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Converting Visible Light into UVC: Microbial Inactivation by Pr3þ-Activated Upconversion Materials Ezra L. Cates, Min Cho, and Jae-Hong Kim* School of Civil and Environmental Engineering, Georgia Institute of Technology, 200 Bobby Dodd Way, Atlanta, Georgia 30332-0373, United States
bS Supporting Information ABSTRACT: Herein we report the synthesis and properties of light-activated antimicrobial surfaces composed of lanthanide-doped upconversion luminescent nano- and microcrystalline Y2SiO5. Unlike photocatalytic surfaces, which convert light energy into reactive chemical species, this work describes surfaces that inactivate microorganisms through purely optical mechanisms, wherein incident visible light is partially converted into germicidal UVC radiation. Upconversion phosphors utilizing a Pr3þ activator ion were synthesized and their visible-to-ultraviolet conversion capabilities were confirmed via photoluminescence spectroscopy. Polycrystalline films were prepared on glass substrates, and the extent of surface microbial inactivation and biofilm inhibition under visible light excitation were investigated. Results show that, under normal visible fluorescent lamp exposure, a sufficient amount of UVC radiation was emitted to inhibit Pseudomonas aeruginosa biofilm formation and to inactivate Bacillus subtilis spores on the dry surfaces. This new application of upconversion luminescence shows for the first time its ability to deter microbial contamination and could potentially lead to new material strategies for disinfection of surfaces and water.
’ INTRODUCTION Minimizing the occurrence and transmission of pathogenic microorganisms in water, food, and the indoor environment has been of utmost priority to human beings since the earliest civilizations. Currently, ever-evolving microbial resistance and the need for more sustainable disease prevention techniques make innovative disinfection technologies vital to the future of both developed and developing countries. Many works over the past recent decades have discussed the benefits of photocatalytic processes, for both water and surface disinfection, wherein catalysts such as TiO2 use light energy to produce redox potential and germ-killing reactive oxygen species.14 Utilization of light energy in particular, either ultraviolet or visible, makes such technologies a more sustainable approach than the use of chemical disinfectants or other energy-intensive means. We herein introduce for the first time in the literature an innovative approach to utilizing electromagnetic energy for the purpose of disinfection, the mechanism of which is fundamentally different from that of photocatalysis. Our strategy is based on the purely optical phenomenon of directly converting visible light into germicidal UVC radiation via the photoluminescence process of upconversion (UC, Figure 1). This anti-Stokes process involves the sequential absorption of two or more photons by a material to reach an excited state that emits one higher-energy photon. UC almost exclusively employs lanthanide ions (Ln3þ) as luminescent activators due to the shielding of 4f electrons from crystal field r 2011 American Chemical Society
effects and vibronic coupling by the outer 5s2 and 5p6 subshells that results in millisecond range excited-state lifetimes.5,6 Furthermore, it is unique among nonlinear optical processes in that it does not require coherent, high-energy electromagnetic fields to occur significantly and can in fact employ nonlaser excitation,7 though laser-based studies abound in the literature.5 The vast majority of these works have progressed within the context of infrared-to-visible conversion due to biomedical and bioimaging applications of the highly efficient Yb3þ-sensitized phosphors.6,810 Additionally, conversion of IR sub-band-gap photons into visible wavelengths for improving solar cell efficiency is a unique application that has inspired many recent studies of UC under low-power excitation (∼1000 W/m2), and several phosphor systems show significant external optical efficiencies (for example, 3.0% at 880 W/m2 excitation)11 despite the quadratic dependence on excitation intensity.1113 This ability to amplify ordinary diffuse IR radiation allows for many interesting fields of research; however, little attention has been devoted to upconversion of abundant visible light energy into UV radiation.
Received: January 17, 2011 Accepted: March 8, 2011 Revised: March 7, 2011 Published: March 23, 2011 3680
dx.doi.org/10.1021/es200196c | Environ. Sci. Technol. 2011, 45, 3680–3686
Environmental Science & Technology
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Figure 1. Utilization of visible-to-ultraviolet upconversion phosphor coating for light-activated antimicrobial materials. Energy diagram depicts excited-state absorption of visible light from the ground-state configuration, G, to the excited states, E1 and E2, to emit a UVC photon upon relaxation.
