Recently, researchers have begun focusing much attention on the fabrication of organic-inorganic nanostructured composite materials used in different fields, such as photovoltaic devices, opthoelectronic circuits, solar cells, lithography, sensors, and medicine [1, 2, 3, 4, 5, 6, 7]. Electrospinning is an important method used extensively among the many methods for fabricating nanofiber composite materials due to their simple structure and low cost. Due to these features, recently, it is used extensively to fabricate nanofiber materials in the tapes used for healing the wounds caused by bacteria and viruses in medical [8, 9, 10, 11, 12]. In these studies, the most used compound by researchers is TiO2 which is one of the inorganic-compound with a photocatalytic property causing the disintegration of organic materials under the sunlight UV. So, this compound has been studied in detai[13, 14, 15]. On the other hand, some researchers have fabricated TiO2 nanofibers doping it with different metals such as aluminum, cerium, platinum, silver etc. because of its being a semiconductor with broadband range achievable by doping with these metals. Among these elements, silver is the most used material doped into TiO2. Many researchers have studied Ag-doped TiO2 nanofibers because they are antibacterial agents in biomedical materials owing to their powerful antibacterial activity [16, 17, 18, 19, 20, 21].
In this report, we have fabricated undoped and doped with different percent of Ag-TiO2 nanofibers by the electrospinning method. Then, we have investigated the anti bacterial activities of the undoped and Ag-doped TiO2 nanofibers on S. aureus bacteria. In addition, to explain the photocatalytic properties of the fabricated nanofiber samples have been characterized by scanning electron microscopy energy dispersive analysis (SEM-EDX), differential thermal agency/thermal gravimetric analysis (DTA/TG), and X-ray diffraction (XRD) methods.
2.1 Fabrication of pure and Ag-doped TiO2 nanofibers
To prepare pure and with different content Ag-doped TiO2 nanofibers (1%, 2%, 3%, 4%, 5%), we used precursor materials of AgNO3 (Merck, 99.9%, Mw = 62.004 g/mol), Ti[OCH(CH3)2]4 (Aldrich, 99.999%, Mw = 284.22 g/mol), CH3CH2OH (Sigma-Aldrich absolute, 99:8% Mw = 46.07 g/mol), and PVP (Aldrich, (C6H9NO)x, Mw = 1,300,000). First, we prepared TiO2 solution which we will use as a stock solution and also to fabricate pure TiO2 nanofibers. In order to prepare stock TiO2 solution, first, 3.6 mL of titanium isopropoxide and 2.4 mL of diethanolamine were added into 100 mL of ethanol. Then, we began to stir this solution with a magnetic stirrer and added 0.24 mL polyethylenglycol dropwise for 1 hour. In the second step, we prepared the solution which will be used for the fabrication of Ag-doped TiO2 nanofibers. We dissolved 10 g of PVP into 100 mL ethanol and stirred this solution (stock PVP solution) about 1 hour with a magnetic stirrer until an homogeneous solution was obtained. Then, 1%, 2%, 3%, 4%, 5% AgNO3 were dissolved into DMF solutions contained in five separate beakers and stirred half an hour with a magnetic stirrer to obtain homogeneous solutions. After preparing these two solutions, we prepared five different solutions as follows; to obtain the solution that we will use for the fabrication of the 1% Ag-doped TiO2 fibers, we took 2.41 g from the stock TiO2 solution and 12.46 g from the stock PVP solution added them together and stirred with the magnetic stirrer until it was homogeneous. Similarly, the other solutions that will be used to fabricate 2%, 3%, 4%, 5% Ag-doped TiO2 nanofibers were obtained following the same processes. Finally, loading these obtained homogeneous solutions and the pure TiO2 solution obtained before into a plastic syringe of the pump of the electrospinning set-up, we fabricated Ag-doped and pure TiO2 nanofiber samples, respectively. All samples have been produced at constant voltage, 20 kV, at constant heights, 8 cm, and with a constant flow rate, 0.1 mL/h.
2.2 Determination of the antibacterial effect of nanofibers
After completing the fabrication of the nanofiber samples, we applied two different methods to see their antibacterial effect on S. aureus bacteria known as Disk Diffusion Method and Baird Parker Agar Plate. We, first, applied the disk diffusion method to see the effect of the concentration of Ag on the bacteria as following: To do this, 1% and 5% silver-doped TiO2 solutions were immersed on each of the cellulose and silicon substrates. Then, 100 μl 0.5 Mc Farland turbidity of S.aureus (ATCC 25923), suspensions were spread on to petri plates. After this step, all samples which will be tested were cut into 6x6 mm pieces and placed on to the plates and incubated aerobically for 24 hours at 35±2oC. After incubation, growth inhibition zone diameters around the samples were measured with a ruler. Different diameters of the inhibition zones around the discs have demonstrated the antimicrobial effects of the tested substance.
