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Self-accelerating sulfur reduction via polysulfide to realize a high-rate sulfidogenic reactor for wastewater treatment
Accepted Manuscript Self-accelerating sulfur reduction via polysulfide to realize a high-rate sulfidogenic reactor for wastewater treatment Liang Zhang, Zefeng Zhang, Rongrong Sun, Shuang Liang, Guang-Hao Chen, Feng Jiang PII:
S0043-1354(17)30990-9
DOI:
10.1016/j.watres.2017.11.062
Reference:
WR 13392
To appear in:
Water Research
Received Date: 16 May 2017 Revised Date:
27 November 2017
Accepted Date: 28 November 2017
Please cite this article as: Zhang, L., Zhang, Z., Sun, R., Liang, S., Chen, G.-H., Jiang, F., Selfaccelerating sulfur reduction via polysulfide to realize a high-rate sulfidogenic reactor for wastewater treatment, Water Research (2017), doi: 10.1016/j.watres.2017.11.062. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
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Self-accelerating sulfur reduction via polysulfide to realize a high-rate
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sulfidogenic reactor for wastewater treatment
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Liang Zhanga,c,1, Zefeng Zhanga,1, Rongrong Suna, Shuang Lianga, Guang-Hao Chend,
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Feng Jianga,b*
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a
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China
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b
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Guangzhou, China
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c
Department of Bioscience, Aarhus University, Aarhus, Denmark
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d
Department of Civil & Environmental Engineering, Chinese National Engineering
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Research Center for Control & Treatment of Heavy Metal Pollution (Hong Kong
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Branch) and Water Technology Center, The Hong Kong University of Science and
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Technology, Clear Water Bay, Hong Kong, China
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School of Chemistry & Environment, South China Normal University, Guangzhou,
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Key Laboratory of Theoretical Chemistry of Environment, Ministry of Education,
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*Corresponding author
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E-mail: [email protected]
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1: The authors contributed equally to this work.
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ACCEPTED MANUSCRIPT Abstract
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Sulfur reduction is a promising alternative to sulfate reduction as it can generate
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sulfide at a low cost for the precipitation of heavy metals or autotrophic
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denitrification in wastewater treatment. However, the extremely low water solubility
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of elemental sulfur limits its bioavailability and results in a low sulfur-reduction rate.
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Polysulfide, which is naturally generated through reactions between sulfur and
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sulfide, can enhance the bioavailability of sulfur and thus contribute to high-rate
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sulfur reduction. Based on this principle, a laboratory-scale sulfur-reducing
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bioreactor was designed in this study for wastewater treatment. After 164 days of
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operation, the sulfide production rate (SPR) in the bioreactor reached 126 mg S/L-h,
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which is significantly higher than those of other sulfate-reducing systems. Moreover,
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dissolved zero-valent sulfur (referred to as polysulfide) was detected in the
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sulfur-reducing reactor when the organics were completely depleted, indicating that
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polysulfide can form naturally and be readily reduced to sulfide in the bioreactor. We
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found that the produced sulfide promoted the formation of more polysulfide, which
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enabled a self-accelerating chain reaction of sulfur reduction via polysulfide. This
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stimulation effect was further validated by the seven-hour batch tests. In the batch
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test without sulfide addition initially, a continuous increase in the hourly SPR was
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observed with increasing sulfide concentration. Furthermore, in the batch tests with
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the addition of 50 to 200 mg S/L sulfide at the beginning, the average SPR in the first
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three hours increased with elevating initial sulfide concentration due to more
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ACCEPTED MANUSCRIPT polysulfide formation and reduction. However, high sulfide concentration (> 250 mg
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S/L) hindered the continuous increase in SPR. Additionally, when polysulfide
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formation was prevented through the addition of Fe2+, the SPR dropped by 97.6%
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compared to that in the presence of polysulfide. This validates the key role of
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polysulfide in the high-rate sulfur reduction process. Overall, the findings suggest
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that high-rate sulfur reduction can be achieved for autotrophic denitrification or
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heavy-metal removal in wastewater treatment.
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Keywords: Indirect sulfur reduction, Polysulfide, Sulfide addition, Sulfidogenic
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process, Metal removal, Autotrophic denitrification
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1. Introduction
Sulfidogenic processes can not only oxidize organics anaerobically and
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efficiently, but also provide sulfide as a source of electron donors for autotrophic
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denitrification in low carbon-to-nitrogen wastewater treatment (Lu et al., 2012,
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Fajardo et al., 2014, van den Brand et al., 2015), thus enabling the natural
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integration of anaerobic mainstream treatment with nitrogen removal (Wu et al.,
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2016). Sulfidogenic processes have also been widely employed for metallurgical
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wastewater treatment through metal sulfide precipitation (Muyzer and Stams, 2008,
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Sánchez-Andrea et al., 2014, Zhang et al., 2016).
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To date, studies have mostly focused on sulfate reduction processes and their
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ability to provide biogenic sulfide for autotrophic denitrification processes or metal
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precipitation. Florentino et al. (2016), on the other hand, suggested that sulfur
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reduction processes can potentially be used for metal removal and recovery from
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acid mine drainage and are more cost effective than sulfate-reducing processes as
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they consume up to 75% less organics theoretically. However, the extremely low
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water solubility of sulfur (5 µg/L at 25 °C) may be a limiting factor for high-rate sulfur
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reduction processes. Inexpensive and common sulfur sources such as sublimated
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and chemical sulfur from sublimation and Claus processes are almost insoluble and
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thus not very bioaccessible to sulfur reducers (Florentino et al., 2015).
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In this study, a promising solution to the aforementioned issues is proposed
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ACCEPTED MANUSCRIPT wherein elemental sulfur is converted to polysulfide for use by sulfur reducers.
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Polysulfide can form naturally from the nucleophilic attack of elemental sulfur by HS-
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in the presence of sulfide, resulting in the nucleophilic cleavage of S8 rings (Equation
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1) (Hedderich et al., 1998, Kletzin et al., 2004, Boyd and Druschel, 2013). Polysulfide
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can be readily reduced by sulfur reducers or sulfate-reducing bacteria (SRB)
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(Equation 2, taking formate as an example) (Schauder and Müller, 1993, Hedderich
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et al., 1998, Liang et al., 2016), and an increase in HS- can generate additional
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polysulfide (Equation 2). This in turn provides more electron acceptors (polysulfide)
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to sulfur reducers, resulting in higher sulfide production. We hypothesize that the
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interaction between polysulfide formation and reduction triggered by sulfide creates
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a self-accelerating chain reaction of sulfur reduction via polysulfide in a bioreactor. A
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high rate of sulfur reduction is achieved until all elemental sulfur and organic matter
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(electron donors) are exhausted. If this hypothesis is correct, a sulfidogenic
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bioreactor capable of sulfur reduction at a very high rate can be developed with
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elemental sulfur as the low-cost sulfur source for efficient metal removal or
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autotrophic denitrification. Although Blumentals et al. (1990) and Boyd and Druschel
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(2013) have suggested through pure culture studies that polysulfide is involved in
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biological elemental sulfur reduction as an intermediate under hydrothermal
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conditions, it remains unknown if high-rate sulfur reduction in wastewater treatment
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can be achieved via polysulfide.
