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B. Adebayo-Tayo et al. Heliyon 5 (2019) e02502 given rise to the biosynthesis of nanoparticles [12]. The wide use of synthesized nanoparticles in human activities create the need for a method which will not pose threat to the environment and humans as a result of their exposure to the nanoparticles, hence the biological method [13, 14]. The use of metallic nanoparticles in modern technologies is based on their unique characteristics such as surface Plasmon features, morphol- ogies and interesting physicochemical properties among others [15]. Silver nanoparticles compare to other metallic nanoparticles as a result of their outstanding antimicrobial properties such as broad spectrum and surface Plasmon resonance are of great importance in modern nano- science and nanotechnology [16, 17]. Unlike standard antimicrobial agents, low doses of silver nanoparticles are needed in the treatment of diseases [18]. The interaction of silver nanoparticles with microorgan- isms gives rise to the release of silver ions which are potential destroyers of microbial cells due to their ability to deactivate enzymes in microbial cell and membrane permeability disruption leading to lysis and apoptosis [19, 20]. Silver nanoparticles also possess antiinflammatory, anti- platelets and antiangiogenesis potentials among other thus making their application in medical field paramount [21]. The use of green algae in the production of silver nanoparticles have been reported to be efficient due to the composition of the microalgae as well as little or no toxic effect of the synthesized nanoparticles and outstanding features of the nano- particles biosynthesized [22]. This work aims at biosynthesizing silver nanoparticles from methanol extract of the green alga; Oscillatoria sp. and characterizing the biosynthesized silver nanoparticles. 2. Materials and methods All chemicals were analytical grade products purchased from Sigma –Aldrich (St.Louis, MO, USA). 2.1. Sample collection and extraction of the bioactive compound from the algae The blue green algae (Oscillatoria sp.) sample was collected from the wall of the walkway of the Department of Theatre Art, University of Ibadan, Ibadan, Nigeria (Latitude 03 434̍ 43̍ E and longitude 07 26̍ 41̋ N) with sterile scalpel from where thick mass was visually observed and transported to the laboratory, Botany Department, University of Ibadan for further analysis. The scraped algal sample was washed under running tap water to remove sand particles adhering to the sample and thereafter rinsed with sterile distilled water. The washed algal biomass was dried at room temperature for one week, ground into powder using blender and stored at room temperature for further analysis. The test pathogens; S. aureus ATCC29213, E. coli ATCC 11775, E. coli ATCC 35218, P. aeruginosa ATCC27853, Citrobacter sp., S. typhi ATCC 14028 and Bacillus cereus were obtained from the Department of Microbiology and Department of Pharmacy, University of Ibadan, Ibadan Oyo state. Brine shrimp eggs were obtained from the Biochemistry Department, University of Ibadan and the Sea water was obtained from the Atlantic Ocean in Lagos, Nigeria. The seawater was 29.4 C, pH 8.2, Salinity 309 þ 0.03PSU and Turbidity 15.5 NTU. For the extraction of the bioactive compound in the algae samples, the algae were washed with distilled water to remove extraneous materials, dried properly and grind into a fine powder. Soxhlet extraction of 200 grams of the dried sample was done using 300 mL methanol as solvent. The marc was filtered and solvents were removed under reduced pressure in a rotary evaporator. Dark brown pastes obtained were weighed and stored in a refrigerator at 4 C. 2.2. Biosynthesis of silver nanoparticles The silver nanoparticles were biosynthesized by adding 10 mL of 1 g/ mL of the methanol extract of Oscillatoria sp. into 90 mL of 0.1 mM of AgNO3 solution [23]. The mixture was kept in the dark at room temperature for 72 h. The samples were used for further analysis. 2.3. Characterization of the biosynthesized OsSNPs Visual detection of the greenly synthesized OsSNPs was done by observing the mixture for a change in colour in comparison to the control samples. UV–Visible spectrophotometric analysis of the OsSNPs solutions were determined at room temperature using UV–Vis spectrophotometer (a Lambda 25-Perkin Elmer, Waltham, MA, USA) with a resolution of 0.5 nm. The absorbance of the sample was read at the wavelengths of 200–800 nm [24]. The chemical structure of the OsSNPs samples was analyzed using FTIR spectroscopy (Shimadzu). Two milligram of the dried samples was taken ground with KBr salt at 25 C and pressed into a mold to form pellet. The spectra were recorded at a wave range of 500–4000 cm-1 and at resolution of 4 cm1 [25]. The morphological structure of the OsSNPs was analyzed by Scanning Electron Microscopy (SEM). The dried samples were coated with gold using a coater (JEOL, Akishima-shi, Japan, and Model number JFC- 1600). The images of SNPs were taken in a SEM (ZEISS EVO-MA 10, Oberkochen, Germany). X ray diffraction patterns (XRD) were obtained in a Siemens Kristalloflex diffractometer using nickel filtered CuKα ra- diation from 4 to 70 (2θ angle). Thermogravimetry (TG) analysis was carried out using dried OsSNPs in SDT 2960 device from TA Instruments. The nanoparticles were heated in open alumina pans from 40 to 600 C, under an oxidant atmosphere (O2), flux of 50 mL/min and a heating rate of 10 C/min were used. The estimation of the silver content in OsSNPs was done using the residue at 600 C. The elemental composition of OsSNPs was determined using an En- ergy Dispersion X-ray (EDX). The dried OsSNPs powder was used for the analysis and the EDX analysis software was sourced from Oxford in- struments analytical. All measurements were performed at accelerated voltage of 10 KV. Particle size analyzer (Zetasizer Nano ZS, Malvern Instruments Limited, Worcestershire, United Kingdom) was used to determine the particle size distribution and surface charge of the OsSNPs. The analysis was done at 25 C with 90 detection angle and 633 nm. Hydrodynamic diameter and polydispersity index were measured as a function of time. Prior to the analysis, the OsSNPs was suspended in sterile water and sonicated for 15 min. 2.4. Antibacterial potential of the OsSNPs The antibacterial potential of the OsSNPs was evaluated using the Agar Well Diffusion method [26] using S. aureus ATCC29213, E. coli ATCC 11775, E. coli ATCC 35218, P. aeruginosa ATCC27853, Citrobacter sp., S. typhi ATCC 14028 and Bacillus cereus as the test pathogens. Escherichia coli ATCC 11775 is a reference type strain which was first isolated from urine sample of a Danish patient's in 1941 [27]. Escherichia coli ATCC 35218 is a strain recommended by NCCLS for used as a quality control organism for the assay of betalactam antibiotics, evaluation of Mueller Hinton agar and the quality control strain for susceptibility testing [28, 29]. The isolate was cultured overnight in peptone water. 18 hrs old culture of the isolate was seeded on Mueller - Hinton Agar (Lab M Ltd., UK) plates. Uniform wells were cut on the dried agar plate using a sterile cork -borer of diameter 7 mm. Each well was filled with 20 μL of the biosynthesized OsSNPs. AgNO3 solution (1 mM) and the methanol extract of Oscillatoria sp. were put into respective wells as negative controls. The inoculated plates were incubated at 37 C for 24 h. After incubation the plates were observed for zones of inhibition (ZOI) around the wells. ZOI diameters (mm) greater than 1 mm were considered positive [30]. 2.5. Antibiofilm activity of OsSNPs The antibiofilm activity of the biosynthesized SNPs was determined according to the method as described by [31] with slight modification. 2PDF Image | Green synthesis of silver nanoparticle Oscillatoria extract
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