Arsenic removal by periphytic biofilm and its application combined with biochar

Accepted Manuscript Arsenic removal by periphytic biofilm and its application combined with biochar Ningyuan Zhu, Jianhong Zhang, Jun Tang, Yan Zhu, Yonghong Wu PII: DOI: Reference:

S0960-8524(17)31113-6 http://dx.doi.org/10.1016/j.biortech.2017.07.026 BITE 18446

To appear in:

Bioresource Technology

Received Date: Revised Date: Accepted Date:

16 May 2017 4 July 2017 6 July 2017

Please cite this article as: Zhu, N., Zhang, J., Tang, J., Zhu, Y., Wu, Y., Arsenic removal by periphytic biofilm and its application combined with biochar, Bioresource Technology (2017), doi: http://dx.doi.org/10.1016/j.biortech. 2017.07.026

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Arsenic removal by periphytic biofilm and its application combined with biochar

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Ningyuan Zhua,c, Jianhong Zhangb, Jun Tanga,c, Yan Zhua,c, Yonghong Wua*

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a

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Chinese Academy of Sciences, 71 East Beijing Road, Nanjing 210008, China

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b

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Corporation, Beijing 100048, China

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c

State Key Laboratory of Soil and Sustainable Agriculture, Institute of Soil Sciences,

Resources & Environment Business Dept., International Engineering Consulting

Graduate School of the Chinese Academy of Sciences, Beijing 100039, China

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*Corresponding author:

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Dr. Yonghong Wu

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71, East Beijing Road, Nanjing 210008 China

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Tel: (+86)-25-8688 1330

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Fax: (+86)-25-8688 1000

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E-mail: [email protected] (Y. Wu).

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Abstract:

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A biochar and periphyton-based system (BPS) comprising of a biochar column and

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a periphyton bioreactor was designed to avoid the toxicity issue associated with

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removing As(III) from wastewater. Results showed that the periphyton can grow when

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As(III) is less than 5.0 mg L-1. The BPS obtained a high As(III) removal rate

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(~90.2-95.4%) at flow rate = 1.0 mL min-1 and initial concentration of As(III) = 2.0 mg

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L-1. About 60% of the As(III) was pre-treated (adsorbed) in the biochar column and the

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removal of the remaining As(III) was attributed to the periphyton bioreactor. The As(III)

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removal process by periphytic biofilm in the initial stage fits a pseudo-second-kinetic

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model. The calcite in the periphytic biofilm surfaces and the -OH and -C=O groups

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were responsible for the As(III) removal. This study indicates the feasibility of the BPS

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for As(III) removal in practice.

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Key words: Periphytic biofilm, As(III) removal, Biochar

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1．Introduction

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Arsenic (As), a ubiquitous element found in the Earth’s crust, has abroad

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applications in the semiconductor industry, alloy manufacturing and agricultural

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production because of its excellent physicochemical and electrochemical properties

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(Chowdhury et al., 2017; Wong et al., 2017). Excessive amounts of As(III) derived from

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natural processes and anthropogenic activities now occur in many environments and

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ecosystems, such as paddy fields (Chowdhury et al., 2017). Around 20 million people

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are affected or threatened by exposure to unsafe As levels in China (Xie et al., 2017),

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and West Bengal where As levels in groundwater are higher than the World Health

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Organization (WHO) guidelines (10 μg L-1) (Gill & O'Farrell, 2015; Shrivastava et al.,

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2017). Inorganic species (arsenate (As(V) at high redox potential value) and arsenite

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(As(III) at low redox potential value)) are the predominant forms of arsenic in aquatic

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environments. As(III), which is more toxic, soluble, and mobile than As(V), can make

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up 67-99% of the total arsenic in groundwater (Giles et al., 2011).

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To combat these high As levels, a variety of As(III) treatment methods, such as

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adsorption (Lata & Samadder, 2016) and microbial removal (Sun et al., 2016), have

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been developed to remove or detoxify arsenic. These promising methods are easy to

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operate, low-cost and environmentally-benign with significant progress being made in

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the last two decades (Hayat et al., 2017; Ungureanu et al., 2015). There are, however,

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some limitations that need to be considered when these current technologies are

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employed in practice, particularly industrial scale applications (Kowalski, 2014).

