Single Cell Level Microalgal green synthesis of silver nanoparticles: Confocal Microscopy and Digital Image Analysis

Year 2019, Volume: 2 Issue: 2, 56 - 60, 06.12.2019

Betül Yılmaz Öztürk , Derviş Öztürk

Abstract

Nanoparticles are attracting
increasing attention due to their unusual and fascinating properties, which are
strongly influenced by their size, morphology and structure. Among the
developed nanoparticles, silver (Ag) nanoparticles are pertaining to have a
wide range of application in the fields of physical, chemical and biological
science. Physical
and chemical methods are used to synthesize such nanomaterials, among the
various known synthesis methods, biosynthesis of silver nanoparticles is
preferred as it is environmentally safe, low cost and less toxic. In
particular, the synthesis of nanoparticles in the cell can be achieved in a
standard size and shape. In the present work, the coccoid green algae Chodatodesmus mucronulatus was used as a reducing agent for the
synthesis of intracellular nanostructure silver particles (Ag-NPs). Algae are
with autofluorescence characteristics. These properties are known to be due to
chlorophyll pigments. In this context, a
confocal laser scanning microscopy (CLSM) based method to assess to show that
the amount of chlorophyll decreases at microalgae is reported. ). During this process, changes in
the amount of chlorophyll a, b and
carotenoid of the Chodatodesmus mucronulatus were examined at 24
hours using UV-Vis spectrophotometer for 3 days. As a result, the amount of
carotenoid, especially with the onset of the reaction, decreased markedly.
After 72 hours of reaction, the amount of carotenoid decreased from 6,54 μg ml^-1
to 0,00 μg / ml, chlorophyll a
decreased 24,46 µg ml^-1 to 0,06 µg ml^-1,chlorophyll b decreased from 11,33 µg ml^-1
to 4,15 µg /ml. This change (pigment amount in cells) was also observed with a
confocal microscope every 24 hours. Using this technique, the effect of in-use
concentrations of chlorophyll autofluorescence was defined. Determination of
mean fluorescence intensity (MFI) per cell by collecting auto-fluorescence from
single cells in x, y and z dimensions permitted evaluation at single-cell
level. According to the results, there is a decrease in the amount of pigment
in the cell. This suggests that the pigments may be capping agents and trigger
nanoparticle synthesis.

Keywords

Microalgae, Chlorophyll, Confocal microscopy, Nanoparticle, Silver

Thanks

This work was studied at Eskisehir Osmangazi University Central Research Laboratory Application and Research Center (ARUM).