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HS− +
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n −1 2− S8 → Sn + H + 8
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−
2−
−
2−
HCO2 + Sn + H2O → HCO3 + HS− + Sn−1 + H +
(2)
Therefore, in this study, we investigated the long-term feasibility of high-rate
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sulfur reduction in a laboratory-scale sulfur-reducing bioreactor. Several batch tests
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were conducted to systematically elucidate the role of polysulfide in biological sulfur
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reduction using a cultivated sulfur-reducing sludge. Finally, the mechanisms and
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conditions for realizing high-rate sulfur reduction are proposed and discussed.
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2. Materials and Methods
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2.1. Setup and operation of the sulfur-reducing bioreactor
The sludge in wastewater treatment plants in Hong Kong generally
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contains large amounts of SRB (Wang et al., 2011, Ye and Zhang, 2013). The sulfate
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levels in sewage are as high as 600–1000 mg/L due to seawater toilet flushing (Jiang
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et al., 2009). Thus, sludge was taken from the Shatin sewage treatment plant and
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seeded into a laboratory-scale sulfur-reducing anaerobic fluidized bed (SRAFB)
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bioreactor to enrich sulfur reducers. The SRAFB reactor was made of Plexiglas with
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an effective volume of 3.02 L as shown in Fig. S1. The bioreactor was fed with
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synthetic wastewater prepared following the method described in our previous
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study (Jiang et al., 2013) and continuously operated for 164 days, divided into six
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stages (from stage 1 to stage 6) based on different hydraulic retention times (HRTs)
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(ranging from 3.0 to 13.5 h as shown in Table S1). Sublimed sulfur particles (20–40
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bioreactor based on the daily sulfur consumption. More detailed operational
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information is provided in the supporting information. During the operation of the
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bioreactor, total organic carbon (TOC), dissolved sulfide, sulfate, thiosulfate, alkalinity,
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volatile fatty acids (VFAs), and pH values in the influent and effluent samples were
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measured daily. The polysulfide level in the bioreactor was monitored by running the
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bioreactor as a sequential batch reactor at the end of the operational period.
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Subsequently, sulfur-reducing sludge samples were taken for further batch tests to
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investigate biological sulfur reduction with and without polysulfide.
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2.2. Polysulfide analysis in the sulfur-reducing bioreactor
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In order to confirm the important role of polysulfide in high-rate sulfur
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reduction, polysulfide levels in the sulfur-reducing bioreactor were monitored. In this
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batch test, the bioreactor was run as a sequential batch reactor. Approximately 840
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mg/L sodium bicarbonate and 50 mg/L TOC diluted with stock synthetic
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wastewater (Jiang et al., 2013) was supplied to the sulfur-reducing bioreactor. The
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batch test lasted for 15 h, but after the first 9.2 h approximately two-thirds of the
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supernatant were renewed with the diluted stock synthetic wastewater to obtain 50
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mg/L TOC because the TOC was completely depleted after 9 h according to the
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results of the pre-tests. The mixed liquid samples were collected every 1.5 h through
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the top opening using a 5 mL syringe and then filtered to measure TOC, sulfide, and
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polysulfide concentrations. At each sampling time point, the pH of the SRAFB reactor
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was also measured.
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2.3. Biological sulfur reduction with polysulfide
To determine the effect of sulfide on polysulfide formation and subsequent
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sulfur reduction, batch tests were conducted in duplicate using the cultivated
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sulfur-reducing sludge. The sludge was washed with deoxygenated deionized water
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and then evenly distributed into ten 2.4 L flasks. The sludge concentration in each
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flask was measured as 0.68 g/L volatile suspended solids (VSS). Initial sulfide
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concentrations of 0, 50, 100, 150, and 200 mg S/L in respective flasks were obtained
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by spiking a concentrated sodium sulfide solution. Organic carbon (100 mg/L glucose)
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and 5 g/L sublimed sulfur were added to each flask to provide sufficient carbon and
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sulfur. The flasks were purged with nitrogen gas and then sealed with rubber
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stoppers to maintain anaerobic conditions. The tests were conducted at an ambient
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temperature of approximately 25°C, and the initial pH was controlled at 7.50±0.02.
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The batch tests were conducted for 7 h, during which 5 mL samples from two
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replicates were collected every two hours through a sampling port on the rubber
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stopper to measure sulfide, polysulfide, sulfate, thiosulfate, and TOC. A pH probe
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was fitted in the flask to measure pH variations with time. The sulfide production
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rate (SPR) was calculated according to Equation 3:
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sulfide production rate (mg S/L-h) =
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∆CS 2−
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∆t
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( ∆t , h) in mg S/L.
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2.4. Biological sulfur reduction without polysulfide
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The test conditions were controlled to compare direct sulfur reduction
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without polysulfide and indirect sulfur reduction via polysulfide. FeCl2 was used to
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completely precipitate the dissolved sulfide to prevent polysulfide formation (Ringel
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et al., 1996). A batch test was conducted in duplicate to evaluate sulfur reduction
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without polysulfide by adding 4.7 mM FeCl2 at the beginning of the test to 200 mL
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flasks. The sludge (0.68 g/L) was washed with deoxygenated deionized water three
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times to remove residual sulfide before FeCl2 was added. A control test without FeCl2
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addition was also performed in duplicate. The other test conditions were kept the
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same as those in section 2.3. The batch tests were performed for 9 h, during which
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samples were collected every 3 h to monitor the changes in sulfide, sulfate,
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thiosulfate, and TOC concentrations. The pH values were measured using a pH probe
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fitted to the flask.
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2.5. Chemical analysis
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The TOC, sulfide, sulfate, and thiosulfate concentrations were determined after
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filtration (Millipore, 0.45 µm). The TOC was analyzed using a TOC analyzer (Shimadzu
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TOC-5000A), and the total sulfide (H2S, HS-, and S2-) concentration was determined
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using the methylene blue method (APHA, 2005). The sulfate and thiosulfate 9
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chromatograph (DIONEX-900), and the VSS was measured according to the standard
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methods (APHA, 2005). The pH was measured using a pH meter (Hach, HQ40D). The
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acid volatile sulfide (AVS) deposited in the sludge was measured using the method
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described by Simpson (2001). Elemental sulfur in the sludge samples was extracted
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based on the method described by McGuire and Hamers (2000) and was measured
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using a high-performance liquid chromatograph (HPLC, Shimadzu LC-16, Japan)
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equipped with a Kromasil column (C18, 5μm, 100 Å) and a UV detector at 254 nm.
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Since the length of the Sn2- chain varied from 2 to 11, it was difficult to determine the
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aggregate concentration of polysulfide. In contrast, the zero-valent sulfur atoms in
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polysulfide ions (also called dissolved zero-valent sulfur) can be analyzed using an
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ultraviolet-visible spectrophotometer (UV-6000PC, Metash, Shanghai, China) at a
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wavelength of 285 nm after filtration (Millipore, 0.22 µm) (Kleinjan et al., 2005). Thus,
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the dissolved zero-valent sulfur in this study was defined as an indicator of
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polysulfide. The mixed liquid samples were taken with a 5 mL syringe and
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immediately filtered through disposable 0.22-μm filters. The filtrate was directly
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deposited in a N2-filled glass tube and immediately analyzed with the
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ultraviolet-visible spectrophotometer.