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Biochar, especially chemically modified biochar such as MnOx-loaded biochar, bismuth

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impregnated biochar and biochar colloid, has a high adsorption capacity for heavy metal

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(e.g. Cu, As, Cd and Cr) contaminants in aqueous solutions (Qian et al., 2016; Song et

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al., 2014; Zhu et al., 2016). However, complicated environmental factors such as the

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existence of competitive ions and the risk of transportation in soil and groundwater

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environments by water flow suppresses their application. Microorganisms possess the

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ability to detoxify arsenic. The surface adsorption of arsenic, oxidation or reduction of

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inorganic arsenic, methylation and demethylation of arsenic and chelation to

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intracellular cysteine-rich polypeptides are the widely acknowledged mechanisms of

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arsenic detoxification. The detoxification processes, however, are affected by forms of

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arsenic species, the composition of microorganisms and the potential interactions

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between microorganisms and nutrients. For example, pure strains of microorganisms,

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such as Dunaliella tertiolecta and MLH-1 a γ-proteobacterium, were poisoned when the

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As concentration was > 1 mg L-1 (Duncan et al., 2013; Oremland et al., 2004).

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Periphytic biofilm is composed of heterotrophic and phototrophic microorganisms,

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embedded in a self-produced mucilage matrix of extracellular polymeric substances

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(EPS) (Wu et al., 2012). The EPS has great potential for sorption of heavy metal ions

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(Wu et al., 2014) and periphyton is an important sink for As (Lopez et al., 2016). The

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first step is the interaction between the cell surface and As based on adsorption followed

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by the penetration of As ions into the cell membrane and then into the cells (Bahar et al.,

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2013). As a result of its affinity for As, periphytic biofilm has been employed in this

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study to remove As(III) from wastewater. In consideration of the possible toxic effects

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of high concentration As(III) on periphytic biofilm, the wastewater was pretreated by an

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adsorption process using biochar.

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The practical utility of the combined physical adsorption-biosorption process has

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not been addressed for As(III) treatment. In the study by Upadhyaya (2010), a

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biologically active carbon (BAC) reactor system was operated for simultaneous As(V)

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and nitrate removal (Upadhyaya et al., 2010). However, to obtain the ideal As(V)

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removal efficiency, acetic acid and Fe2+ should be supplemented as the sole electron

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donor and dissolved oxygen needs to be removed to below 1 mg L-1. On one hand, this

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coupled process decreases the need for a high adsorption dosage input to acquire a high

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removal efficiency for the wastewater treatment. On the other hand, with the

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pretreatment by the adsorption process, this system may possess a high stability for

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toxic heavy metal ion treatment. The objectives of this study were to (i) investigate the

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growth of periphytic biofilm in the presence of As(III), (ii) study the As(III) removal by

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periphytic biofilm, (iii) design a biochar and periphyton-based system (BPS) to remove

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As in practice, and (iv) explore the removal mechanisms of As(III) removal by the

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integrative system. The study is expected to provide a useful approach to As(III)

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removal in practice and provide insight into the mechanisms of As(III) removal by

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periphytic biofilm.

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2. Materials and methods

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2.1 As(III) removal experiments

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Bismuth impregnated biochar was prepared following the method reported by (Zhu

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et al., 2016). The impregnation of bismuth improved the specific surface area of the

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adsorbent through helping to create more micropores. Thus, the prepared bismuth

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impregnated biochar had a 190.4 m2 g-1 (Brunauer-Emmertt-Teller) surface area and 2

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nm average pore diameter. Periphytic biofilms were collected from our biofilm culture

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system described in another of our previous studies (Wan et al., 2016).