After completing this measurement, we applied the Baird Parker Agar Plate method for the fabricated Ag-doped nanofibers. To do this we used the following procedure: Counting of S. aureus bacteria was carried out using Baird Parker agar with spread plate method. Firstly S. aureus ATCC 6538 strain was reproduced in nutrient broth for 24 hours at 37°C. Ag-doped TiO2 nanofiber samples produced on lamella. After, each sample surface was inoculated to be 106 kob / mL bacteria in cm2. The sample from reproduced microorganism was spread homogeneously to the surface with a sterile pipette. After the samples are kept in the appropriate time and environment, samples were taken from the surface with the help of a sterile swap of each sample and the swaps were transferred into tubes with sterile 10 mL ringers in the tubes. After preparing serial dilutions, Baird Parker agar plates were inoculated with each of the dilution of bacteria with the help of a sterile drigalski spatula so sowing with each pair being parallel to each other and spread homogeneously.
The broth to be used in the analyzes was sterilized in an autoclave at 121°C for 20 minutes at 1 atmosphere pressure, then cooled to room temperature. In this way, for each sample to be plated, 3 dilutions of darkness, sunlight, and UV were prepared. After the media was expected to absorb the sample, the media was allowed to incubate for 24 h at 37°C. After incubation, black colonies with white zones around 0.5 mm denier were counted and the number of bacteria in cm2 of samples was calculated according to the following formula [22, 23, 24].
Here; N;, the number of microorganisms in 1 gram or 1 milliliter; C, Total number of colony counted in all petri plates; V; volume transferred to petri plates counted (mL); n1, the petri plate where the first dilution is done is counted: n2, The petri plate which is counted in the second dilution; d, the dilution ratio of the two consecutive dilutions of the census.
Morphology, crystal structure, and thermal properties of the fabricated fibers were performed by scanning electron microscope (SEM) (Leo 1430 VP), X-ray diffraction (XRD) (XRD 6000-Shimadzu) with CuKα radiation (λ = 1.5418 Å) at 40 kV and 100 mA, and differential thermal analysis/thermal gravity (DTA/TG) (Netzsch STA449F3) measurements.
Ethical approval: The conducted research is not related to either human or animals use.
3 Results and Discussion
3.1 SEM measurements
SEM images of 1, 2, and 3, 4, 5 wt % Ag-doped TiO2 nanofibers and their measured average thicknesses are given in Figure 1 and Table 1, respectively. As seen from Figure1, all nanofibers are straight, smooth, and uniform long fibers and their thicknesses are at the nanometer scale. Images of nanofibers taken with scanning electron microscopy (SEM) have been analyzed to study the fiber morphology and to determine distributions of the diameter fiber thicknesses using FibraQuant 1.3 Software.
As seen Table 1 and Figure 1, diameters of Ag/TiO2 nanofiber samples are at the nanometer scale. The measured average diameter of these nanofibers depends on the concentration of Ag. The diameters of the nanofibers increased with increasing Ag concentration (up to 3%). Similar mechanisms and results were reported in our previous study .
3.2 XRD measurements
The XRD patterns of the pure TiO2 nanofibers and 5% Ag doped TiO2 nanofibers thermally treated at 500°C are shown in Figure 2 a, b, respectively. As it is known from the literature that pure TiO2 nanofibers thermally treated below 500°C have the crystalline both anatase and rutile  . As seen from Figure2, there is a decrease in the rutile structure in 5% Ag-doped TiO2 nano fiber compared to pure TiO2 nanofiber.
3.3 DTA /TG measurements
Chemical properties of the sample materials have been determined by DTA/TG measurements. DTA and TG analysis of pure TiO2 and 5 wt % Ag doped TiO2 nanofibers show in Figure 3. As seen from these figures, mass losses of nanofibers are different. The broad endothermal peak in the range of 400-500°C corresponded to the enhanced crystallization of anatase rather than an anatase→rutile phase transition. The TG curve recorded 62.9% significant weight losses in the temperature range 25-600°C, which are mainly due to dehydration of physically absorbed water, volatile organic solvent and crystallization water in the as-prepared Ag–TiO2 samples.
