Materials Science Forum Vols. 798-799 (2014) pp 69-74 Online available since 2014/Jun/30 at www.scientific.net © (2014) Trans Tech Publications, Switzerland doi:10.4028/www.scientific.net/MSF.798-799.69
Silver-doped Zirconia Nanoparticles as Possible Bactericide in Water Filters E. N. L. Rodrigues1,a, E.J.P. Miranda Jr2,b and M. M. Oliveira3,c 1,2
Department of Mechanics and Materials – DMM – IFMA – Brazil 3
a
Department of Chemistry – DAQ – IFMA – Brazil
[email protected],
[email protected],
[email protected],
Keywords: Pechini method, nanoparticles, zirconia, silver, bactericide.
Abstract. The objectives of this study were to produce ZrO2-based nanoparticles doped with silver, using the polymeric precursor method (Pechini), and to verify their bactericidal activity against Escherichia coli and Staphylococcus aureus. Zirconium and silver oxides were chosen due to their bactericidal activity. The oxides were characterized by X-ray diffraction and field emission gunscanning electron microscopy. The support for the oxides was a porous surface used in gravity filters. The method was not effective against the bacterium Escherichia coli in the sample subjected to UV light for 45 min, since it did not eliminate the bacterium, and did not inhibit colony growth and formation. However, the bactericidal activity test proved effective against Staphylococcus aureus, eliminating the bacterium when the sample was subjected to UV light for 45 min, thus preventing colony growth and formation. Introduction The World Health Organization has identified in order to improve access to drinking water is a major challenge in the developing world. Silver impregnated ceramic filters have been designed and developed to achieve the high demand for a water filtration cheaply and effectively especially in low-income population areas where there are practically no water treatment [1]. Ceramic material is traditionally used in gravity filters because it lends itself easily to the creation of a uniform structure of small pores. These ceramic structures can remove many contaminants from water, be exclusion according to pore size or by adherence to the pore walls. Silver is notable for its ability to be effective against bacteria even in small concentrations [2]. Craver and Smith demonstrated that antibacterial activity is enhanced by applying silver nanoparticles [3]. Sondi & Salopek-Sondi suggested that mechanisms antibacterial silver action depend on the direct contact with the cell wall of the bacteria [4]. Li et al. describe how the silver is adsorbed on the surface of the bacterium, interrupting a transfer of electrons of the membrane and effectively disabling the bacterium [5]. Some authors have demonstrated the effectiveness of these gravity filters in reducing Escherichia coli and other bacterial indicators in drinking water, and thus, the incidence of diarrheal diseases [6]. Feng et al. explain how DNA molecules of E. coli condense and lose their ability to replicate when exposed to silver ions. However, the antibacterial mechanism is not fully understood and does not explain the way some antiviral effects can be attributed to silver [7]. One of the differences of this work was the use of zirconia-base nanoparticles doped with silver to modify the surface of porous ceramic material, for the possible elimination of Escherichia coli and Staphylococcus aureus. In this study, we examined the microstructure of the porous ceramic material of the gravity filter after impregnation with nanometric particles, and assessed its bactericidal efficiency against E. coli and S. aureus. In addition, an analysis was made of the ZrO2 nanoparticles doped with silver for use in the impregnation of the filter element.