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To reduce the disturbance of exogenous ions, a simulated wastewater was used in

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the As(III) removal experiment. The components were as follows: 62 ± 0.6 mg L-1 NO3-,

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4.8 ± 0.3 mg L-1 PO43-, 9.2 ± 0.2 mg L-1 HCO3- , 14.4 ± 0.3 mg L-1 SO42- and 35.5 ± 0.5

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mg L-1 Cl-. Batch As(III) removal experiments by periphytic biofilm were conducted as

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follows: 0.25 g (w/w) periphytic biofilm was added into a 250 mL flask with the

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stimulated wastewater. Then, different doses of sodium arsenite were added, leading to

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initial As(III) concentrations of 0, 2.0, 5.0, 10.0 and 15.0 mg L-1. On day 0, 5, 7, and 10,

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each treatment was sampled to analyze the As(III) concentration. All the biofilms were

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collected, centrifuged (3500 r min-1), and weighed at the end of the experiment (day 10).

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All the conical flasks used were sealed using kraft paper and put into a constant

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temperature incubator in a standard light-dark cycle of 12/12 h at 28 ± 1 ˚C in the

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daytime and 20 ± 1 ˚C at night. The experiment was performed in triplicate.

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2.2 As(III) removal mechanism experiments

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Two separate experiments were used to explore the As(III) removal mechanisms.

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The first was an adsorption experiment to determine the adsorption potential of

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periphytic biofilm samples at the beginning of the experiment. The experimental

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process was briefly induced as follows: 1.0 g (w/w) periphytic biofilm was added to 500

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mL As(III) solution in a 1000 mL flask with initial As(III) concentrations set at 5 and 10

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mg L-1. The sorption amounts of As(III) were investigated at different time intervals (0,

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5, 10, 45, 90, 120 and 360 min). The flasks were agitated at 200 rpm in an orbital shaker

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(Incu-Shaker LR, USA) at 25 ˚C.

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The second experiment was to investigate how the periphytic biofilm “reacted”

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with the As(III) during prolonged contact. The experiment followed the previous

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adsorption experiment. Briefly, after 360 mins, the periphytic biofilm cultured in 5.0 mg

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L-1 As(III) was cultured for an additional 14 days. The culture conditions were the same

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as in the As(III) removal experiment. On day 15 the periphytic biofilms were collected

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and freeze dried in a vacuum freeze dryer (LyoQuest-55, Spain). Both mechanism

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exploration experiments were performed in triplicate.

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2.3 As(III) removal in practice experiment

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To remove As(III) in practice, a biochar and periphyton-based system (BPS) was

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designed (Figure 1). The system included two main parts, bismuth modified biochar and

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periphyton bioreactor. The volume of the biochar column reactor was 565 mL (diameter:

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6 cm, height: 20 cm), containing bismuth modified biochar. The dimensions of the spiral

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periphyton bioreactor made of PE pipe were 21.9 m total length and 2.0 mm diameter.

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The detailed parameters of the bioreactor are similar to our previous study (Shangguan

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et al., 2015). The BPS bioreactor also contained two other parts: a clarification tank

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(volume: 1000 mL, depth: 10 cm, length: 10 cm) and an influent tank (volume: 1000

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mL, diameter: 10 cm).

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The biochar column reactor was filled with 20 g of bismuth impregnated biochar

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(height: 10 cm) and quartz sand at both the top and bottom (depth: 3 cm for both) of the

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pillar. The biochar and sand were all washed thoroughly three times with deionized

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water before packing into the column reactor. The periphytic biofilm was inoculated by

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pumping water containing microbial sources into the bioreactor at 28 ± 1 °C under light

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irradiation. After 35 days, periphytic biofilms formed on the inside surface of the PE

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pipe. The system was then used to determine As(III) removal.

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The wastewater was collected from Xuanwu Lake, East China. The water had a pH

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7.8 ± 0.2, dissolved oxygen 4.2 ± 0.1, chemical oxygen demand 38.6 ± 0.3, NO3- -N

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15.3 ± 0.4 mg L-1, NH4+ -N 0.27 ± 0.01 mg L-1, total phosphorus (TP) 0.16 ± 0.02 mg

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L-1, and dissolved phosphorus (DP) 0.12 ± 0.01 mg L-1. The inclusion of periphytic

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biofilm was conducted according to our previous study (Shangguan et al., 2015). Briefly,

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the water with the added periphytic biofilm from our biofilm culture system was

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continuously pumped into the PE pipe for 35 days. This reactor worked in a standard

