna L. (Betulacea) and Prunus divaricata Ledep. subsp. divaricata (Rosaceae) from Usak, Turkey. Bulgarian Journal of Agricultural Science 19, 1204-1207.
14. Childers, R., Danies, G., Myers, K., Fei, Z., Small, I.M., & Fry, W. E. (2015). Acquired resistance to mefenoxam in sensitive isolates of Phytophthora infestans. Phytopathology, 105(3), 342-349.
15. Chowdhury, S., Yusof, F., Faruck, M.O., & Sulaiman, N. (2016). Process optimization of silver nanoparticle synthesis using response surface methodology. Procedia Engineering, 148, 992- 999.
16. Dobrowolski, M. P., Shearer, B.L., Colquhoun, I.J., O’brien, P. A., & Hardy, G.S. (2008). Selection for decreased sensitivity to phosphite in Phytophthora cinnamomi with prolonged use of fungicide. Plant pathology, 57(5), 928-936.
17. Doğanyiğit, Z., Küp, F.Ö., Kaymak, E., Aslı, O., Burçin, O., & Akın, A.T. (2019). Gümüş nanopartiküllerin üzüm çekirdeği ekstraktının endotoksik kalp dokusundaki histolojik değişikliklere ve tnf ve bnp ekspresyonuna etkisi. Bozok Tıp Dergisi, 9(3), 87-96.
18. Eshghi, M., Kamali-Shojaei, A., Vaghari, H., Najian, Y., Mohebian, Z., Ahmadi, O., & Jafarizadeh-Malmiri, H. (2021). Corylus avellana leaf extract-mediated green synthesis of antifungal silver nanoparticles using microwave irradiation and assessment of their properties. Green Processing and Synthesis, 10(1), 606-613.
19. Gurunathan, S., Kalishwaralal, K., Vaidyanathan, R., Venkataraman, D., Pandian, S.R.K., Muniyandi, J., Hariharan, N., & Eom, S.H. (2009). Biosynthesis, purification and characterization of silver nanoparticles using Escherichia coli. Colloids and Surfaces B: Biointerfaces, 74(1), 328-335.
20. Henglein, A. (1989). Small-particle research: physicochemical properties of extremely small colloidal metal and semiconductor particles. Chemical reviews, 89(8), 1861-1873.
21. Hu, J. H., Hong, C. X., Stromberg, E. L., & Moorman, G. W. (2008). Mefenoxam sensitivity and fitness analysis of Phytophthora nicotianae isolates from nurseries in Virginia, USA. Plant Pathology, 57(4), 728-736.
22. Iravani, S., Korbekandi, H., Mirmohammadi, S.V., & Zolfaghari, B. (2014). Synthesis of silver nanoparticles: chemical, physical and biological methods. Research in Pharmaceutical Sciences, 9(6), 385–406.
23. İslam, A., Tüfekçi, F., & Turan, A. (2021). Fındık (1. Bölüm: Genel Özellikler). Ankara, Türkiye, Nobel Akademik Yayıncılık, 1-22 s.
24. Joseph, S., & Mathew, B. (2015). Microwave-assisted green synthesis of silver nanoparticles and the study on catalytic activity in the degradation of dyes. Journal of Molecular Liquids, 204, 184-191.
25. Konvičková, Z., Holišová, V., Kolenčík, M., Niide, T., Kratošová, G., Umetsu, M., & Seidlerová, J. (2018). Phytosynthesis of colloidal Ag-AgCl nanoparticles mediated by Tilia sp. leachate, evaluation of their behaviour in liquid phase and catalytic properties. Colloid and Polymer Science, 296, 677-687.
26. Korkut, D. S., Korkut, S., Bekar, I., Budakçı, M., Dilik, T., & Çakıcıer, N. (2008). The effects of heat treatment on the physical properties and surface roughness of Turkish hazel (Corylus colurna L.) wood. International Journal of Molecular Sciences, 9(9), 1772-1783.
27. Krishnaraj, C., Jagan, E. G., Rajasekar, S., Selvakumar, P., Kalaichelvan, P. T., & Mohan, N. J. C. S. B. B. (2012). Synthesis of silver nanoparticles using Acalypha indica leaf extracts and its antibacterial activity against water borne pathogens. Colloids and Surfaces B: Biointerfaces, 76(1), 50-56.
28. Kroon, L. P. N. M., Brouwer, H., De Cock, A., WAM., & Govers, F. (2012). The Phytophthora genus anno 2012. Phytopathology, 102(4), 348-364.
29. Lee, K. J., Jun, B. H., Choi, J., Lee, Y., Joung, J., & Oh, Y. O. (2007). Environmentally friendly synthesis of organic-soluble silver nanoparticles for printed electronics. Nanotechnology, 18(33), 335601.
30. Lin, Y.L., Wang, W.Y., Kuo, Y.H., & Chen, C. F. (2000). Nonsteroidal constituents from Solanum incanum L. Journal of the Chinese Chemical Society, 47(1), 247-251.