Our results indicate that the ZrO2-based nanoparticles doped with silver used in this study have a promising potential for use as a bactericidal agent in porous filter elements of gravity filters, thus contributing to the elimination of diseases in low income communities where these filters are used. Nevertheless, other variables must be considered, such as the percentage of the dopant, the nanoparticles impregnation method, and the method used in bactericidal tests. Experimental Materials The reagents used in the synthesis of the materials are listed in Table 1. Table 1. Reagents used in the preparation of the polymeric precursors. Reagent Citric acid Zirconium (IV) butoxide Silver nitrate Ethylene glycol
Formula C6H8O7
Supplier Synth
Purity [%] 99.5
Zr[O(CH2)3CH3]4
Aldrich
80
Ag(NO)3 HOCH2CH2OH
Aldrich J. T. Baker
95 99.9
Zirconium and silver citrates were prepared by the Pechini method [8], using a molar ratio of citric acid: metal cations of 3:1, which allows for the formation and stabilization of the metal citrate. In Pechini Method is formatted a polymer precursor in which the interest cations are randomly distributed in the solution. When polymerized, the cations form a solid three-dimensional network, avoiding the precipitation or segregation of ions of the formation of new phases during the synthesis of metal oxide. The materials were then analyzed gravimetrically to determine the concentration of zirconium ions and silver ions in their respective solutions. Resins were prepared from the zirconium and silver citrates by heating them to approximately 60ºC and then adding ethylene glycol in a proportion (by weight) of 60/40 citric acid/ethylene glycol (CA/EG). Methods X-ray diffractograms were recorded using a Siemens D500 X-ray diffractometer coupled to a LiF (100) monochromator, applying a scan rate of 2θ = 2º/ min, using Cu Kα (λ = 1.5406 Å) radiation and varying 2θ from 5º to 80º. Morphological and qualitative analyses were performed using a field emission gun-scanning electron microscope with a JEOL JSM-7500F energy dispersive X-ray spectroscopy (EDX). The bactericidal activity was tested using samples of the filter elements (dimensions of 1.5 cm x 1 cm). The samples were contaminated with two types of bacteria: E. coli (gram-negative) and S. aureus (gram-positive), after which they were exposed to light in a chamber equipped with a lamp emitting light in the ultraviolet (UV) region, and to direct sunlight. Two samples contaminated with E. coli were placed in Petri dishes in a QUIMIS Q216F20M lab test chamber equipped with a 20 W UV lamp, where they were left for 45 min (the time required to activate the silver-doped zirconium oxide). The same procedure was applied to the bacterium S. aureus. The two samples contaminated with S. aureus and exposed to UV light were removed using a swab moistened with sterile saline and transferred to Petri dishes containing 20 mL of BairdParker agar culture medium. These samples were incubated in a bacteriological incubator at 3537ºC and monitored for 48 hours.
To inoculate the samples contaminated with E. coli, they were placed in Petri dishes containing 20 mL of EMB agar culture medium and subjected to the same procedure as the one performed for S. aureus. Results and Discussion X-Ray Diffraction Figure 1 shows X-ray diffractograms of the ZrO2 powders containing concentrations of 0.25% and 0.5% of silver prepared by the Pechini method and calcined at 600 ºC for 2 h. Note that there was no change in the peak positions of the samples, indicating that the majority phase was monoclinic zirconia and the secondary phase tetragonal zirconia. The narrowing of the peaks and the intensity of the diffraction peaks and crystallinity indicate grain size growth. The peaks at around 2θ = 30.3º, 50.3º and 60.2º were attributed to the (011), (112) and (121) reflections of tetragonal zirconia, according to JCPDS card n°. 50-1089, while the peaks at 2θ = 24.3º, 28.2º and 31.5º correspond to the (110), ( 11) and (111) reflections of monoclinic zirconia, according to JCPDS card n°. 37-1484 [9, 10].
Figure 1. Diffractogram of ZrO2 doped with 0.5% of Ag and calcined at 600 °C/2 h. The crystallite size measured from the highest intensity peak using the Debye-Scherrer equation was 13.8 nm for the ZrO2 powder with 0.25% of silver and 18 nm for the powder containing 0.5% of silver calcined at 600ºC for 2 h. The results of this work are consistent with others reported in the literature [11]. Bactericidal tests of the filter elements Contamination and bactericidal activity test against S. aureus Figure 2 illustrates the results, showing Petri dishes with Baird-Parker agar containing samples collected from the surfaces of the filter element impregnated with silver-doped ZnO2.
Figure 2. Bactericidal activity test with the bacterium S. aureus: (a) S. aureus not exposed to UV light, (b) S. aureus exposed to UV light for 45 min, and (c) control. A comparison of samples (a) and (c) in Figure 2 indicates that the method of S. aureus contamination of the porous filter element impregnated with silver-doped ZnO2 was adequate, since sample (a) shows visible growth and colony formation, while the control (c) shows the absence of the bacterium. Sample (b) in Figure 2, which was exposed to UV light for 45 min, shows zero bacterial growth, and hence, no formation of S. aureus colonies. Therefore, in this case, the bactericidal activity of silver-doped ZnO2 was efficient. Scanning Electron Microscopy (SEM) SEM imaging of the porous filter element Figure 3 shows micrographs of the filter element under different levels of magnification after impregnation and heat treatment at 600 ºC for 2 h to produce zirconium oxide doped with 0.25% of silver on the surface of the material. As can be seen, the surface of the filter element impregnated with oxide shows a visible modification. The SEM analysis confirmed the formation of a layer of zirconium oxide doped with silver on the porous surface of the material.
( a )
( b )
( c )
Figure 3. SEM micrographs of the filter element impregnated with zirconium oxide doped with 0.25% of silver: (a) 100X, (b) 500X, and (c) 1000X magnification.
