Environ. Sci. Technol. 2005, 39, 1175-1179
Efficient Visible-Light-Induced Photocatalytic Disinfection on Sulfur-Doped Nanocrystalline Titania J I M M Y C . Y U , * ,† W I N G K E I H O , † J I A G U O Y U , † H O Y I N Y I P , †,‡ PO KEUNG WONG,‡ AND JINCAI ZHAO§ Departments of Chemistry and Biology, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong, China, and Center for Molecular Sciences, Institute of Chemistry, Chinese Academy of Sciences, Beijing, China
Sulfur-doped titanium dioxide exhibits a strong visiblelight-induced antibacterial effect. The new photocatalyst can effectively kill Micrococcus lylae, a common Grampositive bacterium. The relationship between sulfur concentration and the bactericidal activity of S-doped TiO2 was investigated. Results from DMPO spin-trapping electron spin resonance measurements confirm the formation of hydroxyl radicals, which is the origin of the considerable bactericidal activity under visible light irradiation.
Introduction In 1985, Matsunaga and co-workers (1) reported that microbial cells in water could be killed by contact with a TiO2-Pt photocatalyst upon irradiation with UV light for 60120 min. This finding created a new avenue for water sterilization. TiO2-mediated disinfection (2-5) seems to be a promising technique compared to the common disinfection methods such as chlorination and UV disinfection. Chlorination may generate carcinogenic disinfection byproducts, while UV disinfection alone may not be effective for some UV-resistant bacteria. The emergence of more resilient and virulent strains of microorganisms further demonstrates the need for more effective sterilization technologies. Anatase TiO2 absorbs wavelengths in the near-UV region (λ e 390 nm), which is about 3% of the solar spectrum. Due to this inherent limitation, solar energy cannot be utilized efficiently for photocatalytic disinfection. To improve the efficiency, a new approach to broaden the photoresponse of TiO2 by doping with a nonmetal atom has been introduced. Khan et al. have shown efficient photochemical water splitting under visible light by a chemically modified TiO2, in which carbon substitutes for some of the lattice oxygen (6). Kisch et al. have also investigated the daylight photocatalysis by carbon-modified TiO2 (7). Asahi et al. reported theoretical calculations of the band structure of nitrogen-doped TiO2 and its visible light photocatalytic degradation of acetaldehyde and methylene blue (8). They found that nitrogen atoms substituted the lattice oxygen sites and narrowed the band gap by mixing the N2p and O2p states. Reports by Hashimoto et al. demonstrated the visible-light-induced hydrophilicity (9) and photocatalytic decomposition of gaseous 2-propanol * Corresponding author phone: (852) 2609-6268; fax: (852) 26035057; e-mail:
[email protected]. † Department of Chemistry, The Chinese University of Hong Kong. ‡ Deparment of Biology, The Chinese University of Hong Kong. § Chinese Academy of Sciences. 10.1021/es035374h CCC: $30.25 Published on Web 01/19/2005
2005 American Chemical Society
on nitrogen-doped TiO2 (10). They concluded that the lattice oxygen sites were substituted by nitrogen atoms and formed an isolated narrow band above the valence band and narrowed the band gap. Similarly, several others groups have also investigated the photocatalytic and photoelectrochemical properties of nitrogen-doped TiO2 powders and thin films prepared by different methods (11-15). Recently, Umebayashi et al. have succeeded in synthesizing S-doped TiO2, and used it for visible light photocatalytic degradation of methylene blue (16-18). They suggested that sulfur was doped as an anion and replaced the lattice oxygen in TiO2. On the contrary, a report by Ohan et al. found that S atoms were incorporated as cations and replaced Ti ions in the sulfur-doped TiO2 photocatalyst (19-21). Undeniably, doping TiO2 photocatalyst with a nonmetal element becomes a hot research topic, and it opens up new possibilities for the development of solar-induced photocatalytic materials. Previously, our group has reported the doping of fluorine and phosphorus into TiO2 to achieve enhanced photocatalytic activities under UV irradiation (22, 23). We believe that the nonmetal doping is a conceptually simple and promising approach to improve the efficiency of TiO2 disinfection. Although the use of TiO2 in disinfection has been studied extensively, to the best of our knowledge, there has been no report regarding the visible-light-induced bactericidal effects of TiO2 doped with a nonmetal element. The information about this visible-light-induced disinfection is both scientifically and practically important. In the present work, we reveal for the first time that sulfur-doped TiO2 exhibits bactericidal effects on Micrococcus lylae in water under visible light irradiation.