3.4 Disc diffusion method and baird parker agar plate method measurements
The experimental results of diameters of inhibitory zones against bacteria obtained from disc diffusion method for 1% and 5% Ag-doped nanofibers are given in Table 2.
As seen in Table 2, diameters of inhibition zones of Ag-doped TiO2 nanofibers (1% and 5%) on the silicone substrate for S. aureus were measured as 27 mm and 25 mm, respectively. Similarly, diameters of inhibition zones of Ag-doped TiO2 nanofibers (1% and 5%) on the cellulose substrate for S. aureus were measured as 25 mm and 31 mm, respectively. The largest zone diameter (31 mm) was obtained for 5% Ag-doped on the cellulose substrate. The inhibition zone diameters for all samples on culture plates are seen in Figure 4. According to the results, excellent antibacterial effects were obtained against S. aureus.
The experimental results of the number of bacteria obtained from Baird Parker Agar Plate method for Ag-doped TiO2 nanofibers 1%, 2%, 3%, 4% and 5% are given in Table 3. As seen in Table 3, the number of bacteria decreases with increasing silver content under UV, sunlight and dark, until it goes completely to zero. As a result, we can say that Ag-doped TiO2 nanofibers have very good antibacterial effect on S. aureus bacteria.
Undoped and Ag-doped TiO2 nanofibers were produced by using the electrospinning method. The results obtained from the characterization measurements of the samples are given as following;
The antibacterial properties of the produced nanofibers were investigated by using two methods (Baird Parker agar plate and disc diffusion). S. aerus (ATCC 6538) strain was used as the Baird Parker agar plate method and S. aerus (ATCC 25923) strain was used as the disc diffusion method. Increasing silver content, almost all of the bacteria have disappeared under UV and sunlight and the dark.
This was measured using the inhibition zones diameters for all samples on culture plates. According to the results, good antibacterial effects were obtained. The biggest zone diameter was obtained with 31 mm for S. aureus.
The morphology and structure of undoped and Ag-doped TiO2 nanofibers was confirmed by SEM, XRD, and DTA/TG techniques.
This work was supported by Suleyman Demirel University via Research Project Coordination Unit [BAP PROJECT-4196-YL1-14].
Li L., Sui H., Zhao, K., Zhang W., Li X., Liu S., et.al., Preparation of carbon nanofibers supported MoO2 composites electrode materials for application in dye-sensitized solar cells, Electrochim. Acta, 2018, 259, 188-195. Web of ScienceCrossrefGoogle Scholar
Krysovaa H., Trckova-Barakovab J., Prochazkaa J., Zukala A., Maixnerc J., Kavana L., Titania nanofiber photoanodes for dye-sensitized solar cells, Catal. Today, 2014, 230, 234–239. Web of ScienceCrossrefGoogle Scholar
Yamada K., Suwa Y., Katagiri C., Nakayama K., High vertical carrier mobility in the nanofiber films of a phthalocyanine derivative and its application to vertical-type transistors, Org. Electron, 2018, 53, 320–324. CrossrefWeb of ScienceGoogle Scholar
Ai Y,, Loua Z., Chena S., Chen D., Wang M. Z., Jiangd K., Shena G., All rGO-on-PVDF-nanofibers based self-powered electronic skins, Nano Energy, 2017, 35, 121–127. Web of ScienceCrossrefGoogle Scholar
Ramesh S., Kim S. H., Kim J.H., Cellulose–Polyvinyl Alcohol–Nano-TiO2 Hybrid Nanocomposite: Thermal, Optical, and Antimicrobial Properties against Pathogenic Bacteria, Polym. Plast. Technol. Eng., 2018, 57, 669–681. CrossrefWeb of ScienceGoogle Scholar
Malwala D., Gopinath P., Efficient adsorption and antibacterial properties of electrospun CuO-ZnO composite nanofibers for water remediation, J. Hazard. Mater., 2017, 321, 611–621. CrossrefPubMedWeb of ScienceGoogle Scholar