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3. Results
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3.1. Performance of the sulfur-reducing bioreactor
The anaerobic sulfur-reducing reactor was successfully operated with
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sublimated sulfur as its sole source of electron acceptors for 164 days. The SPR
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increased as the HRT of the reactor decreased and reached a maximum of 126 mg
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S/L-h (Fig. 1a). Although sublimed sulfur is nearly insoluble, this rate is significantly
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higher than those reported in previous studies, which achieved SPRs of 24 mg S/L-h
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(Wang et al., 2009), 22 mg S/L-h (Wang et al., 2008), 43 mg S/L-h (Qian et al., 2015),
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45 mg S/L-h (Jiang et al., 2013), and 72 mg S/L-h (Celis-García et al., 2007) using
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sulfidogenic processes based on sulfate/sulfite reduction. More than 80% of the TOC
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was also removed (Fig. S3). In addition, sulfate and thiosulfate were not detected in
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the influent and effluent, indicating that all of the sulfide in the effluent came from
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sulfur reduction.
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As shown in Fig. 1b, the ratio of removed organic carbon to produced sulfide
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(mg C/mg S) was close to the theoretical level of 0.19 mg C/mg S for sulfur reduction,
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as calculated from Equation 4 (Corg means the biodegradable organic carbon). This
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result reveals that biological sulfur reduction prevailed in this reactor.
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Corg + 2S 0 + 2 H 2 O → CO2 + 2 HS − + 2 H +
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3.2. Polysulfide in the sulfur-reducing bioreactor
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An increase in dissolved zero-valent sulfur concentration was clearly
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observed after the complete depletion of organic carbon in the sulfur-reducing 11
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maximum of 55 mg S/L, which is much higher than the solubility of sublimated sulfur
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in water (5 µg/L at 25 °C) (Schauder and Müller, 1993). This result reveals that the
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measured dissolved zero-valent sulfur was most likely polysulfide, namely the S0 in
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Sn2-.
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Dissolved zero-valent sulfur was not detected when TOC was present in the
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sulfur-reducing bioreactor, indicating that the rate of dissimilatory polysulfide
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reduction was much higher than the rate of polysulfide formation. This suggests that
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polysulfide formation could be the rate-limiting step for high-rate sulfur reduction.
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According to Equations 1 and 2, HS- production from sulfur reduction can facilitate
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polysulfide formation. We thus hypothesized that indirect sulfur reduction via
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polysulfide would be accelerated by an increase in sulfide concentration resulting
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from sulfur reduction.
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3.3. Sulfur reduction without sulfide initially
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To verify the aforementioned hypothesis, a batch test was performed in the
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absence of sulfide initially (refer to section 2.3). Without sulfide at the beginning of
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the batch test, the hourly SPR was only 4.2 mg S/L-h during the first hour; however,
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it climbed gradually and reached a maximum of 36.4 mg S/L-h during the seventh
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hour, which represents an approximately 8.7 times increase (Fig. 3). Polysulfide was
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detected during the entire period and reached 9.6 mg S/L by the end of the batch
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batch test, or in subsequent batch tests.
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3.4. Sulfur reduction with sulfide initially
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Batch tests were further performed with different concentrations of sulfide
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added (50, 100, 150, and 200 mg S/L) at the start of the tests to assess the feasibility
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of artificially stimulating sulfur reduction.
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Sulfide addition stimulated polysulfide formation at the start of the tests (Fig.
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S4). Thereafter, the polysulfide concentrations decreased with time in all of the
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flasks due to polysulfide reduction. As a result, the increased initial concentration of
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sulfide enhanced sulfur reduction (Fig. 4). At the start of the tests, the highest hourly
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SPR (53.7 mg S/L-h) was achieved with the highest sulfide dosage (200 mg S/L).
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Within the first 3 h, a linear correlation was observed between the average SPR and
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the initial sulfide dosage (Fig. S5a). The organic reduction rates followed a similar
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trend (Fig. S5b) and the organic compounds were completely depleted six and seven
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hours in the tests when the initial dosages were 150 and 200 mg S/L, respectively
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(Fig. S6).
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However, there is a limit to how much sulfide can stimulate sulfur reduction.
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Once the sulfide concentration in the flasks exceeded 250 mg S/L, the stimulation
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effect of sulfide addition on sulfur reduction weakened over time. For instance, the
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batch tests with 150 and 200 mg/L sulfide additions exhibited high hourly SPRs 13
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in the tests with just 50 mg S/L sulfide added, the hourly SPR just kept on increasing,
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similar to the test without sulfide addition. These results reveal that accelerating
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sulfur reduction only applies at sulfide levels below ~250 mg S/L.
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3.5. Sulfur reduction without polysulfide
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To compare the rates of direct and indirect sulfur reduction, a batch test was
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conducted in which polysulfide formation was prevented by adding Fe2+ (refer to
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section 2.4). In the absence of polysulfide, only 0.7 mg S of AVS had accumulated by
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the end of the batch test, which was just 2.4% of the amount of sulfide produced in
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the batch test without polysulfide control (28.6 mg S) (Fig. 5a). This result indicates
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that the rate of sulfur reduction decreased by 97.6% in the absence of polysulfide.
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Moreover, organic consumption in the batch tests with Fe2+ addition (3%) was
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significantly lower than that in the batch tests without (66%) (Fig. 5b).
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4. Discussion
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4.1. Mechanisms of high-rate sulfur reduction
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A high-rate sulfidogenic process was demonstrated in a sulfur-reducing
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bioreactor using elemental sulfur as its sole source of electron acceptors, although
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elemental sulfur is almost insoluble. We found that polysulfide was present in the
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We therefore hypothesized that the soluble polysulfide was the main intermediate
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which first boosted sulfur bioavailability and then accelerated sulfur reduction. Batch
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tests were further conducted to verify this hypothesis, and the results were
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affirmative. Previous studies (Kleinjan et al., 2005, Sigel and Sigel, 2005, Florentino et
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al., 2016) have suggested that an increase in sulfide concentration enhances
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polysulfide formation. Furthermore, polysulfide can be easily reduced to sulfide by
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sulfur reducers (Schauder and Müller, 1993, Florentino et al., 2016, Liang et al.,
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2016), which in turn promotes polysulfide formation through the abiotic reaction
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between sulfur and sulfide at neutral or alkaline conditions (Boyd and Druschel,
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2013). To the best of our knowledge, the present study is the first to demonstrate
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the application of high-rate sulfur reduction via polysulfide formation in wastewater
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treatment.
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The mechanism of high-rate sulfur reduction is the indirect sulfur reduction
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enabled by polysulfide, which occurs through the pathways proposed in Fig. 6. The
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sulfide produced through sulfur reduction promotes polysulfide formation (Equation
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1), which is the rate-limiting step of the indirect sulfur reduction process. Thereafter,
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the sulfur reducers reduce polysulfide to sulfide (Equation 2), which promotes the
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formation of more polysulfide akin to a chain reaction. This chain reaction
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continuously enhances sulfur reduction until it is suppressed due to the toxicity of
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the high sulfide levels. Accordingly, the rate of sulfide production increased
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this study (see Fig. 3).
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4.2. Conditions for high-rate sulfur reduction
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The results also reveal that the bioavailability of elemental sulfur can be
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significantly enhanced by introducing polysulfide as an intermediate to enable
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high-rate indirect sulfur reduction. Thus, the key prerequisites for high-rate sulfur
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reduction are polysulfide formation and reduction.