In the present work, we achieved visible-to-ultraviolet UC (specifically to UVC wavelengths) through the use of praseodymium as a UC activator. Past spectroscopic studies on Y2SiO5:Pr3þ commercial laser crystals have revealed the UC capabilities of Pr3þ in the yttrium oxyorthosilicate host, which utilizes a metastable 4f intermediate state to populate the 4f5d band through absorption of blue photons, as shown in the lefthand side of Figure 2B14,15 (a detailed excitation spectrum is shown in Hu et al.14). Emission from this band produces a broad spectrum of UV radiation, including in the UVC range (220280 nm), which is well-known to be effectively absorbed by microbial DNA and RNA, causing covalent linkages between nucleic acid components to produce a variety of photoproducts that inhibit transcription.16 This mutagenic action peaks at approximately 270 nm, depending on the organism.17 Using this system as a foundation, we prepared nanocrystalline phosphors and studied the antimicrobial properties of surfaces coated with the UC materials. Additional codopants, Liþ and Gd3þ, were also employed to maximize UC emissions through tailoring the crystal structure and to improve the biocidal effect. Since the optical efficiencies of UC phosphors are generally much lower than those of conventional phosphors (∼1% for the most efficient infrared-to-visible UC materials under laser excitation),18 we chose antimicrobial surfaces as a suitable application to begin with and to prove the concept of light upconversion for disinfection applications, since the target organisms would reside directly on the material surface and thus receive maximum exposure to emitted UV photons. In general, materials that possess antimicrobial properties find application in numerous environmental and public health technologies for prevention of hospital-acquired infections, food contamination, and other pathogen transmissions through inanimate surfaces.19,20 Presently, visible-to-UV conversion is not a welldeveloped field; however, we hypothesized that the aforementioned highly mutagenic effect of germicidal UVC would result in significant microbial inactivation by such surfaces (while maintaining insignificant human exposure), even under the
Figure 2. (A) Upconversion emission spectra of Y2SiO5:Pr3þ phosphor powders with various codopants under 488 nm laser excitation. Inset (*) shows zoomed-out view of Y2SiO5:Pr3þ,Gd3þ,Liþ emission to show height of UVB peak. (B) Upconversion mechanisms of Pr3þ and Gd3þ UV emissions. Solid blue line shows visible light absorption; dotted black line shows nonradiative energy transfer; dotted purple line shows UV photon emission.
relatively low fluxes expected from upconversion of visible light by the Ln3þ-doped systems developed herein.
’ EXPERIMENTAL SECTION Materials. All chemicals were purchased from Sigma Aldrich. Yttrium nitrate was made from Y2O3 (99.999%) by boiling in 1:1 nitric acid (trace metal grade) and evaporating to dryness in an oven at 104 °C. Any dopants were also added at the same time as HNO3, with appropriate amounts of 0.2 M aqueous Ln3þ dopant solutions and 1 M LiNO3 (99.999%) while a stoichiometric amount of Y2O3 was omitted. For Liþ doping, Y3þ was omitted at 1/3 the molar amount of Liþ. Aqueous stock solutions of Pr3þ and Gd3þ (0.2 M) were prepared from Pr(NO3)3 3 6H2O (99.99%) and Gd(NO3)3 3 6H2O (99.99%). Tetraethoxysilane (TEOS, 99.999%) was the silicon source and gelling agent. All water was purified on a Millipore ultrapurification system. Powder Phosphor Synthesis. Optical properties of the UC phosphors were studied mainly by use of nano- or microcrystalline powder materials prepared via sol gel decomposition. Sol gels were made by converting 5.3 g of Y2O3 to the anhydrous nitrate form (with dopants) and then dissolving in 17.25 mL of ethanol (200 proof) and 5.4 mL of water. 3681
dx.doi.org/10.1021/es200196c |Environ. Sci. Technol. 2011, 45, 3680–3686