References

• Agarwal P, Gupta R, Agarwal N 2019. Advances in Synthesis and Applications of Microalgal Nanoparticles for Wastewater Treatment. J Nanotechnol, 2019:1-9.
• Chandler GT, Volz DC 2004. Semiquantitative confocal laser scanning microscopy applied to marine invertebrate ecotoxicology. Mar Biotechnol, 6:128–137.
• Dağlıoğlu Y, Öztürk BY 2019. A novel intracellular synthesis of silver nanoparticles using Desmodesmus sp.(Scenedesmaceae): different methods of pigment change. Rend. Lincei Sci. Fis. Nat., 1-11.
• Dubey M, Bhadauria S, Kushwah B 2009. Green synthesis of nanosilver particles from extract of Eucalyptus hybrida (safeda) leaf. Dig J Nanomater Biostruct, 4:537–543.
• Gomaa EZ 2017. Antimicrobial, antioxidant and antitumor activities of silver nanoparticles synthesized by Allium cepa extract: a green approach. J Genet Eng Biotechnol, 15(1): 49-57.
• Halbhuber K-J, König K 2003. Modern laser scanning microscopy in biology, biotechnology and medicine. Ann Anat Anat Anz, 185:1–20.
• Harter K, Meixner AJ, Schleifenbaum F 2012. Spectro-microscopy of living plant cells. Mol Plant, 5:14–26.
• Jegadeeswaran P, Shivaraj R, Venckatesh R 2012. Green synthesis of silver nanoparticles from extract of Padina tetrastromatica leaf. Dig J Nanomater Bios, 7(3): 991-998.
• Jena J, Pradhan N, Nayak RR, Dash BP, Sukla LB, Panda PK, Mishra BK 2014. Microalga Scenedesmus sp.: a potential low-cost green machine for silver nanoparticle synthesis. J Microbiol Biotechnol, 24:522–533.
• Khatami M, Pourseyedi S 2015. Phoenix dactylifera (date palm) pit aqueous extract mediated novel route for synthesis high stable silver nanoparticles with high antifungal and antibacterial activity. IET Nanobiotechnol., 9(4): 184-190.
• Khatami M, Varma RS, Heydari M, Peydayesh M, Sedighi A, Agha Askari H, Rohani M, Baniasadi M, Arkia S, Seyedi F, Khatami, S. 2019. Copper Oxide Nanoparticles Greener Synthesis Using Tea and its Antifungal Efficiency on Fusarium solani. Geomicrobiol. J., 1-5.
• Ko SH, Park I, Pan H, Grigoropoulos CP, Pisano AP, Luscombe CK, Frechet JMJ 2007. Direct nanoimprinting of metal nanoparticles for nanoscale electronics fabrication. Nano Lett, 7: 1869-1877.
• Li S, Shen Y, Xie A, Yu X, Qiu L, Zhang L, Zhang Q 2007. Green synthesis of silver nanoparticles using Capsicum annuum L. extract. Green Chem., 9: 852-858.Nancharaiah YV, Rajadurai M, Venugopalan VP (2007) Single cell level microalgal ecotoxicity assessment by confocal microscopy and digital image analysis. Environ Sci Technol, 41:2617–2621.
• Neu TR, Lawrence JR 1997. Development and structure of microbial biofilms in river water studied by confocal laser scanning microscopy. FEMS Microbiol Ecol, 24: 11-25.
• Öztürk BY 2019. Intracellular and extracellular green synthesis of silver nanoparticles using Desmodesmus sp.: their Antibacterial and antifungal effects. Caryologia. International Journal of Cytology, Cytosystematics and Cytogenetics, 72(1): 29-43.
• Ramkumar VS, Pugazhendhi A, Gopalakrishnan K, Sivagurunathan P, Saratale GD, Dung TNB, Kannapiran E 2017. Biofabrication and characterization of silver nanoparticles using aqueous extract of seaweed Enteromorpha compressa and its biomedical properties. Biotechnol Rep, 14: 1-7.
• Roldan M, Thomas F, Castel S, Quesada A, Hernandez- Marine M 2004. Noninvasive pigment identification in single cells from living phototrophic biofilms by confocal imaging spectrofluorometry. Appl. Environ. Microbiol., 70: 3745-3750.
• Salem JK, El-Nahhal IM, Najri BA, Hammad TM, Kodeh F 2016. Effect of anionic surfactants on the surface plasmon resonance band of silver nanoparticles: determination of critical micelle concentration. J. Mol. Liq., 223: 771-774.
• Sarafis V 1998. Chloroplasts: a structural approach. J Plant Physiol, 152:248–264.
• Shankar PD, Shobana S, Karuppusamy I, Pugazhendhi A, Ramkumar VS, Arvindnarayan S, Kumar G 2016. A review on the biosynthesis of metallic nanoparticles (gold and silver) using bio-components of microalgae: Formation mechanism and applications. Enzyme Microb. Technol., 95: 28-44.
• Song KC, Lee SM, Park TS, Lee BS 2009. Preparation of colloidal silver nanoparticles by chemical reduction method. Korean J Chem Eng, 26: 153-155.
• Sun C, Lee JSH, Zhang MQ 2008. Magnetic nanoparticles in MR imaging and drug delivery. Adv Drug Delivery Rev., 60: 1252-1265.
• Travan A, Pelillo C, Donati I, Marsich E, Benincasa M, Scarpa T, Semeraro S, Turco G, Gennaro R, Paoletti S 2009. Non-cytotoxic silver nanoparticle-polysaccharide nanocomposites with antimicrobial activity. Biomacromolecules, 10: 1429-1435.
• Venugopal K, Rather HA, Rajagopal K, Shanthi MP, Sheriff K, Illiyas M, RathEr RA, Manikandan E, Uvarajan S, Bhaskar M, Maaza M 2017. Synthesis of silver nanoparticles (Ag NPs) for anticancer activities (MCF 7 breast and A549 lung cell lines) of the crude extract of Syzygium aromaticum. J. Photochem. Photobiol., 167: 282-289.
• Yu Z, Pei H, Jiang L, Hou Q, Nie C, Zhang L 2018. Phytohormone addition coupled with nitrogen depletion almost tripled the lipid productivities in two algae. Biores Technol, 247:904–914.