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Moreover, maintaining neutral or alkaline pH levels is essential for polysulfide
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formation (Florentino et al., 2016). Polysulfide is unstable and easily decomposes to
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form elemental sulfur and sulfide at acidic pH levels (Boyd and Druschel, 2013,
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Florentino et al., 2016). Schauder and Müller (1993) reported that polysulfide can
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barely be detected at pH levels below 6. Therefore the pH level in a high-rate
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sulfur-reducing reactor must be at least 6 and was kept at an average of 6.7 in our
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sulfur-reducing bioreactor (Fig. S7) to maintain suitable conditions for polysulfide
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formation.
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An increase in sulfide concentration in a sulfur-reducing reactor stimulates
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sulfur reduction by facilitating polysulfide formation (Kleinjan et al., 2005, Sigel and
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Sigel, 2005, Florentino et al., 2016). At the start of a batch test, apart from manual
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addition, sulfate reduction may also provide sulfide to promote polysulfide
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formation if sulfate is present in the wastewater. However, too much sulfide (> 250 16
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to the toxicity of unionized hydrogen sulfide to sulfidogenic bacteria. Hydrogen
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sulfide has been found to exert a direct and reversible toxicity effect on SRB (Reis et
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al., 1992) with 100% inhibition of SRB growth at H2S concentrations varying from 477
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to 617 mg/L (Reis et al., 1992, Kolmert et al., 1997, Neculita et al., 2007).
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In addition, preventing polysulfide formation by adding Fe2+ evidently reduced
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the sulfur reduction rate. Although elemental sulfur could be reduced without
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polysulfide, it occurred at a very low rate compared to the rate of indirect sulfur
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reduction. This result suggests that high-rate sulfur reduction cannot be achieved
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without polysulfide. Thus, we propose that a two-stage system consisting of a
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high-rate
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independently would be more suitable for metal-laden wastewater treatment than
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the single-stage system presented by Florentino et al. (2016). In addition, the sulfur
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reduction process can provide sufficient electron donors (sulfide) for subsequent
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autotrophic denitrification in low carbon-to-nitrogen wastewater treatment.
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5. Conclusions
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This study achieved high-rate sulfur reduction in a sulfur-reducing bioreactor
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and investigated its mechanisms, which are discussed in this paper. The main findings
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are: 17
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•
The high-rate sulfur reduction was achieved using insoluble sublimated sulfur as the sole source of electron acceptors. The highest rate of sulfide production
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attained in the bioreactor was 126 mg S/L-h, which is significantly higher than
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those previously reported for other sulfidogenic processes.
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The high-rate sulfur reduction was attributed to indirect sulfur reduction driven by polysulfide. The sulfide produced through sulfur reduction induced both
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polysulfide formation and reduction to produce more sulfide, thus enabling the
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self-acceleration of sulfur reduction. •
•
Neutral or alkaline conditions, along with a low metal concentration in the influent, are essential for high-rate sulfur reduction.
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The continuous increase in sulfide production rate was observed until the sulfide concentration in the bioreactor exceeded 250 mg/L.
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These findings suggest that high-rate sulfur reduction can be achieved at a low cost
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and represents a promising technology for autotrophic denitrification or the removal
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of heavy metals in wastewater treatment.
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Acknowledgements
The authors acknowledge the support from the National Natural Science
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Foundation of China (51178914 and 51638005), the Guangdong Provincial Science
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and Technology Planning Project (2016A050503041 and 2017B050504003), and the
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Hong Kong Innovation and Technology Commission (ITC-CNERC14EG03).
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Liang, S., Zhang, L. and Jiang, F. (2016) Indirect sulfur reduction via polysulfide contributes to serious odor problem in a sewer receiving nitrate dosage. Water Res. 100, 421-428.
Lu, H., Wu, D., Jiang, F., Ekama, G.A., van Loosdrecht, M. and Chen, G.H. (2012) The demonstration of a novel sulfur cycle based wastewater treatment process: Sulfate reduction, autotrophic denitrification, and nitrification integrated (SANI®) biological nitrogen removal process. Biotechnol. Bioeng. 109(11), 2778-2789. McGuire, M.M. and Hamers, R.J. (2000) Extraction and quantitative analysis of elemental sulfur from sulfide mineral surfaces by high-performance liquid chromatography. Environ. Sci. Technol. 19
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Wang, A., Ren, N., Wang, X. and Lee, D. (2008) Enhanced sulfate reduction with acidogenic sulfate-reducing bacteria. J. Hazard. Mater. 154(1), 1060-1065. Wang, J., Lu, H., Chen, G.-H., Lau, G.N., Tsang, W. and van Loosdrecht, M. (2009) A novel sulfate reduction, autotrophic denitrification, nitrification integrated (SANI) process for saline wastewater treatment. Water Res. 43(9), 2363-2372.
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Wang, J., Shi, M., Lu, H., Wu, D., Shao, M.-F., Zhang, T., Ekama, G.A., van Loosdrecht, M.C. and Chen, G.-H. (2011) Microbial community of sulfate-reducing up-flow sludge bed in the SANI® process for saline sewage treatment. Appl. Microbiol. Biotechnol. 90(6), 2015-2025. Wu, D., Ekama, G.A., Chui, H.-K., Wang, B., Cui, Y.-X., Hao, T.-W., van Loosdrecht, M.C. and Chen, G.-H.
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(2016) Large-scale demonstration of the sulfate reduction autotrophic denitrification nitrification integrated (SANI®) process in saline sewage treatment. Water Res. 100, 496-507.
Ye, L. and Zhang, T. (2013) Bacterial communities in different sections of a municipal wastewater treatment plant revealed by 16S rDNA 454 pyrosequencing. Appl. Microbiol. Biotechnol. 97(6), 2681-2690.
Zhang, L., Lin, X., Wang, J., Jiang, F., Wei, L., Chen, G. and Hao, X. (2016) Effects of Lead and Mercury on Sulfate-Reducing Bacterial Activity in a Biological Process for Flue Gas Desulfurization Wastewater Treatment. Sci. Rep. 6.
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ACCEPTED MANUSCRIPT Figure captions Fig. 1. The performance of the sulfur-reducing bioreactor: (a) the average sulfide production and sulfide production rates for the six stages (from stage 1 to stage 6), (b) the C/S ratios for the six stages (the dashed line represents the theoretical C/S
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ratio of 0.19)
Fig. 2. The variations in TOC, sulfide and polysulfide concentrations in the sulfur-reducing bioreactor during the batch test. Organic carbon was added at 9.2 h
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as indicated by the orange arrow.
Fig. 3. (a) The hourly sulfide production rate; and (b) sulfide production and TOC
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removal as a function of time without sulfide addition at the start of the batch tests (sulfur reduction with polysulfide) (the data points are averages of two replicates). Fig. 4. (a) The hourly sulfide production rates and (b) sulfide concentration as a function of time under varying sulfide addition conditions at the start of the batch
replicates).
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tests (sulfur reduction with polysulfide) (the data points are averages of two
Fig. 5. (a) Sulfide production at pH 7.5 and pH 7.5+Fe2+ in the 9 h batch tests (the AVS
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was measured to quantify the sulfide concentration in the batch test with Fe2+ addition); (b) the corresponding TOC consumed in the control tests (the data points
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are averages of two replicates). Fig. 6. The hypothetical pathway of biological elemental sulfur reduction under neutral or alkaline conditions.
