Nanochemistry and biotechnology 
«Green» synthesis and bioimaging applications of Ag2S nanoparticles 
Borovaya M.N1., Horiunova I.I1., Naumenko A.P2., Blume Y.B1., Yemets A.I1
1. Institute of Food Biotechnology and Genomics, NAS of Ukraine. Osypovskogo Street, 2a, Kyiv-04123, Ukraine.
E-mail: yemets.alla@nas.gov.ua, marie0589@gmail.com
2Faculty of Physics, Taras Shevchenko National University. Prospect acad. Glushkova, 4, Kyiv-03022, Ukraine. 

Ag2S is one of the most important transition metal sulfide semiconductor nanomaterials due to its direct and narrow band gap (1.5 eV) and good chemical stability. It possesses unique semiconducting, optical, and electrical properties and is highly stable. Owing to these features, silver sulfide nanoparticles are broadly used in solar cell batteries, thermoelectric sensors, infrared detectors, and superionic conductors. These nanoparticles are non-toxic to living organisms [1]. The great potential of practical usage of Ag2S nanoparticles brought into existence numerous protocols for their preparation. Various physical and chemical methodologies were developed for silver sulfide nanoparticles production such as molecular beam and lithography techniques, homogeneous precipitation route etc. Most of these methods are cumbersome and ecologically unfriendly. They involve the use of toxic and combustible precursors, are accomplished under oxygen and/or a water-free atmosphere requiring high temperatures and pressures, and the resulting nanoparticles can become unstable or agglomerate upon interaction with biological media components or biomolecules. Therefore it is important to develop environmentally friendly synthesis techniques that can generate particles with the desired properties [2]. The appropriate narrow bandgap and low toxicity of Ag2S quantum dots facilitated them to be a promising contrast agent for fluorescence imaging. So the purpose of this study is to develop a new eco-friendly method for «green» synthesis of Ag2S nanoparticles by basidiomycetes fungus Pleurotus ostreatus and investigate their luminescent properties for bioimaging applications. 

Ag2S nanoparticles were synthesised using Pleurotus ostreatus. Cultivation of P. ostreatus was carried out in 100 mL Erlenmeyer flask with 50 mL of Glucose Yeast Peptone liquid medium under 28˚С for 10 days. On the tenth day the surface mycelium was aseptically separated from the culture liquid and washed repeatedly with sterile distilled water (not less than 5 times) to eliminate the residual culture medium. 0,025 М solutions of AgNO3 and Na2S were poured into the flasks with fungal mycelium in order to produce Ag2S. After adding sodium sulphide, the solution turned brown. Obtained solution was filtered through nitrocellulose Millipore (USA) filter (pore diameter - 0.45 mm) for further optical and structural studies. Absorption spectra of obtained nanoparticles were measured using a spectrophotometer Specord UV-VIS. Characterization of Ag2S nanoparticles was carried out using electron microscope JEOL JEM-2100F (Japan). It was observed that synthesized nanoparticles were spherical in shape, have a size from 10 to 25 nm.
Absorption maximum corresponded to 470 nm. 
For bioimaging studies 4-day roots of Allium cepa were fixed in acetoalcohol in a ratio of 1:3 for an hour. Then roots were treated by freshly prepared Ag2S nanoparticles for 24 h under 60˚C.  Luminescent properties, penetration and localization of Ag2S quantum dots in root cells were investigated by laser scanning microscope 510 META (Carl Zeiss, Germany) with lens 40X using the following filters: LP 505; LP 560, excitation wavelength - 488 nm. Obtained results demonstrated that silver sulfide nanoparticles can be used as an alternative to organic dyes. They have intense luminescence in the wavelength range of 540-633 nm (green-red region of the spectrum). Produced quantum dots were 10-25 nm in diameter. Ag2S nanoparticles are localized mostly in the nucleus of epidermal and meristematic cells of the root tips. Therefore, the present work demonstrates an efficient biosynthesis of extracellular highly stable, spherical silver sulfide nanoparticles produced by «green» synthesis with the use of fungi mycelium. Obtained quantum dots were spherical within 10-25 nm in diameter. They are promising luminescent probes for cell biology applications considering their luminescent properties and less toxicity to the environment.

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