Electrochemical Carbon-Nanotube Filter Performance toward Virus

Dec 23, 2011 - The MWNT-filter surface morphology was examined using a high-resolution field emission scanning electron microscope (FE-SEM) (Hitachi S...
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Electrochemical Carbon-Nanotube Filter Performance toward Virus Removal and Inactivation in the Presence of Natural Organic Matter Md. Saifur Rahaman,† Chad D. Vecitis,‡ and Menachem Elimelech*,† †

Department of Chemical and Environmental Engineering, Yale University, New Haven, Connecticut 06520-8286, United States School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts 02138, United States

‡

S Supporting Information *

ABSTRACT: The performance of an electrochemical multiwalled carbon nanotube (EC-MWNT) filter toward virus removal and inactivation in the presence of natural organic matter was systematically evaluated over a wide range of solution chemistries. Viral removal and inactivation were markedly enhanced by applying DC voltage in the presence of alginate and Suwannee River natural organic matter (SRNOM). Application of 2 or 3 V resulted in complete (5.8 to 7.4 log) removal and significant inactivation of MS2 viral particles in the presence of 5 mg L−1 of SRNOM or 1 mg L−1 of alginate. The EC-MWNT filter consistently maintained high performance over a wide range of solution pH and ionic strengths. The underlying mechanisms of enhanced viral removal and inactivation were further elucidated through EC-MWNT filtration experiments using carboxyl latex nanoparticles. We conclude that enhanced virus removal is attributed to the increased viral particle transport due to the applied external electric field and the attractive electrostatic interactions between the viral particles and the anodic MWNTs. The adsorbed viral particles on the MWNT surface are then inactivated through direct surface oxidation. Minimal fouling of the EC-MWNT filter was observed, even after 4-h filter runs with solutions containing 10 mg L−1 of natural organic matter and 1 mM CaCl2. Our results suggest that the EC-MWNT filter has a potential for use as a high performance point-of-use device for the removal of viruses from natural and contaminated waters with minimal power requirements.

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INTRODUCTION Access to safe drinking water is fundamental to maintaining the basic standards of public health. According to a World Bank estimation, nearly 1.1 billion individuals lack access to safe water,1,2 which results in millions of deaths annually by waterborne diseases in developing countries.3 Only a few lowcost point-of-use technologies are available that provide high removal levels of a wide range of microbial pathogens and particularly of viruses. Therefore, there is a critical need to develop technologies that provide high treatment levels, while remaining affordable and convenient to use.4 Carbon nanotubes (CNTs) have been considered for environmental remediation due to their unique properties, such as high surface area, improved mechanical strength, and high thermal and electrical conductivity.5−12 For example, CNTs were found to be effective in the removal of a range of organic contaminants.9,11,13,14 Recently, multiwalled and singlewalled carbon nanotube (MWNT and SWNT) filters have been developed and found to be effective for complete bacterial removal by physical straining and for multilog viral removal through depth filtration.15−17 Although both the MWNT and SWNT filters exhibited complete bacterial removal, the MWNT filter was found to be more effective at removing viruses from relatively pure solutions than the SWNT filter.17 However, the MWNT-filter performance was greatly compro© 2011 American Chemical Society

mised in the presence of natural organic matter (NOM) and other organic macromolecules such as alginate.17 Electrochemical inactivation of waterborne pathogens has long been studied as an alternative to conventional water disinfection.18−23 However, most previous studies have focused on indirect oxidation of pathogens by electrochemically generated active chlorine species and/or other oxidants.24 The major drawback of indirect electrochemical oxidation is that the generated chlorine species react with natural organic matter, already present in the water, and produce harmful disinfection byproducts.20 Promoting direct oxidation is advantageous as the pollutants are oxidized at the anode surface, therefore avoiding production of any harmful byproducts.25 Due to their high electrical conductivity, application of CNTs as an electrode material is an active field of research for fuel cells and batteries.26−29 In an analogous exploitation of their unique electronic properties for water treatment, CNTs were employed in the enhanced electrochemical inactivation of removed pathogens.24,30 In our recent study, we demonstrated Received: Revised: Accepted: Published: 1556