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Fig. 1. The performance of the sulfur-reducing bioreactor: (a) the average sulfide
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production and sulfide production rates for the six stages (from stage 1 to stage 6), (b) the C/S ratios for the six stages (the dashed line represents the theoretical C/S
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Fig. 2. The variations in TOC, sulfide and polysulfide concentrations in the sulfur-reducing bioreactor during the batch test. Organic carbon was added at 9.2 h
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Fig. 3. (a) The hourly sulfide production rate; and (b) sulfide production and TOC removal as a function of time without sulfide addition at the start of the batch tests
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Fig. 4. (a) The hourly sulfide production rates and (b) sulfide concentration as a function of time under varying sulfide addition conditions at the start of the batch
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tests (sulfur reduction with polysulfide) (the data points are averages of two
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Fig. 5. (a) Sulfide production at pH 7.5 and pH 7.5+Fe2+ in the 9 h batch tests (the AVS
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Fig. 6. The hypothetical pathway of biological elemental sulfur reduction under
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ACCEPTED MANUSCRIPT Highlights High-rate sulfide production was achieved in a sulfur-reducing bioreactor
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Self-accelerating sulfur reduction was observed
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The mechanism of self-accelerating sulfur via polysulfide was characterized
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High sulfide concentration (>250 mg S/L) weakened the self-acceleration of sulfur
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reduction
S0043-1354(17)30990-9
DOI:
10.1016/j.watres.2017.11.062
Reference:
WR 13392
To appear in:
Water Research
Received Date: 16 May 2017 Revised Date:
27 November 2017
Accepted Date: 28 November 2017
Please cite this article as: Zhang, L., Zhang, Z., Sun, R., Liang, S., Chen, G.-H., Jiang, F., Selfaccelerating sulfur reduction via polysulfide to realize a high-rate sulfidogenic reactor for wastewater treatment, Water Research (2017), doi: 10.1016/j.watres.2017.11.062. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
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Self-accelerating sulfur reduction via polysulfide to realize a high-rate
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sulfidogenic reactor for wastewater treatment
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Liang Zhanga,c,1, Zefeng Zhanga,1, Rongrong Suna, Shuang Lianga, Guang-Hao Chend,
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Feng Jianga,b*
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a
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China
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b
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Guangzhou, China
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c
Department of Bioscience, Aarhus University, Aarhus, Denmark
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d
Department of Civil & Environmental Engineering, Chinese National Engineering
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Research Center for Control & Treatment of Heavy Metal Pollution (Hong Kong
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Branch) and Water Technology Center, The Hong Kong University of Science and
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Technology, Clear Water Bay, Hong Kong, China
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School of Chemistry & Environment, South China Normal University, Guangzhou,
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Key Laboratory of Theoretical Chemistry of Environment, Ministry of Education,
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*Corresponding author
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E-mail: [email protected]
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1: The authors contributed equally to this work.
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ACCEPTED MANUSCRIPT Abstract
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Sulfur reduction is a promising alternative to sulfate reduction as it can generate
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sulfide at a low cost for the precipitation of heavy metals or autotrophic
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denitrification in wastewater treatment. However, the extremely low water solubility
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of elemental sulfur limits its bioavailability and results in a low sulfur-reduction rate.
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Polysulfide, which is naturally generated through reactions between sulfur and
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sulfide, can enhance the bioavailability of sulfur and thus contribute to high-rate
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sulfur reduction. Based on this principle, a laboratory-scale sulfur-reducing
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bioreactor was designed in this study for wastewater treatment. After 164 days of
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operation, the sulfide production rate (SPR) in the bioreactor reached 126 mg S/L-h,
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which is significantly higher than those of other sulfate-reducing systems. Moreover,
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dissolved zero-valent sulfur (referred to as polysulfide) was detected in the
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sulfur-reducing reactor when the organics were completely depleted, indicating that
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polysulfide can form naturally and be readily reduced to sulfide in the bioreactor. We
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found that the produced sulfide promoted the formation of more polysulfide, which
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enabled a self-accelerating chain reaction of sulfur reduction via polysulfide. This
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stimulation effect was further validated by the seven-hour batch tests. In the batch
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test without sulfide addition initially, a continuous increase in the hourly SPR was
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observed with increasing sulfide concentration. Furthermore, in the batch tests with
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the addition of 50 to 200 mg S/L sulfide at the beginning, the average SPR in the first
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three hours increased with elevating initial sulfide concentration due to more
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ACCEPTED MANUSCRIPT polysulfide formation and reduction. However, high sulfide concentration (> 250 mg
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S/L) hindered the continuous increase in SPR. Additionally, when polysulfide
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formation was prevented through the addition of Fe2+, the SPR dropped by 97.6%
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compared to that in the presence of polysulfide. This validates the key role of
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polysulfide in the high-rate sulfur reduction process. Overall, the findings suggest
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that high-rate sulfur reduction can be achieved for autotrophic denitrification or
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heavy-metal removal in wastewater treatment.
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Keywords: Indirect sulfur reduction, Polysulfide, Sulfide addition, Sulfidogenic
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process, Metal removal, Autotrophic denitrification
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1. Introduction
Sulfidogenic processes can not only oxidize organics anaerobically and
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efficiently, but also provide sulfide as a source of electron donors for autotrophic
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denitrification in low carbon-to-nitrogen wastewater treatment (Lu et al., 2012,
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Fajardo et al., 2014, van den Brand et al., 2015), thus enabling the natural
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integration of anaerobic mainstream treatment with nitrogen removal (Wu et al.,
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2016). Sulfidogenic processes have also been widely employed for metallurgical
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wastewater treatment through metal sulfide precipitation (Muyzer and Stams, 2008,
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Sánchez-Andrea et al., 2014, Zhang et al., 2016).
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To date, studies have mostly focused on sulfate reduction processes and their
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ability to provide biogenic sulfide for autotrophic denitrification processes or metal
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precipitation. Florentino et al. (2016), on the other hand, suggested that sulfur
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reduction processes can potentially be used for metal removal and recovery from
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acid mine drainage and are more cost effective than sulfate-reducing processes as
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they consume up to 75% less organics theoretically. However, the extremely low
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water solubility of sulfur (5 µg/L at 25 °C) may be a limiting factor for high-rate sulfur
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reduction processes. Inexpensive and common sulfur sources such as sublimated
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and chemical sulfur from sublimation and Claus processes are almost insoluble and
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thus not very bioaccessible to sulfur reducers (Florentino et al., 2015).