Experimental Section Preparation of Photocatalysts. Sulfur-doped TiO2 was prepared by the following method: A 3 g sample of triblock copolymer HO(CH2CH2O)20(CH2CH(CH3)O)70(CH2CH2O)20H (average molecular weight ca. 4400, designated EO20PO70EO20 or P123) and thiourea (the sulfur source) at different molar ratios were dissolved in 170 mL of absolute ethanol under vigorous stirring. A Ti precursor solution (prepared by mixing 0.02 mol of titanium tetraisopropoxide (Ti(OCH(CH3)2)4, TTIP), 0.01 mol of acetylacetone, and 30 mL of ethanol) was then mixed with the above solution, and the resulting solution was stirred for 2 h. The solution thus obtained was hydrolyzed in an acidic medium (pH 1-2 as controlled by the addition of dilute HCl). After being stirred for 3 h, the sol solution was dried at 100 °C for 24 h to obtain a gel, then thermal-treated in air at a heating rate of 3 °C/ min, and kept at 500 °C for 1 h to remove the P123 triblock copolymer template. The solid was ground to a powder with an agate mortar and denoted as ST. Characterization. X-ray photoelectron spectroscopy (XPS) measurements were performed on a PHI Quantum 2000 XPS system with a monochromatic Al KR source and a charge neutralizer. All the binding energies were referenced to the C1s peak at 284.8 eV of the surface adventitious carbon. The X-ray diffraction (XRD) patterns, obtained on a Bruker D8 Advance X-ray diffractometer using Cu KR radiation at a scan rate of 0.05 deg (2θ) s-1, were used to identify the phase constitutions in samples and their crystallite size. The accelerating voltage and the applied current were 40 kV and 40 mA, respectively. The crystallite size was calculated from X-ray line broadening analysis by the Scherrer formula. UVvis diffuse reflectance spectra were achieved using a UV-vis spectrophotometer (Cary 100 scan spectrophotometers, Varian). Transmission electron microscopy (TEM) images VOL. 39, NO. 4, 2005 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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FIGURE 1. High-resolution XPS spectra of the S2p region for the pure TiO2 and S-doped TiO2 with different atomic percentages of sulfur. were taken with a JEOL 2010 transmission electron microscope operated at a 200 kV accelerating voltage. The Brunauer-Emmett-Teller (BET) surface areas (S(BET)) of the powder samples were determined by nitrogen adsorptiondesorption isotherm measurements at 77 K on a Micromeritics ASAP 2010 nitrogen adsorption apparatus. All the samples measured were degassed at 180 °C before the actual measurements. Electron spin resonance (ESR) spectra were obtained using a Bruker model ESP 300E ESR spectrometer. The settings for the ESR spectrometer were center field 3480.00 G, microwave frequency 9.79 GHz, and power 5.05 mW. Preparation of the Bacterial Culture. M. lylae, a Grampositive bacterium that was isolated in our laboratory, was used as a model bacterium in the experiments. It was incubated in a 10% trypticase soy broth (TSB) at 30 °C with 200 rpm agitation for 24 h. The culture was washed with a 0.9% saline solution by centrifugation at 12000 rpm for 5 min at 25 °C, and the pellet was resuspended in saline. The cell suspension was diluted in a centrifuged tube to the required cell concentration (3 × 107 cfu/mL). Measurements of Bactericidal Activity. The photocatalyst was added to a 0.9% saline solution in a conical flask and homogenized by sonication. The suspension was then sterilized by autoclaving at 120 °C for 20 min and mixed with the prepared cell suspension after cooling. The final photocatalyst concentration was adjusted to 0.2 mg/mL, and the final bacterial cell concentration was 3 × 106 cfu/mL. The visible light was obtained from a 100 W tungsten halogen lamp with a glass filter that cut off light with wavelengths shorter than 420 nm. The intensity of the illumination was 47 mW cm-2 on the catalyst surface during the experiment. Each set of experiments was performed in duplicate. The reaction mixture was stirred (380 rpm) with a magnetic stirrer to prevent settling of the photocatalyst. A bacterial suspension without photocatalyst was irradiated as a control, and the reaction mixture with no visible light irradiation was used as a dark control. Before and during light irradiation, an aliquot of the reaction mixture was immediately diluted with 0.9% saline solution and plated on TSB agar. The colonies were counted after incubation at 37 °C for 48 h.