Zhang Z., Wu Y., Wang Z., Zhang X., Zhao Y., Sun L., Electrospinning of Ag Nanowires/polyvinyl alcohol hybrid nanofibers for their antibacterial properties, Mater. Sci. Eng. C, 2017, 78, 706–714. Web of ScienceCrossrefGoogle Scholar
Hassan M. S., Amna T., Al-Deyab S. S., Kim H. C., Oh T. H., Khil M. S., Toxicity of Ce2O3/TiO2 composite nanofibers against S. aureus and S. typhimurium: A novel electrospun material for disinfection of food pathogens, Colloids Surf. A Physicochem. Eng. Asp., 415, 2012, 268– 273. CrossrefWeb of ScienceGoogle Scholar
Nthumbi R. M., Ngila J. C., Electrospun and functionalized PVDF/PAN nanocatalyst-loaded composite for dechlorination and photodegradation of pesticides in contaminated water, Environ. Sci. Pollut. Res., 2016, 23, 20214–20231. Web of ScienceCrossrefGoogle Scholar
Barakat N. A. M., Kanjwal M. A., Al-Deya S. S., Chronakis I. S., Kim H. Y., Influences of Silver-Doping on the Crystal Structure, Morphology and Photocatalytic Activity of TiO2 Nanofibers, Mater. Sci. Appl., 2011, 2, 1188-1193. Google Scholar
Ramesha S., Kima H. S., Kim J. H., Cellulose–Polyvinyl Alcohol–Nano-TiO2 Hybrid Nanocomposite: Thermal, Optical, and Antimicrobial Properties against Pathogenic Bacteria, Polym-Plast. Technol., 2018, 57, 669–681. CrossrefWeb of ScienceGoogle Scholar
Caratão B., Carneiro E., Sá P., Almeida B., Carvalho S., Properties of Electrospun TiO2 Nanofibers, J. Nanotechnol., 2014,1-5. Google Scholar
Cicek Bezir N., Evcin A., Kayalı R., Kasıkcı Ozen M., Oktay A., Investigation of structural, electronic and optical properties of pure and Ag-doped TiO2 nanofibers fabricated by electrospinning, Cryst. Res. Technol., 2016, 51, 65–73. CrossrefWeb of ScienceGoogle Scholar
Evcin A., Ç. Bezir N., Kayalı R., Kasıkçı M., Oktay A., Characteristic Properties of Dy-Eu–Ag co-Doped TiO2 Nanoparticles Prepared by Electrospinning Processes, Acta. Phys. Pol. A, 2015, 128, 303-306. CrossrefWeb of ScienceGoogle Scholar
Mishra S., Ahrenkiel S. P., Synthesis and Characterization of Electrospun Nanocomposite TiO2 Nanofibers with Ag Nanoparticles for Photocatalysis Applications, J. Nanomater., 2012, 1-6. Web of ScienceGoogle Scholar
Aboelzah A., Azad A. M., Vijay G., Necrosis of Staphylococcus aureus by the Electrospun Fe- and Ag-Doped TiO2 Nanofibers, Int. Sch. Res. Notices, 2012, 1-11. Google Scholar
Kanjwal M. A., Barakat N. A. M., Sheikh F. A., Khil M. S., Kim H. Y., Functionalization of Electrospun Titanium Oxide Nanofibers with Silver Nanoparticles: Strongly Effective Photocatalyst, Int. J. Appl. Ceram. Technol., 2010, 7, E54–E63. Web of ScienceGoogle Scholar
Barakat N. A. M., Kanjwal M. A., Al-Deyab S. S., Chronakis I. S., Kim H. Y., Influences of Silver-Doping on the Crystal Structure, Morphology and Photocatalytic Activity of TiO2 Nanofibers, Mater. Sci. Appl., 2011, 2, 1188-1193. Google Scholar
J. T. Nickerson, A. J. Microbiology of Food and Food Prossesing. American Elsevier Publishing Company. New York. USA. (Sinskey), 1974. Google Scholar
Anonim, Census of milk and products and coliforms. Part 1, Colony Counting Technique at 30°C. General Directorate of Turkish Standards, Ankara, TS 6930, 1989. Google Scholar
K. Halkman, Food Microbiology Applications, Başak Printing and Promotion Services Limited Company, Ankara, 2005. Google Scholar
About the article
Published Online: 2018-08-20
Conflict of interest: Authors state no conflict of interest.
Citation Information: Open Chemistry, Volume 16, Issue 1, Pages 732–737, ISSN (Online) 2391-5420, DOI: https://doi.org/10.1515/chem-2018-0081.
© 2018 Nalan Çiçek Bezir et al., published by De Gruyter. This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 License. BY-NC-ND 4.0