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In this study, a promising solution to the aforementioned issues is proposed
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Polysulfide can form naturally from the nucleophilic attack of elemental sulfur by HS-
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in the presence of sulfide, resulting in the nucleophilic cleavage of S8 rings (Equation
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1) (Hedderich et al., 1998, Kletzin et al., 2004, Boyd and Druschel, 2013). Polysulfide
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can be readily reduced by sulfur reducers or sulfate-reducing bacteria (SRB)
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(Equation 2, taking formate as an example) (Schauder and Müller, 1993, Hedderich
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et al., 1998, Liang et al., 2016), and an increase in HS- can generate additional
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polysulfide (Equation 2). This in turn provides more electron acceptors (polysulfide)
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to sulfur reducers, resulting in higher sulfide production. We hypothesize that the
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interaction between polysulfide formation and reduction triggered by sulfide creates
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a self-accelerating chain reaction of sulfur reduction via polysulfide in a bioreactor. A
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high rate of sulfur reduction is achieved until all elemental sulfur and organic matter
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(electron donors) are exhausted. If this hypothesis is correct, a sulfidogenic
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bioreactor capable of sulfur reduction at a very high rate can be developed with
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elemental sulfur as the low-cost sulfur source for efficient metal removal or
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autotrophic denitrification. Although Blumentals et al. (1990) and Boyd and Druschel
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(2013) have suggested through pure culture studies that polysulfide is involved in
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biological elemental sulfur reduction as an intermediate under hydrothermal
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conditions, it remains unknown if high-rate sulfur reduction in wastewater treatment
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can be achieved via polysulfide.
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HS− +
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n −1 2− S8 → Sn + H + 8
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−
2−
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2−
HCO2 + Sn + H2O → HCO3 + HS− + Sn−1 + H +
(2)
Therefore, in this study, we investigated the long-term feasibility of high-rate
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sulfur reduction in a laboratory-scale sulfur-reducing bioreactor. Several batch tests
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were conducted to systematically elucidate the role of polysulfide in biological sulfur
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reduction using a cultivated sulfur-reducing sludge. Finally, the mechanisms and
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conditions for realizing high-rate sulfur reduction are proposed and discussed.
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2. Materials and Methods
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2.1. Setup and operation of the sulfur-reducing bioreactor
The sludge in wastewater treatment plants in Hong Kong generally
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contains large amounts of SRB (Wang et al., 2011, Ye and Zhang, 2013). The sulfate
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levels in sewage are as high as 600–1000 mg/L due to seawater toilet flushing (Jiang
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et al., 2009). Thus, sludge was taken from the Shatin sewage treatment plant and
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seeded into a laboratory-scale sulfur-reducing anaerobic fluidized bed (SRAFB)
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bioreactor to enrich sulfur reducers. The SRAFB reactor was made of Plexiglas with
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an effective volume of 3.02 L as shown in Fig. S1. The bioreactor was fed with
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synthetic wastewater prepared following the method described in our previous
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study (Jiang et al., 2013) and continuously operated for 164 days, divided into six
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stages (from stage 1 to stage 6) based on different hydraulic retention times (HRTs)
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(ranging from 3.0 to 13.5 h as shown in Table S1). Sublimed sulfur particles (20–40
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bioreactor based on the daily sulfur consumption. More detailed operational
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information is provided in the supporting information. During the operation of the
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bioreactor, total organic carbon (TOC), dissolved sulfide, sulfate, thiosulfate, alkalinity,
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volatile fatty acids (VFAs), and pH values in the influent and effluent samples were
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measured daily. The polysulfide level in the bioreactor was monitored by running the
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bioreactor as a sequential batch reactor at the end of the operational period.
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Subsequently, sulfur-reducing sludge samples were taken for further batch tests to
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investigate biological sulfur reduction with and without polysulfide.
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2.2. Polysulfide analysis in the sulfur-reducing bioreactor
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In order to confirm the important role of polysulfide in high-rate sulfur
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reduction, polysulfide levels in the sulfur-reducing bioreactor were monitored. In this
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batch test, the bioreactor was run as a sequential batch reactor. Approximately 840
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mg/L sodium bicarbonate and 50 mg/L TOC diluted with stock synthetic
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wastewater (Jiang et al., 2013) was supplied to the sulfur-reducing bioreactor. The
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batch test lasted for 15 h, but after the first 9.2 h approximately two-thirds of the
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supernatant were renewed with the diluted stock synthetic wastewater to obtain 50
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mg/L TOC because the TOC was completely depleted after 9 h according to the
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results of the pre-tests. The mixed liquid samples were collected every 1.5 h through
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the top opening using a 5 mL syringe and then filtered to measure TOC, sulfide, and
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polysulfide concentrations. At each sampling time point, the pH of the SRAFB reactor
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was also measured.
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2.3. Biological sulfur reduction with polysulfide
To determine the effect of sulfide on polysulfide formation and subsequent
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sulfur reduction, batch tests were conducted in duplicate using the cultivated
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sulfur-reducing sludge. The sludge was washed with deoxygenated deionized water
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and then evenly distributed into ten 2.4 L flasks. The sludge concentration in each
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flask was measured as 0.68 g/L volatile suspended solids (VSS). Initial sulfide
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concentrations of 0, 50, 100, 150, and 200 mg S/L in respective flasks were obtained
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by spiking a concentrated sodium sulfide solution. Organic carbon (100 mg/L glucose)
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and 5 g/L sublimed sulfur were added to each flask to provide sufficient carbon and
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sulfur. The flasks were purged with nitrogen gas and then sealed with rubber
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stoppers to maintain anaerobic conditions. The tests were conducted at an ambient
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temperature of approximately 25°C, and the initial pH was controlled at 7.50±0.02.
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The batch tests were conducted for 7 h, during which 5 mL samples from two
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replicates were collected every two hours through a sampling port on the rubber
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stopper to measure sulfide, polysulfide, sulfate, thiosulfate, and TOC. A pH probe
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was fitted in the flask to measure pH variations with time. The sulfide production
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rate (SPR) was calculated according to Equation 3:
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sulfide production rate (mg S/L-h) =
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∆CS 2−
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∆t
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( ∆t , h) in mg S/L.
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2.4. Biological sulfur reduction without polysulfide
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The test conditions were controlled to compare direct sulfur reduction
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without polysulfide and indirect sulfur reduction via polysulfide. FeCl2 was used to
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completely precipitate the dissolved sulfide to prevent polysulfide formation (Ringel
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et al., 1996). A batch test was conducted in duplicate to evaluate sulfur reduction
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without polysulfide by adding 4.7 mM FeCl2 at the beginning of the test to 200 mL
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flasks. The sludge (0.68 g/L) was washed with deoxygenated deionized water three
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times to remove residual sulfide before FeCl2 was added. A control test without FeCl2
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addition was also performed in duplicate. The other test conditions were kept the
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same as those in section 2.3. The batch tests were performed for 9 h, during which
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samples were collected every 3 h to monitor the changes in sulfide, sulfate,
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thiosulfate, and TOC concentrations. The pH values were measured using a pH probe
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fitted to the flask.
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2.5. Chemical analysis
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The TOC, sulfide, sulfate, and thiosulfate concentrations were determined after
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filtration (Millipore, 0.45 µm). The TOC was analyzed using a TOC analyzer (Shimadzu
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TOC-5000A), and the total sulfide (H2S, HS-, and S2-) concentration was determined
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using the methylene blue method (APHA, 2005). The sulfate and thiosulfate 9
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chromatograph (DIONEX-900), and the VSS was measured according to the standard
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methods (APHA, 2005). The pH was measured using a pH meter (Hach, HQ40D). The
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acid volatile sulfide (AVS) deposited in the sludge was measured using the method
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described by Simpson (2001). Elemental sulfur in the sludge samples was extracted
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based on the method described by McGuire and Hamers (2000) and was measured
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using a high-performance liquid chromatograph (HPLC, Shimadzu LC-16, Japan)
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equipped with a Kromasil column (C18, 5μm, 100 Å) and a UV detector at 254 nm.