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using a ZetaPALS analyzer (Brookhaven Instruments Corp., Holtsville, NY). MS2 Bacteriophage Preparation and Characterization. MS2 bacteriophage (ATCC 15597-B1), along with their bacterial host Escherichia coli 15597, were purchased from the American Tissue Culture Collection (ATCC). The stock viral solution for the filtration experiments was prepared by suspending the freeze-dried phage pellet in 10 mL of DI water, which results in a viral concentration of 1010 PFU mL−1. Stock solution was further diluted in order to prepare the desired solutions for viral filtration experiments (106 to 107 PFU mL−1). For EPM measurements of the viral particles, where a much higher virus concentration is needed, MS2 purification was performed according to a previously published method33 with minor modifications. Specifically, a concentrated phage suspension (1 mL) was added to the soft agar-host E. coli solution, and the mixture was poured directly over an agar plate. After 24 h of incubation at 37 °C, 5 mL of NaHCO3 solution (1 mM) was poured into the plate and set aside for 5 min. The solution was then poured off into a plastic tube and vortexed for 20 s. Subsequently, the mixture was centrifuged at 5,000 rpm for 15 min. The supernatant was then separated and centrifuged again at 5,000 rpm for 1 min. Finally, the supernatant was filtered through a 0.22-μm PES syringe filter (Millex-GP, SLGP033RS, Millipore), and the filtrate was analyzed for viral concentration. Using this purification protocol, a highly concentrated stock viral solution (1011 PFU mL−1) was obtained. This stock solution was used for both viral EPM and size (hydrodynamic diameter by dynamic light scattering) measurements using a ZetaPALS analyzer (Brookhaven Instruments Corp., Holtsville, NY). Electrolyte and Organic Matter Solutions. Separate electrolyte stock solutions of monovalent and divalent salts were prepared by dissolving reagent grade NaCl and CaCl2•2H2O (J.T. Baker, NJ) salts in ultrapure DI water. The desired solutions for all runs were prepared by diluting and/or mixing the stock solutions at an ambient pH of 5.5. The solution pH was adjusted by adding 0.1 M NaOH or 0.1 M HCl. Prior to each viral experiment, the electrolyte solution was autoclaved in order to ensure its sterility. Suwannee River natural organic matter (SRNOM) and Suwannee River humic acid (SRHA) (International Humic Substances Society, St. Paul, MN) stock solutions were prepared by dissolving 100 mg of the respective dry powder in 200 mL of DI water and stirring the solution overnight in the dark. The pH of the stock solutions was raised to 8.5 by adding 0.1 mM NaOH. The stock solutions were then vacuum filtered through a 0.22-μm cellulose acetate membrane (Corning Inc., Corning, NY) and were subsequently stored in the dark at 4 °C. The total organic carbon contents of both the SRNOM and SRHA stock solutions were determined using a TOC analyzer (TOC-VCSH, Shimadzu, Kyoto, Japan). SRNOM and SRHA have been used extensively as model natural organic matter, and their physicochemical characteristics are well established.34,35 Alginate stock solution (0.5 g/L) was prepared by dissolving alginic acid sodium salt (Sigma-Aldrich, St. Louis, MO) in DI water and filtering the solution through a 0.45-μm cellulose acetate membrane (Corning Inc., Corning, NY). The alginate stock solution was stored at 4 °C until use. For all viral experiments, solutions with the desired concentrations of MS2 and organic matter were obtained by adding appropriate amounts of MS2 and organic matter stock solutions to the electrolyte solutions. The solution pH was adjusted just before

a novel electrochemical depth-filtration process where direct oxidation is promoted by filtering the microbial pathogens through an anodic CNT filter medium.24 The electrochemical CNT filter demonstrated complete removal and significant inactivation of both viruses and bacteria by applying 2 or 3 V DC potential. However, this pathogen filtration study was conducted using a relatively pure aquatic matrix of 10 mM NaCl, which is not applicable to natural waters that ubiquitously contain dissolved organic matter. Therefore, it is of paramount importance to investigate whether NOM impacts the performance of the electrochemical CNT filter under more relevant environmental conditions. In this study, we investigate the removal and inactivation of viruses by an electrochemical multiwalled carbon nanotube (EC-MWNT) filter in the presence of natural organic matter and alginate over a broad range of solution chemistries. A marked increase in viral removal is observed upon application of DC voltage, and the underlying mechanism of the enhanced viral removal is elucidated through more controlled filtration experiments using carboxyl latex nanoparticles as a viral surrogate. The organic fouling propensity of the EC-MWNT filter was also evaluated by performing extended filtration experiments with natural organic matter.

