We investigated the effect of coagulant basicity on bacteriophage removal from river water by polyferric chloride (PFC). PFC at three basicities (basicity 0.9–2.1) was prepared by means of base titration (NaOH was added to ferric chloride (FeCl3) solution) and the virus removal efficiencies of those PFCs were compared with that of FeCl3 (basicity 0). The virus removal efficiencies of the PFCs were equal to or less than that of FeCl3 at both pH 6 and pH 8. This suggests that, unlike aluminum-based coagulants, increasing the basicity of iron-based coagulants does not improve virus removal efficiency. Furthermore, the relative abundance of monomeric iron(III) species in the PFCs decreased, whereas that of precipitated iron(III) species increased with increasing basicity, as assessed with a ferron method. Colloid charge density also decreased with increasing basicity. Therefore, it is likely that the reduction in the abundance of monomeric iron(III) species led to the reduction in colloid charge density, which then reduced virus removal efficiency. Thus, the development of novel iron-based coagulants with increased virus removal efficiency may not be possible by simply increasing the basicity of the coagulant.
Coagulation is a drinking-water treatment process that removes contaminants such as particulate and organic matter. Aluminum and iron salts are widely used coagulants that destabilize and combine contaminants into large flocs that can be easily separated from the source water by sedimentation and filtration processes. Waterborne enteric viruses that do not settle from suspension under the influence of gravity can also be removed by coagulation with aluminum- and iron-based coagulants (Abbaszadegan et al. 2007; Mayer et al. 2008; Shirasaki et al. 2010; Shin & Sobsey 2015). Therefore, coagulation is an essential part of the multiple-barrier approach to controlling the risk to human health posed by virus contamination in drinking water.
The efficiency of virus removal by coagulation is strongly influenced by factors such as source water quality, the nature and concentrations of coagulants used, and the mixing conditions (Hijnen & Medema 2010). The quality of source drinking water is being deteriorated by continued population growth and urbanization, resulting in increased contamination of drinking-water sources with wastewater, biosolids, and emerging contaminants (Shi et al. 2012). This deterioration reduces the overall efficiency of commercially available aluminum- or iron-based coagulants, and thus the efficiency of virus removal. Therefore, coagulation processes that effectively and inexpensively remove contaminants, including viruses and precursors of disinfection by-products (i.e. natural organic matter (NOM)), are urgently needed.
Prepolymerized aluminum coagulants (e.g. polyaluminum chloride (PACl)) and prepolymerized iron coagulants (e.g. polyferric chloride (PFC)) with improved coagulation efficiency have been developed. Since coagulant basicity is an important factor in determining the relative abundances of the aluminum or iron hydrolyte species produced in these coagulants, and in turn the coagulation efficiency (Lei et al. 2009), the influence of the aluminum or iron hydrolysis ratio (basicity = (OH−)/(Al3+) or (OH−)/(Fe3+)) on the coagulation process has been investigated (Yan et al. 2007; Lei et al. 2009; Yang et al. 2011; Kimura et al. 2013). We recently investigated the effect of basicity on the ability of PACl to remove viruses from water by comparing the virus removal efficiencies of PACl at several different basicities and reported that the PACls with higher basicities (basicity ≥ 2.1) had greater virus removal efficiency than did commercially available aluminum-based coagulants (aluminum chloride solution or alum, basicity 0) or the PACls with lower basicities (basicity, 0.9, 1.5, or 1.8) (Shirasaki et al. 2014). However, the relationship between the basicity and virus removal efficiency of PFC is yet to be fully investigated.
Here, we investigated the relationship between the basicity and virus removal efficiency of PFC by comparing the virus removal efficiencies of PFC at several different basicities with that of ferric chloride (FeCl3) solution.