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light-dark cycle of 12/12 h at 28 ± 1 ˚C in the daytime and 20 ± 1 ˚C at night. The flow

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rate was controlled at 1 mL min-1. To facilitate comparisons with the results of the static

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experiments above, the initial As(III) concentrations in the wastewater were adjusted to

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2.0 and 5.0 mg L-1 by dilution using water or directly added sodium arsenite. To support

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the periphytic biofilm growth, Woods Hole (WC) media were also added into the

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wastewater (v/v = 11 mL/L). The medium contained NaNO3 (85.10 g L-1), CaCl2·2H2O

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(36.76 g L-1), MgSO4·7H2O (36.97 g L-1), NaHCO3 (12.6 g L-1), Na2SiO3·9H2O (28.42

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g L-1), K2HPO4 (8.71 g L-1), H3BO3 (24 g L-1), trace elements solution Na2EDTA·2H2O

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(4.36 g L-1), FeCl3·6H2O (3.15 g L-1), CuSO4·5H2O (2.50 mg L-1), ZnSO4·7H2O (22 mg

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L-1), CoCl2·6H2O (10 mg L-1), MnCl2·4H2O (180.00 mg L-1), Na2MoO4·2H2O (6.30 mg

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L-1), Na3VO4 (18.00 mg L-1), vitamin B (135.00 mg L-1), thiamine (335.00 mg L-1), and

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biotin (25.00 g L-1). The influent from the stock tank (1) enters the biochar column (3)

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via the pump (2), enters into the periphytic biofilm reactor (4), and then discharges

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(Figure 1). Effluents A (effluent of the biochar column reactor) and B (the final effluent

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of the BPS reactor) were sampled to analyze arsenic concentrations. The detailed initial

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concentrations of the main components of the wastewater and operation parameters are

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presented in Table 1.

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2.4 Analytic methods and statistics

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The pseudo-first-order kinetic model and pseudo-second-order kinetic models were

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selected to investigate the experimental data to further determine the adsorption process.

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These two model are described as follows:

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(1)

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(2) 8

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Where t is the sampling time (min), qt is the adsorption amount at a given time (mg

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g-1), qe is the adsorption amount at equilibrium (mg g-1), and k1 (1.0 min-1) and k2 (mg

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min-1) are the rate constants of the two adsorption models, respectively. The

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pseudo-first-order kinetic model hypothesized that the adsorption rate varies directly

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with the concentrations of the reactants while the pseudo-second-order kinetic model

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assumed the relationship between the adsorption rate and the concentrations of the

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adsorbate is linear (Rodrigues & Silva, 2016; Simonin, 2016).

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The As(III) concentrations were determined by High Performance Liquid

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Chromatography-Inductively

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7700x-JP12502215, Agilent Technologies, USA). Surface functional groups of

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periphytic biofilm were detected by Fourier transform infrared spectroscopy (FTIR)

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(Nicolet IS10, Thermo Electron Co, USA) at a spectral range of 4000 cm-1 to 400 cm-1

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with a resolution of 8 cm-1. X-ray diffraction (XRD) patterns of periphytic biofilm were

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measured by a Siemens D-501 diffractometer with Ni filter and graphite

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monochromator (Ultima IV- JD2643W, RIGAKU, Japan). After the periphytic biofilms

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were collected and freeze dried on day 15, the dry periphytic biofilms were milled and

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sized into powder. Then, 100 mg periphytic biofilm powder was tiled on a silicon slice

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for XRD analysis. For FTIR analysis, samples were prepared by mixing 1 mg of

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periphytic biofilm with 100 mg of spectroscopy grade KBr.

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Coupled

Plasma-Mass

Spectrum

(HPLC-ICP-MS,

Statistically significant differences between the treatment and the control were

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evaluated using One-way ANVOA. The α was set at 0.05 for all analyses.