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MATERIALS AND METHODS MWNT Filter Media Preparation and Characterization. Multiwalled carbon nanotubes (MWNTs) with >95% purity by weight were purchased from NanoTechLabs Inc. (Yadkinville, NC). According to the manufacturer, the MWNTs have a bulk density of 2.1 g/cm3 and an average diameter and length of 15 nm and 100 μm, respectively. A detailed characterization of the MWNTs was performed in our previous studies.24,31 MWNTs (as received) were suspended in dimethyl sulfoxide (DMSO) at a concentration of 0.5 mg/mL. The MWNT suspension was then probe sonicated (Branson, Sonifier 450) for 15 min, followed by cooling for another 15 min to room temperature. The MWNT filter was prepared by depositing 3 mg of MWNTs on a 5-μm PTFE membrane (Millipore, Omnipore, JMWP) through vacuum filtration of 6 mL MWNT suspension, resulting in a MWNT loading of 0.32 mg/cm2. The filter was then washed by filtering through it 50 mL of ethanol, a 50-mL solution of equal parts ethanol and deionized (DI) water, and 300 mL of DI water to remove any residual DMSO before use. The MWNT-filter surface morphology was examined using a high-resolution field emission scanning electron microscope (FE-SEM) (Hitachi S-4500, Hitachi, USA). MWNT stock solution for electrophoretic mobility (EPM) measurements was prepared using a successive sonicationclarification protocol.32 MWNTs (as received) were suspended in DI water at a concentration of 0.5 g/L. The suspension was then sonicated continuously using an ultrasonication probe (Misonix 3000, Misonix Inc., Farmingdale, NY) for 30 min, and the stable supernatant was separated after cooling the suspension at room temperature for 15 min. In a similar fashion, 5 more cycles of sonication-clarification were performed, and the resulting mixture was used as a stock MWNT suspension for EPM measurements. The stock MWNT suspension was diluted approximately 20-fold for the EPM measurements. Prefiltered salt solutions, natural organic matter, and acids or bases were added as necessary to obtain the desired solution chemistry prior to EPM measurements 1557

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were suspended in 10 mM NaCl solution at pH 9, and 10 mL of the suspension was flowed through the MWNT filter for each individual run. Filtration experiments were carried out at different approach velocities and different applied potentials (DC voltage). Effluent samples were collected in glass tubes, and influent and effluent samples were analyzed for total organic carbon content using a TOC analyzer (TOC-VCSH, Shimadzu, Kyoto, Japan). Assessing Organic Fouling of Electrochemical MWNT Filter. The fouling characteristics of the MWNT filter were studied using two different model organic foulants: SRNOM and SRHA. Figure S2 presents a schematic of the experimental setup for examining organic fouling of the electrochemical MWNT filter. The MWNT filter was first equilibrated by pumping a background electrolyte solution (7 mM NaCl and 1 mM CaCl2) through it for 30 min until a constant flow rate of 4 mL min−1 (280 L m−2 h−1) was attained. An electrolyte solution (pH 5.5) containing 10 mg TOC L−1 of NOM was then pumped at a constant permeate flux of 280 L m−2 h−1. The inlet pressure of the filter was monitored by a differential pressure transducer between the pump and the filter. The measured pressure drop was recorded continuously by a computer. The total mass of permeate was also monitored continuously by an analytical balance in order to determine the permeate water flux. All fouling experiments were conducted at room temperature (23 °C).