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Since the length of the Sn2- chain varied from 2 to 11, it was difficult to determine the
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aggregate concentration of polysulfide. In contrast, the zero-valent sulfur atoms in
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polysulfide ions (also called dissolved zero-valent sulfur) can be analyzed using an
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ultraviolet-visible spectrophotometer (UV-6000PC, Metash, Shanghai, China) at a
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wavelength of 285 nm after filtration (Millipore, 0.22 µm) (Kleinjan et al., 2005). Thus,
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the dissolved zero-valent sulfur in this study was defined as an indicator of
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polysulfide. The mixed liquid samples were taken with a 5 mL syringe and
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immediately filtered through disposable 0.22-μm filters. The filtrate was directly
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deposited in a N2-filled glass tube and immediately analyzed with the
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ultraviolet-visible spectrophotometer.
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3. Results
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3.1. Performance of the sulfur-reducing bioreactor
The anaerobic sulfur-reducing reactor was successfully operated with
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sublimated sulfur as its sole source of electron acceptors for 164 days. The SPR
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increased as the HRT of the reactor decreased and reached a maximum of 126 mg
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S/L-h (Fig. 1a). Although sublimed sulfur is nearly insoluble, this rate is significantly
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higher than those reported in previous studies, which achieved SPRs of 24 mg S/L-h
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(Wang et al., 2009), 22 mg S/L-h (Wang et al., 2008), 43 mg S/L-h (Qian et al., 2015),
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45 mg S/L-h (Jiang et al., 2013), and 72 mg S/L-h (Celis-García et al., 2007) using
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sulfidogenic processes based on sulfate/sulfite reduction. More than 80% of the TOC
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was also removed (Fig. S3). In addition, sulfate and thiosulfate were not detected in
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the influent and effluent, indicating that all of the sulfide in the effluent came from
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sulfur reduction.
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As shown in Fig. 1b, the ratio of removed organic carbon to produced sulfide
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(mg C/mg S) was close to the theoretical level of 0.19 mg C/mg S for sulfur reduction,
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as calculated from Equation 4 (Corg means the biodegradable organic carbon). This
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result reveals that biological sulfur reduction prevailed in this reactor.
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Corg + 2S 0 + 2 H 2 O → CO2 + 2 HS − + 2 H +
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3.2. Polysulfide in the sulfur-reducing bioreactor
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(4)
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An increase in dissolved zero-valent sulfur concentration was clearly
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observed after the complete depletion of organic carbon in the sulfur-reducing 11
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maximum of 55 mg S/L, which is much higher than the solubility of sublimated sulfur
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in water (5 µg/L at 25 °C) (Schauder and Müller, 1993). This result reveals that the
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measured dissolved zero-valent sulfur was most likely polysulfide, namely the S0 in
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Sn2-.
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Dissolved zero-valent sulfur was not detected when TOC was present in the
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sulfur-reducing bioreactor, indicating that the rate of dissimilatory polysulfide
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reduction was much higher than the rate of polysulfide formation. This suggests that
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polysulfide formation could be the rate-limiting step for high-rate sulfur reduction.
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According to Equations 1 and 2, HS- production from sulfur reduction can facilitate
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polysulfide formation. We thus hypothesized that indirect sulfur reduction via
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polysulfide would be accelerated by an increase in sulfide concentration resulting
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from sulfur reduction.
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3.3. Sulfur reduction without sulfide initially
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To verify the aforementioned hypothesis, a batch test was performed in the
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absence of sulfide initially (refer to section 2.3). Without sulfide at the beginning of
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the batch test, the hourly SPR was only 4.2 mg S/L-h during the first hour; however,
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it climbed gradually and reached a maximum of 36.4 mg S/L-h during the seventh
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hour, which represents an approximately 8.7 times increase (Fig. 3). Polysulfide was
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detected during the entire period and reached 9.6 mg S/L by the end of the batch
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batch test, or in subsequent batch tests.
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3.4. Sulfur reduction with sulfide initially
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Batch tests were further performed with different concentrations of sulfide
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added (50, 100, 150, and 200 mg S/L) at the start of the tests to assess the feasibility
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of artificially stimulating sulfur reduction.
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Sulfide addition stimulated polysulfide formation at the start of the tests (Fig.
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S4). Thereafter, the polysulfide concentrations decreased with time in all of the
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flasks due to polysulfide reduction. As a result, the increased initial concentration of
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sulfide enhanced sulfur reduction (Fig. 4). At the start of the tests, the highest hourly
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SPR (53.7 mg S/L-h) was achieved with the highest sulfide dosage (200 mg S/L).
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Within the first 3 h, a linear correlation was observed between the average SPR and
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the initial sulfide dosage (Fig. S5a). The organic reduction rates followed a similar
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trend (Fig. S5b) and the organic compounds were completely depleted six and seven
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hours in the tests when the initial dosages were 150 and 200 mg S/L, respectively
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(Fig. S6).
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However, there is a limit to how much sulfide can stimulate sulfur reduction.
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Once the sulfide concentration in the flasks exceeded 250 mg S/L, the stimulation
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effect of sulfide addition on sulfur reduction weakened over time. For instance, the
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batch tests with 150 and 200 mg/L sulfide additions exhibited high hourly SPRs 13
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in the tests with just 50 mg S/L sulfide added, the hourly SPR just kept on increasing,
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similar to the test without sulfide addition. These results reveal that accelerating
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sulfur reduction only applies at sulfide levels below ~250 mg S/L.
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3.5. Sulfur reduction without polysulfide
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To compare the rates of direct and indirect sulfur reduction, a batch test was
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conducted in which polysulfide formation was prevented by adding Fe2+ (refer to
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section 2.4). In the absence of polysulfide, only 0.7 mg S of AVS had accumulated by
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the end of the batch test, which was just 2.4% of the amount of sulfide produced in
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the batch test without polysulfide control (28.6 mg S) (Fig. 5a). This result indicates
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that the rate of sulfur reduction decreased by 97.6% in the absence of polysulfide.
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Moreover, organic consumption in the batch tests with Fe2+ addition (3%) was
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significantly lower than that in the batch tests without (66%) (Fig. 5b).
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4. Discussion
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4.1. Mechanisms of high-rate sulfur reduction
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A high-rate sulfidogenic process was demonstrated in a sulfur-reducing
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bioreactor using elemental sulfur as its sole source of electron acceptors, although
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elemental sulfur is almost insoluble. We found that polysulfide was present in the
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We therefore hypothesized that the soluble polysulfide was the main intermediate
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which first boosted sulfur bioavailability and then accelerated sulfur reduction. Batch
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tests were further conducted to verify this hypothesis, and the results were
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affirmative. Previous studies (Kleinjan et al., 2005, Sigel and Sigel, 2005, Florentino et
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al., 2016) have suggested that an increase in sulfide concentration enhances
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polysulfide formation. Furthermore, polysulfide can be easily reduced to sulfide by
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sulfur reducers (Schauder and Müller, 1993, Florentino et al., 2016, Liang et al.,
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2016), which in turn promotes polysulfide formation through the abiotic reaction
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between sulfur and sulfide at neutral or alkaline conditions (Boyd and Druschel,
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2013). To the best of our knowledge, the present study is the first to demonstrate
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the application of high-rate sulfur reduction via polysulfide formation in wastewater
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treatment.