Contamination and bactericidal activity test against E. coli The surface of the filter element impregnated with silver-doped ZnO2 was analyzed, and the results of the bactericidal activity test against E. coli are shown in Table 2. Table 2. Result of the bactericidal activity test of silver-doped ZnO2 against Escherichia coli. Exposure Baird-parker agar culture Sample time medium Sample contaminated with E. coli not exposed to + UV light Sample contaminated with E. coli exposed to 45 min + UV light Control (EMB medium without sample)
-
Note: The (+) and (-) signs indicate, respectively, visible contamination and absence of contamination by E. coli. The results described in Table 2 are illustrated in Figure 4, which shows Petri dishes with Baird-Parker agar containing samples collected from the surfaces of the filter element impregnated with silver-doped ZnO2.
Figure 4. Bactericidal activity test with the bacterium E. coli: (a) E. coli not exposed to UV light, (b) E. coli exposed to UV light for 45 min, and (c) control. The results indicate that the gravity filter sample impregnated with silver-doped ZnO2 did not show the desired bactericidal activity in the sample contaminated with E. coli and exposed to UV light for 45 min, since this sample showed growth and formation of colonies of the bacterium. The reason for this may be the fact that the exposure time of the sample to UV light was insufficient to activate the oxide on the surface of the filter element, and the fact that E. coli is a gram-negative bacterium that is more resistant to this oxide. The control (c) showed the absence of the bacterium under study. Conclusions (1) The X-ray diffraction analysis indicated the presence of majority phase monoclinic zirconia and secondary phase tetragonal zirconia in the silver-doped powders prepared by the Pechini method and calcined in a conventional muffle furnace. (2) The SEM analysis of the filter elements of gravity water filters confirmed the formation of a layer of zirconium oxide doped with silver on the porous surface of the material.
(3) The bactericidal test of the filter element whose surface was impregnated with silver-doped zirconium oxide showed that the method was effective in the case of the sample contaminated with S. aureus and exposed to UV light for 45 minutes, since the bacterium was eliminated, preventing the formation of colonies. (4) The method was not effective in eliminating bacteria in the sample contaminated with E. coli and exposed to UV light for 45 minutes. This study indicated that zirconia nanoparticles doped with silver show a promising potential for application in the elimination of the bacterium Staphylococcus aureus. In the case of the bacterium Escherichia coli, the method can be improved by continuing the research and study of other variables, such as the amount of dopant, in this case silver, among other factors. References [1] WORLD HEALTH ORGANIZATION. Guidelines for Drinking-Water Quality. First Addendum to Third Edition, Volume 1 Recommendations, Geneva, 2006. [2] HILL, W.R.; PILLSBURY, D.M. Argyria: the pharmacology of silver. Baltimore: Williams and Wilkins Co; 1939. [3] CRAVER, V.A.; SMITH, J.A. Sustainable colloidal-silver-impregnated ceramic filter for pointof-use water treatment. Environ SciTechnol 42 (2008) 927–33. [4] SONDI, I.; SALOPEK, S.B. Silver nanoparticles as antimicrobial agent: a case study on E. coli as a model for Gram-negative bacteria. J. Colloid Interface Sci. 275 (2004) 177–182. [5] LI Q.; MAHENDRA, S.; LYON, D.Y.; BRUNET, L.; LIGA, M.; LI, D. Antimicrobial nanomaterials for water disinfection and microbial control: potential applications and implications. Water Res. 42 (2008) 4591–4602. [6] BROWN, J.; SOBSEY, M.D.; PROUM, S. Use of ceramic water filters in Cambodia. Water Sci. Technol. 50 (2008) 111–115. [7] FENG, Q.L.; WU, J., CHEN, Q.; CUI F.Z.; KIM, T.N.; KIM, J.O. A mechanistic study of the antibacterial effect of silver ions on Escherichia coli and Staphylococcus aureus. J. Biomed Mater. Res. 52 (2000) 662–668. [8] PECHINI, M. P. Method of preparing lead and alkaline-earth titanates and niobates and coating method using the same to form a capacitor-us PAT., 3.330.697, 1967. [9] JIAO, X.; CHEN, D.; XIAO, L. Effects of organic additives on hydrothermal zirconia nanocrystallites, J. Cryst. Growth 258 (2003) 158. [10] RAY, J.C.; PATI, R.K.; PRAMANIK, P. Chemical synthesis and structural characterization of nanocrystalline powders of pure zirconia and yttria stabilized zirconia (YSZ), J. Eur. Ceram. Soc. 20 (2000) 1289. [11] LIANG, J.; JIANG, X.; LIU, G.; DENG, Z.; ZHUANG, J.; LI, F.; LI, Y. Mater. Res. Bull. 38 (2003) 161.