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3. Results and discussion

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3.1 As(III) removal from simulated wastewater

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3.1.1 Adsorption of As(III) by periphytic biofilm at the initial stage

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Periphytic biofilm possess a strong ability to remove heavy metal ions from

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wastewater through adsorption-desorption processes (Yang et al., 2016). Adsorption

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kinetic experiments were conducted to investigate the capacity of As(III) entrapment by

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periphytic biofilm. Figure 2 shows the change in As(III) concentration in wastewater

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throughout the initial stage (360 mins). When the As(III) concentration was initially

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10.0 mg L-1, the concentration decreased rapidly after the first 30 min reaching an As(III)

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concentration of 8.2 mg L-1. The concentration of As(III) then increased and approached

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the initial As(III) concentration at 360 min. This desorption process might be due to the

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microorganisms in the periphytic biofilms being poisoned with high-intensity As(III).

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Interestingly, the concentration of As(III) also declined rapidly in the initial 30 min

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for the 5.0 mg L-1 As(III) treatment. The As(III) continued to decrease to 3.2 mg L-1 and

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the equilibrium time was reached around 120 min. These results indicated that

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periphytic biofilm possess considerable capacity for As(III) adsorption when the As(III)

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has not accumulated to a toxic level (<10 mg L-1). Periphytic biofilm may also remove

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As by surface adsorption through its porous structure (Wu et al., 2014).

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To explore the process of As(III) trapped by periphytic biofilm at an initial

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concentration of 5.0 mg L-1 As(III), pseudo-first-order and pseudo-second-order kinetic

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models were employed to fit the As removal in the first 360 mins. The

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pseudo-second-kinetic model fitted the As(III) removal process well (k2 = 0.18, qe =

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0.89 and R2 = 0.999), indicating that chemisorption dominated the As(III) removal by

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periphytic biofilm within

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consistent with the adsorption of Cu by periphytic biofilm (Yang et al., 2016),

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suggesting periphytic biofilm has the potential to entrap As from wastewater.

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3.1.2 Removal of As(III) by periphytic biofilm

the initial 360 mins (Takaya et al., 2016). This result was

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Arsenic removal by periphytic biofilm decreased with increasing concentration of

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As(III) (Figure 2b). The As(III) removal efficiency in the 2.0 and 5.0 mg L-1 treatments

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were 95.8% and 60.4%, respectively, on day 14 while less than 20% of As(III) was

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removed in the 10 and 15 mg L-1 treatments. This further implies that the periphytic

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biofilm growth might be negatively affected by high concentration As (> 10 mg L-1).

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The average biomass of periphytic biofilm in the control was 2.0 g while the

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average biomasses in the treatments were 1.8, 1.7, 1.2 and 0.8 g for 2.0, 5.0, 10.0 and

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15.0 mg L-1 As(III) treatments, respectively. There was no significant difference

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between the low-intensity As(III) treatments (2.0 and 5.0 mg L-1) and the control (p >

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0.05). A high concentration of As(III) (10.0 and 15.0 mg L-1 As(III) treatments)

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significantly inhibited the growth of periphytic biofilm (p < 0.05), which offers a good

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explanation for the quick desorption at the initial As(III) concentration of 10.0 mg L-1.

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Microorganisms such as green alga, Chlorella vulgaris, and phytoplankton, Chlorella

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salina, possess the ability to grow in the presence of toxic As ions (Baker &

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Wallschläger, 2016; Karadjova et al., 2008). However, these kinds of pure strains only

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grew at a considerably low As concentration level. Periphytic biofilm proved to be

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capable of growth in the presence of 2.0 and 5.0 mg L-1 As ions. This could be attributed

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to its diverse community composition and porous structure including EPS (Flemming &

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Wingender, 2010; Tlili et al., 2017). The results also showed that the use of periphytic

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biofilm on its own is unfeasible for treating wastewater with high As(III) concentration.

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This suggests that pretreatment is needed before processing high-intensity As(III)

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removal using periphytic biofilm.

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3.2 As(III) removal in practice

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3.2.1 Performance of the biochar column reactor

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The average As(III) removal rates within 72 h for 2.0 mg L-1 As(III) and 36 h for

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5.0 mg L-1 As(III) were about 60.1% and 39.6%, respectively, when the influent velocity

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was 1 mL min-1. Breakthrough curves (represented by Ct/C0 versus breakthrough time)

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are used for indicating adsorption efficiency with time (Xu et al., 2013). The

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breakthrough curves were affected by the initial As(III) concentrations (Figure 3a),

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occurring at 72 h and 36 h for the 2.0 and 5.0 mg L-1 As(III) treatments, respectively.