MATERIALS AND METHODS
Source water and coagulants
River water was sampled from the Toyohira River (Sapporo, Japan) on 4 December 2012 and subjected to water quality analyses (Table 1). FeCl3 solution (basicity 0) was prepared by dissolution of reagent-grade iron(III) chloride hexahydrate (FeCl3•6H2O; Wako Pure Chemical Industries, Osaka, Japan) in Milli-Q water (Milli-Q Advantage; Millipore Corp., Billerica, MA, USA). PFCs with basicities of 0.9, 1.5 and 2.1 (PFC-0.9, PFC-1.5, and PFC-2.1, respectively) were prepared by means of a base titration method as described previously (Lei et al. 2009). Briefly, a certain amount of 0.1 M FeCl3 solution was transferred to a glass beaker and then the solution was titrated with 0.5 M NaOH at a constant rate (4 μL/s) with a peristaltic pump until the target basicity was reached. Throughout the titration, the solution in the beaker was kept at 20 °C and stirred at 630 rpm with a magnetic stirrer. Immediately before use, the coagulants were diluted with Milli-Q water to the concentration required for each experiment.
|Dissolved organic carbon (DOC) (mg/L)||1.0|
|Dissolved organic carbon (DOC) (mg/L)||1.0|
The relative proportions of the iron(III) hydrolyte species in the coagulants were determined by means of the ferron method described by Gao et al. (2007), with minor modifications, after the coagulant was diluted with Milli-Q water to a concentration of 2.8 g-Fe/L (i.e. 0.05 M-Fe). Based on the kinetic differences between the reactions of the iron(III) species and the ferron reagent (8-hydroxy-7-iodoquinoline-5-sulfonic acid; Wako Pure Chemical Industries), iron(III) hydrolyte species were categorized as monomeric species, polymeric species, or precipitated species (Fea, Feb, and Fec, respectively) (Gao et al. 2007). After addition of the ferron reagent to the diluted coagulant, the mixture was magnetically stirred for 10 s at 400 rpm, and then the absorbance at 600 nm was measured with a UV-1700 PharmaSpec spectrophotometer (Shimadzu Corp., Kyoto, Japan) at predetermined reaction times. The iron(III) hydrolyte species were operationally divided into three categories as follows: Fea, species that reacted with ferron within 30 s; Feb, species that reacted with ferron within 120 min (absorbance at 120 min minus the absorbance due to Fea); and Fec, species that did not react with ferron (Fec = Fet – (Fea + Feb), where Fet = total Fe). To obtain Fet, we adjusted the pH of the diluted coagulant to approximately 0.5 with ultrapure nitric acid (Kanto Chemical Co., Tokyo, Japan), heated the mixture for 3 h at 85 °C in a muffle furnace, cooled it to room temperature, and then analyzed it by means of the ferron method as described for Fea.
Colloid titration method
The positive colloid charge of each coagulant was determined by means of a colloid titration method after dilution of the coagulant with Milli-Q water to 2.1 − 4.1 mg-Fe/L, as we reported previously (Shirasaki et al. 2014).
Escherichia coli phage MS2 (NBRC 102619), an F-specific RNA bacteriophage, was obtained from the National Institute of Technology and Evaluation Biological Research Center (Kisarazu, Japan). MS2 is a prototype member of the genus Levivirus in the Leviviridae family. The genome of this bacteriophage contains a single molecule of linear, positive-sense, single-stranded RNA, which is encapsulated in an icosahedral protein capsid with a diameter of 24–26 nm (Fauquet et al. 2005). Because of its morphological similarities to hepatitis A virus and poliovirus, the removal of which is important during drinking-water treatment, MS2 is widely used as a surrogate for waterborne enteric viruses in coagulation experiments (Abbaszadegan et al. 2007; Mayer et al. 2008; Shirasaki et al. 2010, 2014; Matsushita et al. 2011; Kreissel et al. 2014; Shin & Sobsey 2015). MS2 was propagated and purified prior to the preparation of a bacteriophage stock solution as described previously (Shirasaki et al. 2010).