the experiments by adding 0.1 M NaOH or HCl, as necessary. Alginate has been used extensively as a model polysaccharide and hydrophilic organic matter, and its physicochemical characteristics are well established.35 Electrochemical Filtration and Quantification of Virus Removal and Inactivation. The electrochemical MWNT filter device (Figure S1) is a slightly modified version of the original design described in our previous study.24 Briefly, a thin stainless steel sheet is perforated to allow for flow through and is used as the cathode. Beneath it is a silicone rubber insulating ring and a titanium ring in contact with MWNT filter that acts as the anode. In order to prevent any leakage of the viral solution from the edge of the MWNT filter, a high-purity silicone rubber foam film gasket (0.254 mm thick; 33 mm inner diameter) is placed on the plastic base of the filtration cell, and the MWNT filter is placed on top of it. The cell was attached to a DC power supply (Agilent E3646A, Santa Clara, CA) that displayed the current. For viral experiments, MS2 bacteriophage solutions were prepared by adding stock MS2 phage to an electrolyte solution with the desired solution chemistry (pH, ionic strength, and organic carbon content). MS2 viral solution (10 mL) was then filtered using a peristaltic pump at a constant permeate flux of 140 L m−2 h−1. Filter effluent samples were collected in autoclaved glass tubes, and the virus concentration was determined by the plaque forming unit (PFU) method (EPA Method 1601). To measure the viruses desorbed from the MWNT filter, the filter was removed from the cell after having been challenged with virus influent and rinsed with 10 mL of the appropriate electrolyte solution. The filter was then placed in 10 mL of DI water in an autoclaved glass tube and bath sonicated for 2 min to remove and suspend the MWNTs from the PTFE membrane. The resulting suspension, containing MWNTs and viruses, was then analyzed for virus concentration by the PFU method. At least two experiments were performed for each set of experimental conditions. Transmission Electron Microscopy (TEM). Viral solutions (10 mL) containing 109 PFU mL−1 in 10 mM NaCl (pH 3.0) were filtered through MWNT filters both in the presence (2 V) and absence (0 V) of DC voltage. Then, the MWNT filter was removed from the cell and rinsed with 10 mL of electrolyte solution. Next, the filter was placed in 10 mL of 1 mM NaHCO3 solution in an autoclaved glass tube and was bath sonicated for 2 min to suspend the MWNTs from the PTFE membrane. The suspension was then stored at 4 °C until TEM analysis was performed. Prior to TEM analyses, the samples were diluted (100-fold) in order to capture clear images of viral particles adsorbed onto the MWNTs. TEM analyses of the samples were conducted at CAMCOR (University of Oregon, Eugene, OR). The diluted samples (5 μL) were applied to freshly cleaned, holey-carbon-coated copper grids, washed with DI water, and negatively stained with 2% uranyl acetate. The grids were then dried in air and imaged using a FEI Tecnai TEM (FEI Tecnai G2 Spirit TEM STEM). Latex Particle Removal Tests. In order to elucidate the underlying mechanism of enhanced viral removal during application of DC voltage, a viral surrogate (i.e., carboxyl latex particles, 40 nm in diameter, Invitrogen) was used to challenge the electrochemical filter. Due to difficulties in determining low concentration of latex particles in the effluent, a very high influent concentration of 7.6 × 1012 particles per mL (410 mg L−1) was used for all experiments. Latex particles

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RESULTS AND DISCUSSION Properties of MS2 Viral Particles and MWNT Filter Media. The physicochemical properties of both the filter media and the viral particles play a significant role in the overall filter performance. The electrokinetic characteristics of both MWNTs and MS2 viral particles in the presence of SRHA under viral filtration solution chemistries and the hydrodynamic diameter of the viral particles are presented in Table S1. Both the MWNTs and the viral particles are negatively charged over the range of solution chemistries examined. The viral particles were well-dispersed and largely unaggregated, except in the case of electrolyte solutions containing divalent Ca2+ ions. Overall, the results suggest that in the absence of applied voltage, the viral particles will experience repulsive interactions with the MWNT filter media. As expected, the EPM of MWNTs became more negative with increasing pH and less negative with increasing ionic strength (Table S1). Overall, the EPMs of MWNTs in the presence of NOM were slightly more negative than the reported literature values in the absence of NOM,17,32 consistent with previously reported observations.32 With increasing pH, both MWNTs and the adsorbed NOM became more negative through dissociation of carboxyl functional groups, which explains the substantial increase in the magnitude of EPM, from −1.82 × 10−8 to −3.59 × 10−8 m2 V−1 s−1 over the pH range from 3 to 9. Increasing the monovalent salt (NaCl) concentration (from 1 mM to 100 mM) made the EMP values much less negative through compression of the diffuse double layer and reduction of Stern potential. Likewise, addition of divalent cations (Ca2+) caused a slight reduction in EPM through adsorption and charge neutralization. FE-SEM images of the MWNT filter, prepared by depositing DMSO-suspended MWNTs on a 5-μm PTFE membrane, are presented in Figure S3. As shown in the SEM images, MWNT deposition on the PTFE membrane was uniform, resulting in a highly porous filter matrix. Permeability tests, performed in our 1558

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Figure 1. Comparison of the performance of MWNT filter with and without application of a DC voltage (0, 2, and 3 V). (A) Log MS2 bacteriophage removal in the presence of alginate and NOM. Influent solutions containing either 1 mg L−1 alginate or 5 mg L−1 SRNOM (as TOC) in 10 mM NaCl solution (pH 5.5) were spiked with 106−107 PFU/mL of MS2 phages. In each experiment, 10 mL of MS2 viral solution was filtered through the MWNT filter at a constant permeate flux 140 L m−2 h−1. Each data point represents the mean of at least two measurements with error bars representing standard deviations. At both 2 and 3 V, no culturable viruses were detected in the effluent (100% removal; hence, no error bar). (B) Number of culturable MS2 phages desorbed from the MWNT filter. After each viral filtration experiment, the MWNT filter was removed from the holder, placed in 10 mM NaCl, and subsequently bath sonicated for 2 min. The solution containing MS2 and MWNT suspension was analyzed for culturable MS2 phages through PFU assay. Error bars represent one standard deviation.