31. Luther, W. (2006). International Strategy and Foresight Report on Nanoscience and Nanotechnology. Final Report, 49p.
32. Meng, Q. X., Cui, X. L., Bi, Y., Wang, Q., Hao, J.J., & Liu, X.L. (2011). Biological and genetic characterization of Phytophthora capsici mutants resistant to flumorph. Plant Pathology, 60(5), 957-966.
33. Mishra, S., & Singh, H. B. (2015). Biosynthesized silver nanoparticles as a nanoweapon against phytopathogens: exploring their scope and potential in agriculture. Applied Microbiology and Biotechnology, 99, 1097-1107.
34. Molnar, T. J. (2011). Corylus. C. Kole (Ed.) Wild crop relatives: genomic and breeding resources. Forest Trees içinde (15-48 ss). Springer, Germany.
35. Mondal, P., & Purkait, M.K. (2017). Green synthesized iron nanoparticle-embedded pH-responsive PVDF-co-HFP membranes: optimization study for NPs preparation and nitrobenzene reduction. Separation Science and Technology, 52, 2338–2355.
36. Noroozi, M., Zakaria, A., Moksin, M. M., Wahab, Z.A., & Abedini, A. (2012). Green formation of spherical and dendritic silver nanostructures under microwave irradiation without reducing agent. International Journal of Molecular Sciences, 13(7), 8086-8096.
37. Polat, S., & Güney, Y. (2015) Türk fındığı’nın (Corylus colurna) Türkiye’deki yeni bir yayılış alanı. Akademik Sosyal Araştırmalar Dergisi, 3(18), 449-460.
38. Poulose, S., Panda, T., Nair, P.P., & Théodore, T., (2014). Biosynthesis of Silver Nanoparticles. Journal of Nanoscience and Nanotechnology, 14, 2038–2049.
39. Pourmortazavi, S.M., Taghdiri, M., Makari, V., & Rahimi-Nasrabadi, M. (2015). Procedure optimization for green synthesis of silver nanoparticles by aqueous extract of Eucalyptus oleosa. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 136, 1249-1254.
40. Rai, M., & Yadav, A. (2013). Plants as potential synthesizer of precious metal nanoparticles: progress and prospects. IET Nanobiotechnology, 7(3), 117-124.
41. Reddy, L. V. A., Wee, Y. J., Yun, J. S., & Ryu, H. W. (2008). Optimization of alkaline protease production by batch culture of Bacillus sp. RKY3 through Plackett–Burman and response surface methodological approaches. Bioresource Technology, 99(7), 2242-2249.
42. Rodriguez-Sanchez, L., Blanco, M.C., & López-Quintela, M.A. (2000). Electrochemical synthesis of silver nanoparticles. The Journal of Physical Chemistry B, 104(41), 9683-9688.
43. Riethmüller, E., Könczöl, Á., Szakál, D., Végh, K., Balogh, GT., & Kéry, Á. (2016). HPLC-DPPH screening method for evaluation of antioxidant compounds in Corylus species. Natural Product Communications, 11(5), 4-641.
44. Saleeb, N., Robinson, B., Cavanagh, J., Ross, J., Munir, K., & Gooneratne, R. (2020). Antioxidant enzyme activity and lipid peroxidation in Aporrectodea caliginosa earthworms exposed to silver nanoparticles and silver nitrate in spiked soil. Environmental Toxicology and Chemistry, 39(6), 1257-1266.
45. Siddiqi, K.S., Husen, H. & Rao, R.A.K. (2018). A review on biosynthesis of silver nanoparticles and their biocidal. Journal of Nanobiotechnology, 16:14. https://doi.org/10.1186/s12951-018-0334-5
46. Sreeram, K. J., Nidhin, M., & Nair, B. U. (2008). Microwave assisted template synthesis of silver nanoparticles. Bulletin of Materials Science, 31, 937-942.
47. Thakkar, K.N., Mhatre, S. S., & Parikh, R. Y. (2010). Biological synthesis of metallic nanoparticles. Nanomedicine: Nanotechnology, Biology and Medicine, 6(2), 257-262.
48. Tosun, S., (2012). Cadde (Yol) Ağacı Olarak Amerika’da ve Avrupa’da Popülerleşen Türk Fındığı (Corylus colurna L.). Orman ve Av Dergisi, 3, 22-25.
49. Türkkan, M. (2013). Antifungal effect of various salts against Fusarium oxysporum f. sp. cepae, the causal agent of Fusarium basal rot of onion. Journal of Agricultural Sciences, 19(3), 178-187.
50. Wacławek, S., Gončuková, Z., Adach, K., Fijałkowski, M., & Černík, M. (2018). Green synthesis of gold nanoparticles using Artemisia dracunculus extract: control of the shape and size by varying synthesis conditions. Environmental Science and Pollution Research, 25(24), 24210- 24219.
51. Wawra, S., Belmonte, R., Löbach, L., Saraiva, M., Willems, A., & van West, P. (2012). Secretion, delivery and function of oomycete effector proteins. Current Opinion in Microbiology, 15(6), 685-691.
52. Yiğit, U., & Türkkan, M. (2023). Antifungal activity and optimization procedure of microwave-synthesized silver nanoparticles using linden (Tilia rubra subsp. caucasica) flower extract. International Journal of Chemistry and Technology, 7(1), 195-207.