Batch coagulation experiments were conducted with 1,000 mL of bacteriophage-spiked river water in square plastic beakers at 20 °C. The bacteriophage stock solution was added to the river water at approximately 108 plaque forming units (PFU)/mL (C0), and the spiked water was then mixed with an impeller stirrer. After enough HCl or NaOH was added to the water to bring the final pH to 6 or 8, coagulant was injected into the water at a dosage of 2.24, 4.48, or 11.2 mg-Fe/L (i.e. 40, 80, or 200 μM-Fe). The water was stirred rapidly for 1 min (G = 200 s–1, 136 rpm) and then slowly for 10 min (G = 20 s–1, 29 rpm) and left at rest for 60 min to allow the floc particles generated to settle. A sample of the supernatant was then used for quantification of bacteriophage concentration (Cs) and turbidity. A portion of the supernatant was also filtered through a polycarbonate membrane filter (nominal pore size, 0.4 μm; Isopore, Millipore Corp.) and then used for quantification of bacteriophage concentration in the filtrate (Cf). In addition, a portion of the supernatant was filtered through a polytetrafluoroethylene membrane filter (nominal pore size, 0.45 μm; Dismic-25HP; Toyo Roshi Kaisha, Ltd, Tokyo, Japan) and then used for quantification of the ultraviolet absorbance at 260 nm (an indication of NOM concentration) and for measurement of the residual iron concentration after settling. Turbidity and ultraviolet absorbance at 260 nm were quantified with a 2100AN turbidity meter (Hach Company, Loveland, CO, USA) and a UV-1700 PharmaSpec spectrophotometer, respectively. After ultrapure nitric acid (Kanto Chemical Co.) was added to the membrane filtrate (1% v/v), the iron concentration was determined by means of inductively coupled plasma–mass spectrometry (Agilent 7700 series, Agilent Technologies, Inc., Santa Clara, CA, USA).
Infectious bacteriophages were enumerated by determining the number of PFUs according to the double-layer method (Adams 1959) with E. coli (NBRC 13965) as the bacterial host. The average plaque count of triplicate plates prepared from one sample was considered the infectious bacteriophage concentration for that sample.
Real-time reverse transcription-polymerase chain reaction method
Viral RNA of bacteriophages was quantified by means of a real-time reverse transcription-polymerase chain reaction (RT-PCR) method as described previously (Shirasaki et al. 2010). This method detects all bacteriophages regardless of their infectivity or the presence of aggregates.
RESULTS AND DISCUSSION
Effect of coagulant basicity on the removal of turbidity and UV260-absorbing NOM, and on the residual iron concentration
At the Moiwa drinking water treatment plant (Sapporo, Japan), which is the treatment plant for the water source used in the present study, the coagulation process is conducted by using PACl at an approximate pH of 7. Since the optimum pH for iron-based coagulants is generally lower than that for aluminum-based coagulants (American Water Works Association 1990), we decided to use a pH of 6 in the present study. In addition, because approximately 1‒2 mg-Al/L (i.e. 40‒80 μM-Al) of PACl is usually dosed in the Moiwa drinking water treatment plant, 80 μM-Fe (i.e. 4.48 mg-Fe/L) of coagulant dosage was used in the present study at pH 6.
Effect of coagulant basicity on bacteriophage removal
The removal ratios for total MS2, as evaluated by means of an RT-PCR method, were also larger when FeCl3 or PFC-0.9 was used than when PFC-1.5 or PFC-2.1 was used, not only after settling but also after membrane filtration (Figure 2(b)). These results indicate that virus removal efficiency was reduced at higher coagulant basicities. Since this trend of increasing basicity reducing the effectiveness of the PFC was also observed for the removal of UV260-absorbing NOM (Figure 1), it is likely that the mechanisms that destabilize the dispersion of viruses and NOM during the coagulation process, that is, the coordination reactions between coagulant species and the carboxyl groups of the virus surface proteins or NOM, are similar for the two types of contaminant (Bratby 2006).
First, virus removal efficiencies of PFCs were similar to or lower than that of FeCl3 (basicity 0), suggesting that increasing the basicity of iron-based coagulants does not improve virus removal efficiency. Therefore, the development of novel iron-based coagulants for efficient virus removal may not be achievable simply by increasing the basicity of the coagulant.
Second, monomeric iron(III) species likely play a more important role than precipitated iron(III) species in virus removal during coagulation with iron-based coagulants, because the virus removal ratio decreased as the amount of monomeric iron(III) species in the PFCs decreased (that is, as precipitated iron(III) species increased). Finally, the reduction of the colloid charge density of the PFCs used in the present study with increasing coagulant basicity was likely associated with the reduction in virus removal efficiency.
This research was supported in part by a Grant-in-Aid for Young Scientists A (no. 25709044, 2013), a Grant-in-Aid for Scientific Research S (no. 24226012, 2012), and a Grant-in-Aid for Scientific Research B (no. 15H04064, 2015) from the Japan Society for the Promotion of Science. Support was also provided by a Health and Labour Sciences Research Grant (Research on Health Security Control) from the Ministry of Health, Labour and Welfare of Japan, and by a grant from the Kurita Water and Environment Foundation (no. 14A007, 2014).