Figure 2. Effect of solution pH on MS2 bacteriophage removal and inactivation with and without application of a 2 V DC voltage. (A) Log MS2 bacteriophage removal in the presence of 5 mg L−1 SRHA (as TOC). Influent solution (10 mM NaCl) contained 106−107 PFU/mL of MS2 phage and 5 mg L−1 of SRHA (as TOC). Feed solutions at the desired pH were applied to the MWNT filter at a constant permeate flux of 140 L m−2 h−1. Duplicate runs were performed for each experimental condition, and the average values are presented along with error bars that represent one standard deviation. At pH 3 to 7, no culturable viruses were detected in the effluent (100% removal; hence, no error bar) in the presence of 2 V DC voltage. (B) Number of culturable MS2 phages desorbed from the MWNT filter. The sample preparation for the PFU assay was the same as described in Figure 1. Error bars represent one standard deviation.

earlier study,17 demonstrated that the uniformly deposited MWNT filter matrix generated high water fluxes at relatively low pressures, achieving an average permeability of 11900 ± 440 L m−2 h−1 bar−1 over the water flux range of 570 to 1640 L m−2 h−1. The EPM of the viral particles in the presence of NOM became more negative with increasing pH and less negative with increasing monovalent salt (NaCl) concentration and addition of calcium ions. Overall, the EPMs of the viral particles in the presence of NOM were less negative than those in the reported data in the absence of NOM17 for the pH range tested, except at pH 3, which may be due to the displacement of the electrokinetic plane of shear by adsorbed NOM molecules on the viral capsid.36 At pH 3, which is lower than the reported isoelectric point (pI) of the MS2 phage (3.1−

3.9),37,38 the viral particles showed slightly negative EPM due to adsorbed humic macromolecules. The viral particles were mostly unaggregated with an apparent hydrodynamic diameter ranging from 25 to 38 nm (Table S1). The measured MS2 viral particle range is in close agreement with the reported range of 24 to 26 nm.39 For solutions containing Ca2+ ions, the viral particles slightly aggregated as indicated by the measured mean diameter of 53 nm. The presence of humic acid in solution likely prevented the aggregation of the viral particles due to steric stabilization imparted by the adsorbed NOM molecules. Applied Voltage Markedly Enhances MWNT-Filter Performance. Remarkable improvement in viral removal was achieved by application of a DC potential in the presence of both alginate and SRNOM (Figure 1A). For example, log viral 1559

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Figure 3. Effect of solution ionic strength on MS2 bacteriophage removal and inactivation with and without application of 2 V DC voltage. MS2 viral solutions with three different ionic strengths (1, 10, and 100 mM NaCl) were used at pH 7. In each case, the solution contained 106−107 PFU/mL of MS2 phage and 5 mg L−1 SRHA (as TOC). MS2 viral solution (10 mL) was filtered at a constant permeate flux of 140 L m−2 h−1. (A) MS2 log removal as a function of solution ionic strength. (B) Number of culturable MS2 phages desorbed from the MWNT filter. Error bars represent one standard deviation, calculated from the results of duplicate runs at the specific experimental conditions.

consistent with our previous study on viral removal by a MWNT filter in the absence of NOM.17 Steric repulsion and increased electrostatic repulsion with increasing pH reduced the extent of viral particle deposition onto the MWNTs, which resulted in lower log removal. In the presence of DC potential, the effect of pH on log viral removal was relatively small, except at pH 9. The DC potential also resulted in a significant improvement in the MWNT-filter performance at all pHs (Figure 2A). No culturable viruses were detected in the effluent for the pH range used, except at pH 9, which exhibited 3.65 log removal (Table S2). This high level of MS2 viral removal over a wide range of pH values and in the presence of NOM suggests that the EC-MWNT filter can be effectively used for real-world environmental applications. A significant increase in viral inactivation was also achieved by applying DC potential over the pH range studied (Figure 2B and Table S2). For instance, at pH 7 and 2 V, only 2,900 PFU mL−1 of culturable viruses were desorbed from the filter, compared to 171,500 PFU mL−1 in the absence of DC potential. The incomplete inactivation of the viruses in the presence of 2 V DC potential may be due to the shielding effect of the NOM coating on the viral particles and/or MWNTs protecting them from oxidation. In agreement with the proposed NOM oxidative shielding effect, the number of culturable viruses desorbed from the electrolyzed filter increased with decreasing pH, due to increased NOM adsorption at lower pH values.40 (b). Impact of Solution Ionic Strength. In the absence of DC potential, the presence of SRHA adversely affected the MWNT filter performance over the range of ionic strengths investigated (1 to 100 mM NaCl) (Figure 3A). The MWNT filter achieved a maximum log viral removal of 2.2 at an ionic strength of 100 mM, and the removal decreased with decreasing ionic strength, consistent with our previous study in the absence of NOM.17 However, with the application of a 2 V DC potential, the MWNT filter performance increased remarkably, attaining complete removal of all influent viruses at all three ionic strengths. The results suggest that ionic strength does not impact viral removal by the EC-MWNT filter at voltages lower than 2.3 V, where the one-electron oxidation of chloride to reactive chlorine radicals does not take place.19