Results and Discussion X-ray Photoelectron Spectroscopy. In Figure 1, the highresolution XPS spectra of the S2p region show that the S atoms are in the state of S6+ in all S-doped TiO2 samples, with a peak around 170 eV (24). It should be emphasized that the as-prepared powders were washed with deionized water and 1 M HCl aqueous solution several times to remove the surface-adsorbed species such as sulfuric acid. Moreover, in 1176
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FIGURE 2. High-resolution XPS spectra of the S2p region for S-doped TiO2 (a) before and (b) after Ar+ sputtering (0.1 nm/min) for 15 min. Figure 2, the peak at 170 eV was still observed after sputtering with Ar+ ion to a depth of 1.5 nm. These results strongly indicate that the S atoms are incorporated into the bulk phase of TiO2 nanoparticles. In addition, it is important to note that no peaks were found around 160-163 eV, which corresponds to the Ti-S bond when S atoms replaced O atoms on the TiO2 surface. (25) Umebayashi et al. (16-18) used theoretical calculations to depict a narrowing of the band gap when oxygen was replaced by sulfur in anatase TiO2. They found that the sulfur dopant was in the anionic form when TiS2 was used as the starting material. In their preparation, most of the sulfur in TiS2 was oxidized and the residual sulfur would naturally remain as S2-. In our work, however, we used titanium tetraisopropoxide and thiourea as the titanium and sulfur precursors, respectively. Anionic sulfur doping may be difficult to carry out because S2- (1.7 Å) has a significantly larger ionic radius compared to that of O2- (1.22 Å). This leads to the large formation energy required for the substitution forming Ti-S bonding instead of Ti-O bonding. Thus, the substitution of Ti4+ by S6+ is chemically more favorable than replacing O2- with S2-, which is confirmed by our XPS results. Similarly, Ohno et al. (19-21) also obtained cationic sulfur-doped TiO2 and reported the photocatalytic degradation of methylene blue under visible light. It can be concluded that the ionic form of the sulfur dopant is dependent on the preparation routes. It should be mentioned that when the 6+ sulfur ions replace the 4+ titanium ions in the lattice, a charge imbalance is created. The extra positive charge is probably neutralized by the hydroxide ions. Energy-Dispersed X-ray (EDX) Microanalysis, X-ray Diffraction, and Transmission Electron Microscopy. TEM allows us to gain insight into the size, structure, and morphology of the particles. The TEM images of both doped and undoped samples are shown in Figure 3. The spherical particles possess uniform size with average diameters of 11.9 and 8.2 nm for the pure and S-doped TiO2, respectively. Figure 4 displays the EDX microanalysis spectra of the pure and S-doped TiO2. Three areas were selected respectively from both the pure TiO2 (areas A-C) and the S-doped TiO2 (areas D-F) samples in Figure 3 for analysis. Signals corresponding to sulfur were detected in the S-doped TiO2 but not in the pure TiO2. Measurement results from XRD (not shown here) demonstrate that all the S-doped TiO2 is of anatase crystalline phase. In addition, from Table 1, the S-doped TiO2 has a larger BET surface area compared to that of pure TiO2. The crystal sizes of the S-doped samples are smaller than that of
FIGURE 3. TEM images of pure TiO2 and S-doped TiO2 (1.96 atom %). Regions A-F were selected for EDX microanalysis.
FIGURE 4. EDX microanalysis spectra of the pure TiO2 (areas A-C) and S-doped TiO2 (areas D-F). The X-ray excitation energy for S is about 2.3 keV. pure TiO2, which is consistent with the TEM results. These suggest that the incorporation of sulfur inhibits the crystalline growth of the embedded anatase TiO2 during calcination, resulting in a smaller crystal size for the doped materials (23). Diffuse Reflectance UV-Vis Spectroscopy. Figure 5 shows the UV-vis diffuse reflectance spectra of the pure TiO2 and S-doped TiO2 powders. Noticeable shifts of the absorbance shoulder from a wavelength below 400 nm to the visible light region were observed for the S-doped TiO2. Moreover, the
absorbance increases with the atomic percentage of S dopant. Undoubtedly, these results reveal that the sulfur cations are indeed incorporated into the lattice of TiO2, thus altering its crystal and electronic structures. Electron Spin Resonance Spectroscopy. Figure 6 illustrates the ESR spectrum of the DMPO-OH• spin adduct with pulsed laser illumination at λ ) 532 nm. In the dark (0 s), no signal can be detected. Under irradiation, the characteristic 1:2:2:1 quadruple peaks of the DMPO-OH• adduct were observed, and their intensity increased with the VOL. 39, NO. 4, 2005 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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TABLE 1. Crystal Sizes, BET Surface Areas, and Bactericidal Activities of Pure TiO2 and S-Doped TiO2 with Different Atomic Percentages of Sulfur
sample
crystal sizea/ nm
BET surface areab/ m2/g
residual survival of M. lylae after 1 h of visible light irradiationc/%
pure TiO2 ST(1.23 atom %) ST(1.41 atom %) ST(1.66 atom %) ST(1.96 atom %)
11.2 10.3 9.2 9.1 8.5
72.9 100.3 91.9 98.5 113.4
96.5 84.6 54.9 12.1 3.3
a Average crystal sizes of pure TiO 2 and S-doped TiO2 were determined by XRD using the Scherrer equation. b BET surface area calculated from the linear part of the BET plot (P/P0 ) 0.05-0.3). c Survival ratio of M. lylae under visible light irradiation (>420 nm). The experiment was done in duplicate.