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The mechanism of high-rate sulfur reduction is the indirect sulfur reduction
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enabled by polysulfide, which occurs through the pathways proposed in Fig. 6. The
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sulfide produced through sulfur reduction promotes polysulfide formation (Equation
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1), which is the rate-limiting step of the indirect sulfur reduction process. Thereafter,
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the sulfur reducers reduce polysulfide to sulfide (Equation 2), which promotes the
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formation of more polysulfide akin to a chain reaction. This chain reaction
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continuously enhances sulfur reduction until it is suppressed due to the toxicity of
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the high sulfide levels. Accordingly, the rate of sulfide production increased
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this study (see Fig. 3).
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4.2. Conditions for high-rate sulfur reduction
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The results also reveal that the bioavailability of elemental sulfur can be
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significantly enhanced by introducing polysulfide as an intermediate to enable
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high-rate indirect sulfur reduction. Thus, the key prerequisites for high-rate sulfur
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reduction are polysulfide formation and reduction.
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Moreover, maintaining neutral or alkaline pH levels is essential for polysulfide
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formation (Florentino et al., 2016). Polysulfide is unstable and easily decomposes to
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form elemental sulfur and sulfide at acidic pH levels (Boyd and Druschel, 2013,
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Florentino et al., 2016). Schauder and Müller (1993) reported that polysulfide can
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barely be detected at pH levels below 6. Therefore the pH level in a high-rate
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sulfur-reducing reactor must be at least 6 and was kept at an average of 6.7 in our
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sulfur-reducing bioreactor (Fig. S7) to maintain suitable conditions for polysulfide
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formation.
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An increase in sulfide concentration in a sulfur-reducing reactor stimulates
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sulfur reduction by facilitating polysulfide formation (Kleinjan et al., 2005, Sigel and
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Sigel, 2005, Florentino et al., 2016). At the start of a batch test, apart from manual
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addition, sulfate reduction may also provide sulfide to promote polysulfide
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formation if sulfate is present in the wastewater. However, too much sulfide (> 250 16
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to the toxicity of unionized hydrogen sulfide to sulfidogenic bacteria. Hydrogen
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sulfide has been found to exert a direct and reversible toxicity effect on SRB (Reis et
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al., 1992) with 100% inhibition of SRB growth at H2S concentrations varying from 477
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to 617 mg/L (Reis et al., 1992, Kolmert et al., 1997, Neculita et al., 2007).
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In addition, preventing polysulfide formation by adding Fe2+ evidently reduced
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the sulfur reduction rate. Although elemental sulfur could be reduced without
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polysulfide, it occurred at a very low rate compared to the rate of indirect sulfur
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reduction. This result suggests that high-rate sulfur reduction cannot be achieved
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without polysulfide. Thus, we propose that a two-stage system consisting of a
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high-rate
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independently would be more suitable for metal-laden wastewater treatment than
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the single-stage system presented by Florentino et al. (2016). In addition, the sulfur
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reduction process can provide sufficient electron donors (sulfide) for subsequent
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autotrophic denitrification in low carbon-to-nitrogen wastewater treatment.
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metal
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5. Conclusions
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This study achieved high-rate sulfur reduction in a sulfur-reducing bioreactor
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and investigated its mechanisms, which are discussed in this paper. The main findings
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are: 17
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•
The high-rate sulfur reduction was achieved using insoluble sublimated sulfur as the sole source of electron acceptors. The highest rate of sulfide production
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attained in the bioreactor was 126 mg S/L-h, which is significantly higher than
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those previously reported for other sulfidogenic processes.
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•
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The high-rate sulfur reduction was attributed to indirect sulfur reduction driven by polysulfide. The sulfide produced through sulfur reduction induced both
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polysulfide formation and reduction to produce more sulfide, thus enabling the
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self-acceleration of sulfur reduction. •
•
Neutral or alkaline conditions, along with a low metal concentration in the influent, are essential for high-rate sulfur reduction.
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The continuous increase in sulfide production rate was observed until the sulfide concentration in the bioreactor exceeded 250 mg/L.
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These findings suggest that high-rate sulfur reduction can be achieved at a low cost
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and represents a promising technology for autotrophic denitrification or the removal
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of heavy metals in wastewater treatment.
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Acknowledgements
The authors acknowledge the support from the National Natural Science
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Foundation of China (51178914 and 51638005), the Guangdong Provincial Science
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and Technology Planning Project (2016A050503041 and 2017B050504003), and the
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Hong Kong Innovation and Technology Commission (ITC-CNERC14EG03).
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ACCEPTED MANUSCRIPT Figure captions Fig. 1. The performance of the sulfur-reducing bioreactor: (a) the average sulfide production and sulfide production rates for the six stages (from stage 1 to stage 6), (b) the C/S ratios for the six stages (the dashed line represents the theoretical C/S
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ratio of 0.19)
Fig. 2. The variations in TOC, sulfide and polysulfide concentrations in the sulfur-reducing bioreactor during the batch test. Organic carbon was added at 9.2 h
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as indicated by the orange arrow.
Fig. 3. (a) The hourly sulfide production rate; and (b) sulfide production and TOC
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removal as a function of time without sulfide addition at the start of the batch tests (sulfur reduction with polysulfide) (the data points are averages of two replicates). Fig. 4. (a) The hourly sulfide production rates and (b) sulfide concentration as a function of time under varying sulfide addition conditions at the start of the batch
replicates).
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tests (sulfur reduction with polysulfide) (the data points are averages of two
Fig. 5. (a) Sulfide production at pH 7.5 and pH 7.5+Fe2+ in the 9 h batch tests (the AVS
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was measured to quantify the sulfide concentration in the batch test with Fe2+ addition); (b) the corresponding TOC consumed in the control tests (the data points
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are averages of two replicates). Fig. 6. The hypothetical pathway of biological elemental sulfur reduction under neutral or alkaline conditions.
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Fig. 1. The performance of the sulfur-reducing bioreactor: (a) the average sulfide
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production and sulfide production rates for the six stages (from stage 1 to stage 6), (b) the C/S ratios for the six stages (the dashed line represents the theoretical C/S
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ratio of 0.19)
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Fig. 2. The variations in TOC, sulfide and polysulfide concentrations in the sulfur-reducing bioreactor during the batch test. Organic carbon was added at 9.2 h
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Fig. 3. (a) The hourly sulfide production rate; and (b) sulfide production and TOC removal as a function of time without sulfide addition at the start of the batch tests
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Fig. 4. (a) The hourly sulfide production rates and (b) sulfide concentration as a function of time under varying sulfide addition conditions at the start of the batch
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Fig. 5. (a) Sulfide production at pH 7.5 and pH 7.5+Fe2+ in the 9 h batch tests (the AVS
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Fig. 6. The hypothetical pathway of biological elemental sulfur reduction under
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ACCEPTED MANUSCRIPT Highlights High-rate sulfide production was achieved in a sulfur-reducing bioreactor
•
Self-accelerating sulfur reduction was observed
•
The mechanism of self-accelerating sulfur via polysulfide was characterized
•
High sulfide concentration (>250 mg S/L) weakened the self-acceleration of sulfur
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reduction