removals by the MWNT filter were only 2.37 and 1.52 in the presence of 5 mg L−1 of SRNOM and 1 mg L−1 of alginate, respectively. However, under identical experimental conditions, no culturable viruses were detected in the effluent (i.e., complete removal) when a 2 (1 mA) or 3 V DC (10 mA) potential was applied. This observation is remarkable considering the ubiquity of NOM and biomacromolecules in natural and wastewater-impacted waters. Proposed mechanisms for the marked enhancement in virus removal in the presence of applied voltage are discussed later in the paper. The extent of electrochemical inactivation of adsorbed viruses was also evaluated by determining the number of culturable viruses desorbed from the MWNT filter (Figure 1B). A significant number of culturable viruses, 362,500 and 94,250 PFU mL−1, were desorbed from the MWNT filter when the influent viral solution contained 5 mg L−1 of SRNOM and 1 mg L−1 of alginate, respectively. However, with application of 2 V DC voltage, the number of desorbed and culturable viruses was reduced significantly to 792 and 40 PFU mL−1 for solutions containing SRNOM and alginate, respectively. Furthermore, no desorbed and culturable viruses from the MWNT filter were observed when a 3 V DC potential was applied during viral filtration. The high levels of inactivation of adsorbed viruses by applied DC voltage is likely attributed to the direct oxidization of the adsorbed viral capsid by the anodic MWNT filter as shown in our previous study for bacteria adsorbed to MWNTs. Viral Removal and Inactivation in the Presence of NOM under Different Solution Chemistries. We have shown earlier that changes in solution chemistry (pH, ionic strength, and Ca2+ ions) significantly impact the electrokinetic properties of the MWNTs and the MS2 viral particles, which in turn affects the interaction between the viral particles and the MWNTs. Here, we study how these factors influence viral removal and inactivation by the MWNT filter with and without applied voltage. (a). Impact of Solution pH. In the absence of DC potential, the MWNT-filter performance was greatly compromised by the presence of SRHA over the pH range tested (Figure 2A). As anticipated by the MWNT and virus electrokinetic properties, viral removal decreased with increasing solution pH. For example, log viral removal decreased from 2.22 to 1.26 as the solution pH rose from 3 to 9. This trend is 1560

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Figure 4. Effect of divalent cations (Ca2+) on MS2 virus removal and inactivation with and without application of a 2 V DC voltage. MS2 viral solutions (pH 7) having different solution chemistries were used (1 mM CaCl2 + 7 mM NaCl or 10 mM NaCl). In each case, the solution contained 106−107 PFU/mL of MS2 phage and 5 mg L−1 of SRHA (as TOC). MS2 viral solution (10 mL) was filtered at a constant permeate flux of 140 L m−2 h−1. (A) MS2 log removal at different solution chemistries. (B) Number of culturable MS2 phages desorbed from the MWNT filter. Error bars represent one standard deviation calculated from the results of duplicate runs at the specific experimental conditions.

Figure 5. (A) Removal of carboxyl latex particles (40 nm in diameter) at different approach velocities, with and without 2 V DC voltage. (B) Dependence of log removal of latex particles on V−2/3, where V is the approach velocity. Solution conditions: 10 mM NaCl solution at pH 9 and 7.6 × 1012 particles per mL (410 mg L−1) of latex particles. The latex particle suspension (10 mL) was filtered through the MWNT at three different approach velocities (280, 350, and 560 L m−2 h−1). At least two runs were conducted for each experimental condition, and the average values are presented along with the error bars (one standard deviation).

