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

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

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