Environmental Science & Technology A stoichiometric amount of TEOS was then added, followed by an additional 10 min of stirring. The solutions in beakers were heated at ∼70 °C until a clear gel formed. The gels were placed in an oven at 104 °C for 17 h to form a tacky xerogel, which was ground to a powder with mortar and pestle and placed in alumina crucibles. The samples were heated to 1000 °C at a ramp of 8 °C/min in a muffle furnace with air atmosphere and held at 1000 °C for 3 h, then cooled naturally down to room temperature. Dopant concentrations were adjusted to optimize upconversion efficiency, which corresponded to 1.0 mol % Pr3þ and Gd3þ for samples without lithium and 1.2 mol % Pr3þ and Gd3þ for samples with Liþ. Upconversion emission was optimized at a Liþ concentration of 7.2 mol %. The resulting powders appeared off-white for samples without Liþ, while the samples with Liþ were bright white with a light greenish tint attributed to the Pr3þ. Coating Procedure. To coat substrates for antimicrobial testing, 12.7 mm 12.7 mm (0.25 in2) silica glass squares were first roughened with 150-grit sandpaper and washed clean. They were then dipped manually into a sol solution, prepared in the same manner as the sol for the powder phosphors except that it was stirred gently overnight and then aged for an additional day. One side of each square was wiped clean and the film on the other side was allowed to dry for 1 h at room temperature and then 1 h in an oven at 104 °C. They were then transferred to a furnace and heated in the same manner as the powder phosphor. Characterization. Stokes emission spectra were obtained by preparing 0.3% (w/w) phosphor powder suspensions in ethanol via sonication and analyzed on a Shimadzu spectrofluorophotometer equipped with xenon arc lamp. Upconversion emission spectra were obtained with an argon laser (Stellar Pro-L-ML, Modulaser Inc.) with 488 nm line filter as the excitation source to produce a 140 mW beam. The beam was focused onto the sample at an approximate 45° angle, which was in the form of finely ground powder compressed into a small hexagonal mold mounted parallel to the table. The emission was collected normal to the particle surface with UV antireflective focusing lenses and directed through an optical chopper and a short-pass filter (>400 nm cutoff) to remove laser wavelengths and then into a 0.25 m monochromator (Oriel Cornerstone, Newport Corp.). The signal was detected by an Oriel photomultiplier tube, sensitive from 180 to 700 nm, and processed by an Oriel Merlin radiometry system (Newport Corp.) and personal computer. Phase-sensitive detection was employed with the Merlin system and the optical chopper, placed between the sample and the monochromator inlet and operating at 120 Hz. Unactivated Y2SiO5:Liþ powder was used to obtain a background spectrum, which was subtracted from all sample spectra. X-ray diffraction analysis was performed on the as-made powder samples in a Scintag XGEN-4000 diffractometer using Cu KR radiation. Scanning electron microscopy (SEM) of lithium-doped powders was performed with a JEOL JSM-6490LV SEM at 1520 kV after sputter coating the samples with gold for 30 s. For imaging of non-lithium-doped powders and surface coatings, we used a JEOL JSM-6500F Field Emission SEM at an accelerating voltage of 5 kV. Biofilm Growth and Imaging. Pseudomonas aeruginosa PAO1 was grown in tryptic soy broth (TSB) at 37 °C. Cells were harvested by centrifugation at 1000 g for 10 min and washed twice with phosphate-buffered saline (PBS; pH 7.0). A PAO1 suspension was prepared by resuspending the cell pellet in
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50 mL of PBS. Biofilms were grown in a Centers for Disease Control and Prevention (CDC) reactor (Biosurface Technologies Inc., Bozeman, MT). The CDC reactor contained four rods that each held three glass coupons. Phosphor-coated and control glass samples were affixed to two of these rods, on opposing sides. An overnight culture was prepared by incubating PAO1 in 1/10 diluted TSB for 20 h at 37 °C. The sterile reactor was inoculated with 3.5 mL of an overnight culture that had been added to 350 mL of 1/100 diluted TSB. The initial PAO1 population in this batch medium was about 106 colony-forming units (cfu)/mL. Two “cool white” 13 W compact fluorescent bulbs were placed on opposite sides of the reactor, in line with the outward-facing samples, and UV filters were placed in between (cutoff