hinder viral deposition on the MWNT filter. However, upon application of a 2 V DC potential, the filter performance increased markedly, attaining complete removal of all influent viruses, both in the presence and absence of Ca2+ ions in solution. As with the previous solution chemistries studied (ionic strength and pH), Ca2+ ions do not impact the performance of the EC-MWNT filter when a voltage of 2 V is applied. Almost complete inactivation of the viruses (only 3 PFU mL−1 desorbed from the filter) was achieved in the presence of Ca2+ ions and application of a 2 V DC voltage (Figure 4B). Conversely, under identical experimental conditions, 2145 PFU mL−1 of culturable viruses were desorbed when the electrolyte solution contained only NaCl. This also suggests that calcium bridging between viral particles and MWNT surface is minor in the presence of SRHA. Otherwise, the number of culturable viruses desorbed from the filter would be higher in the presence of Ca2+ ions since calcium bridging to the SRHA would likely shield the viruses from possible inactivation. Mechanisms of Enhanced Viral Removal and Inactivation in the Presence of DC Voltage. We hypothesize that

The application of DC potential also resulted in a significant improvement in viral inactivation over the range of ionic strengths investigated (Figure 3B). For example, after filtering viral particles in 100 mM NaCl solution without DC potential, the desorbed viruses (210,000 PFU mL−1) from the MWNT filter were significantly higher than the desorbed viruses after filtering with a 2 V DC potential (4200 PFU mL−1). The results also suggest a negligible effect of solution ionic strength on viral inactivation in the presence of DC voltage, consistent with our previous study on electrochemical bacterial inactivation,24 and indicate that viral inactivation primarily occurs through a direct electro-oxidation mechanism. (c). Effect of Divalent Cations. In the absence of DC potential, the MWNT-filter performance increased only slightly from 1.73 to 1.92 log viral removal upon addition of Ca2+ to the SRHA solution (Figure 4A). This observation is consistent with our previous study on the decreased MWNT filter performance in the presence of NOM.17 The relatively minor increase in viral removal in the presence of Ca2+ and SRHA suggests that calcium bridging may not be a major viral removal mechanism and could be overwhelmed by repulsive steric interactions that 1561

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Figure 6. TEM images of MS2 viral particles adsorbed to MWNT surface after viral filtration experiments: (A, B) without application of DC voltage and (C, D) with application of 2 V DC voltage. Arrows are shown in C and D for oxidized viral particles. Bar indicates 20 nm.

an applied 2 V DC, the slope of the straight line becomes steeper, suggesting the influence of the applied DC potential on the latex particle transport. The difference in particle removal was higher at lower approach velocities, and it diminished as the approach velocity increased. This observation suggests that at higher approach velocities the electrostatic force exerted by the applied DC potential becomes less significant and that convective-diffusive transport dominates, in agreement with our hypothesis for increased transport due to electrophoretic mobility. The hypothesis is also supported by an increase in particle removal with increasing applied DC voltage at a constant approach velocity (Figure S4A). With increasing applied voltage from 2 to 4 V, latex particle removal increased from 1.48 to 2.06 log. Once the particles are in close vicinity of the MWNT collectors, attractive electrostatic interactions between the negatively charged particles and the positively charged anode (MWNTs) result in successful attachment of the latex particles onto the MWNT surface. As shown in Figure S4B, the removal efficiency was greatly impacted when the polarity of the MWNT filter was reversed. The particle log removal decreased from 2.4 to 0.82 when the polarity was changed from +2 V to −2 V. Upon reversing the polarity, the MWNTs become negatively charged, and the repulsive electric double layer interactions dominate and in turn inhibit the particle attachment. (b). Viral Inactivation. After deposition on the nanotubes, the viral particles in contact with the MWNT anode are inactivated through direct oxidation. Since all experiments were performed at 2 V, indirect oxidation through anodic production of oxidants, such as hydroxyl radicals and free chlorine, is very

the marked increase in the removal and inactivation of viruses under different solution chemistries in the presence of electric potential is attributed to electrophoresis and subsequent electrolysis of the viruses. Electrophoresis, the migration of viral particles by the externally applied DC voltage, enhances viral transport to the vicinity of the MWNT surface, where attractive electrostatic interactions between the positively charged anodic MWNTs and the negatively charged viral particles facilitate successful attachment. Once attached, the viral particles are oxidized at the anodic MWNT surface through direct oxidation. The underlying mechanisms of enhanced viral removal and inactivation, in the light of the proposed hypothesis, are discussed below. (a). Viral Removal. The EC-MWNT filtration of viruses involves two steps: transport and subsequent attachment of the viral particles to the MWNT surface. To test the aforementioned hypothesis for the enhanced viral removal, filtration experiments with a viral surrogate, carboxyl latex particles (40 nm in diameter), were performed under varying approach velocities and applied DC potentials. In the absence of DC potential, increasing approach velocity resulted in a lower particle removal (Figure 5A) as expected from depth-filtration theory.17,41 A similar trend with increasing approach velocity was observed in the presence of a 2 V DC potential, but particle removal was much higher than the case with no voltage for all approach velocities (Figure 5A). Log removal of latex particles as a function of V−2/3 (V being the approach velocity) is presented in Figure 5B. The linear dependence on V−2/3 indicates the importance of convectivediffusive transport in the removal of latex particles in both the presence and absence of DC potential. However, in the case of 1562