FIGURE 7. Survival ratio of M. lylae vs visible light irradiation time (>420 nm) for pure TiO2 powders and S-doped TiO2 (1.96 atom %).
FIGURE 5. UV-vis diffuse reflectance spectra of pure TiO2 and S-doped TiO2 with different sulfur contents.
FIGURE 8. Images of M. lylae colonies on an agar plate before and after visible light irradiation.
FIGURE 6. ESR signals of the DMPO-OH• adduct in sulfur-doped TiO2/DMPO dispersion. The signals were recorded before illumination (0 s) and after illumination for 10, 20, and 40 s at 532 nm with a Quanta-Ray Nd:YAG pulsed laser operated in the continuous mode at 10 Hz frequency. irradiation time. This confirms the formation of hydroxyl radicals from the S-doped TiO2 sample (26, 27). Antibacterial Activity. The bactericidal activities of the samples were evaluated by the killing of (M. lylae in water under visible light irradiation on the basis of the decrease in the colony number of M. lylae formed on an agar plate. According to Figures 7 and 8, M. lylae can be almost completely killed within 1 h on S-doped TiO2 nanoparticles under visible light irradiation. Neither pure TiO2 nor S-doped TiO2 in the dark shows any bactericidal effects on M. lylae, 1178
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FIGURE 9. Residual survival of M. lylae after 1 h of visible light irradiation as a function of sulfur content of the S-doped TiO2 photocatalyst. indicating that the photocatalyst itself is not toxic to M. lylae. Thus, the bactericidal effect is ascribed to the photocatalytic reaction of the S-doped TiO2. In addition, Figure 9 reveals that the bactericidal effect on M. lylae increases with the atomic percentages of sulfur dopant. The results are in good agreement with the visible light absorption of the S-doped TiO2 nanoparticles shown in Figure 5. Moreover, this
relationship can further confirm that the visible light bactericidal effect originated from the sulfur dopant in the photocatalysts. The phenomenon of band-gap narrowing in nonmetal atom doped TiO2 has been reported in the literature (8, 1618). Visible light activity in anionic N- or S-doped TiO2 may be caused by band-gap narrowing from mixing the N2p or S3p states with O2p states, respectively. Hashimoto et al. (9, 10) provided an alternative explanation that a localized N2p state formed above the valence band was the origin for the visible light activity of the nitrogen-doped TiO2. More detailed studies are required to conclude whether our S6+ doping can indeed create intra-band-gap states close to the conduction band edges and thus induces visible light absorption at the sub-band-gap energy. This would be similar to the situation of conventional transition-metal ion doping (28, 29). In addition, our ESR results first confirm the generation of hydroxyl radicals from S-doped TiO2 under visible light irradiation. The photocatalysts are activated at the sub-bandgap energy when exposed to visible light and generate reactive oxygen species such as hydroxyl radicals. These hydroxyl radicals cause various damage to living organisms. Possible mechanisms for the bactericidal effect of TiO2 photocatalysis have been proposed (4, 5, 30-33). In conclusion, this study demonstrates a novel approach for the efficient utilization of visible light in killing bacteria through doping sulfur into TiO2 photocatalyst. The formation of hydroxyl radicals from the S-doped TiO2 under visible light irradiation leads to considerable bactericidal effects on M. lylae. Further study about the formation of new energy levels by the S dopant is currently in progress.
Acknowledgments This work was supported by grants from the Research Grants Council of the Hong Kong Special Administrative Region, China (Project Nos. N_CUHK433/00 and CUHK4325/03M).
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Received for review December 10, 2003. Revised manuscript received September 28, 2004. Accepted November 30, 2004. ES035374H
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