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Figure 7. Fouling characteristics of the MWNT filter with different types of natural organic matter (SRHA, Suwannee River humic acid and SRNOM, Suwannee River natural organic matter). (A) Pressure drop and permeate flux as a function of volume of solution filtered. (B) SEM micrograph of the MWNT filter after filtering 1 L of foulant solution. Filters were conditioned by pumping the background electrolyte solution (120 mL) for 30 min and then 10 mg L−1 (as TOC) of the specific natural organic matter in 7 mM NaCl plus 1 mM CaCl2 solution at a constant flow rate of 4 mL/min (or permeate flux of 280 L m−2 h−1). The solution pH was maintained at 5.5, and the temperature was fixed at 23 °C. Bar indicates 1 μm.

unlikely.24,42 Hence, the direct oxidation of the viral capsid is the most plausible explanation for the enhanced viral inactivation in the presence of DC potential. The virus inactivation could possibly be evidenced by the slight morphological changes of adsorbed MS2 viruses as seen in TEM images (Figure 6). The oxidation of the viral particles may also occur through redox transformations of the nucleic acids.43 A detailed mechanistic description of pathogen (bacterial) inactivation through direct oxidation by the ECMWNT filter was presented in our previous study.24 Insignificant Fouling of MWNT Filter by Natural Organic Matter. Both SRHA and SRNOM were used to determine the organic fouling propensity of the MWNT filter. The EC-MWNT filter exhibited minimal fouling by natural organic matter (Figures 7A and S5). The pressure drop across the MWNT filter remained nearly constant over the fouling experiments (210 and 600 min for Figures 7A and S5, respectively) performed by filtering electrolyte solutions containing 10 mg L−1 SRNOM/SRHA (as TOC) and 1 mM Ca2+ ions at a constant permeate flux of 280 L m −2 h−1. No fouling was observed even in an extended fouling test (600 min or 2400 mL filtered volume) with 10 mg L−1 SRHA (as TOC) and 1 mM Ca2+ ions, operated at a constant permeate flux of 280 L m −2 h−1 (Figure S5). Furthermore, SEM images of ECMWNT filters showed a minimal accumulation of NOM on the top surface of the filter (Figure 7B). Generally, microfiltration (MF), which separates particles from solutions by a sieving mechanism, is prone to organic fouling through NOM adsorption and cake layer formation of NOM aggregates on the membrane surface.44,45 However, our EC-MWNT filter, while having water permeability typical of MF, exhibits superior performance compared to MF as it removes viruses completely and does not foul in the presence of NOM over the wide range of solution conditions tested. Implications. The high performance of the EC-MWNT filter in the presence of NOM over a wide range of solution chemistries underscores its potential for application as a pointof-use technology for water purification. The extraordinary removal/inactivation rate of viruses in the presence of high levels of NOM along with the low fouling propensity brings the EC-MWNT filter one step closer to real-world applications.

Additional studies should examine the potential self-cleaning of the filter by electrochemical oxidation of adsorbed organic matter and the inhibition of biofilm formation through inactivation of retained microbes by direct oxidation. The long-term performance of the filter with natural waters should also be investigated.

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ASSOCIATED CONTENT

S Supporting Information *

Schematic and pictures of the electrochemical MWNT filter apparatus (Figure S1); schematic of the experimental setup for EC-MWNT filter fouling experiments (Figure S2); SEM images of the MWNT filter (Figure S3); carboxyl latex particle removal by the EC-MWNT filter at different applied voltages and approach velocities (Figure S4); long-term fouling characteristics of the EC-MWNT filter with SRHA (Figure S5); characteristics of MS2 viral particles and MWNT filter media under different solution chemistries (Table S1); and number balance of virus removal and approximate percent recovery and inactivation of viruses adsorbed to the MWNT filter at different solution pH conditions (Table S2). This material is available free of charge via the Internet at http:// pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Author

*Phone: (203) 432-2789. E-mail: menachem.elimelech@yale. edu.

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ACKNOWLEDGMENTS M.S.R. thanks the Natural Sciences and Engineering Research Council of Canada (NSERC) for his postdoctoral fellowship. We also acknowledge the support of the National Science Foundation under Research Grant CBET-0828795.

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REFERENCES

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