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This is because the higher influent concentrations generated higher driving forces for

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mass transfer and larger concentration gradients, thereby easily reaching saturation and

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breakthrough time (Abdolali et al., 2017; Baral et al., 2009). The enhancement of As(III)

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removal was in inverse proportion to the influent flow rates with different flow rates

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changing the shape of the breakthrough curves (Figure 3b). Faster saturation and an

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earlier breakthrough time were observed at higher flow rates. The mass transfer

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fundamental could explain these results (Russo et al., 2016). Achieving the high

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removal efficiency of the single adsorption technology requires a high dosage input of

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adsorbent because of the existence of competitive ions and the effect of temperature and

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pH in the real wastewater treatment

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the disturbances, we designed this type of biochar column where the wastewater

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effluent from the biochar system is subsequently treated by the periphytic bioreactor.

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3.2.2 Performance of the BPS

(Silvetti et al., 2017). Considering the costs and

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The running of the BPS was divided into six stages. Stage 1 was to stabilize the

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BPS and no data are presented. Figure 4 summarizes the As(III) concentrations in the

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influent and effluent of the BPS during stages 2–6. The influence of initial As(III)

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concentration on As removal was investigated during stage 2 and 3 (36-54 d) at influent

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flow rate of 1.0 mL min-1. During stage 2, the As(III) removal rate increased from 81%

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to 95% (36-39 d) when the effluent As(III) concentration was 2.0 mg L-1. On day 39-43,

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the BPS As(III) removal rate decreased dramatically. This could be attributed to the

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saturation of the bismuth activated carbon. When the biochar column reactor was

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renewed, the removal rate increased immediately. The initial concentration of As(III)

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was then adjusted to 5 mg L-1 on day 51 which led to a decreasing As(III) removal rate.

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The total removal efficiency of As(III) decreased to 60%, which might be associated

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with the adsorption saturation rate of biochar and periphytic biofilm. Indeed, both the

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lab experiment in this study and other studies showed that increasing initial

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concentrations of As(III) produce a corresponding decrease in As(III)removal efficiency

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(Wang et al., 2013).

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Experiments were then performed to investigate the effect of flow rate (ranging

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from 0.5 to 2.0 mL min-1) on As(III) removal in stages 4-6 (55-77 d). During stage 4

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(54-60 d), the effluent As(III) was adjusted to 2.0 mg L-1 and the flow rate adjusted to

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0.5 mL min-1, which led to an increased As(III) removal rate. Then, the flow rate was

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increased to 2.0 mL min-1 in stage 5 (60-64 d), after which the As(III) removal rate

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decreased. A low flow rate resulted in the formation of a porous periphytic biofilm

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while biofilm formed a dense structure at a high flow rate (Wang et al., 2011). A higher

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flow rate means a shorter hydraulic retention time between the target As and the biochar

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particles (Russo et al., 2016; Sun et al., 2017) and the periphytic biofilm in the PE pipe

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in light of the increasing turbidity of flow derived from the increased flow tare. The

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shorter hydraulic retention time between the dense structure of periphytic biofilm and

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As resulted in the decrease of removal efficiency. The 0.5 mL min-1 flow rate was too

297

beneficial to the growth of periphytic biofilm in the spiral PE pipe (in the periphytic

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biofilm reactor) resulting in the pipe clogging on day 60. The BPS reactor was operated

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with a 2.0 mg L-1 As(III) concentration and 1.0 mL min-1 flow rate in stage 6 (65-77 d),

300

achieving stable As(III) removal rates (90.2-95.4%).

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In comparison with the studies of (Tang et al., 2017) and (Qi et al., 2015), the BPS

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system achieved a high As(III) removal efficiency, indicating cost and efficiency

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advantages for its application. The removal efficiency was also higher than the

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performance of a biofilter based on As(III)-oxidizing bacteria (Alcaligenes,

305

Pseudomonas) (Yang et al., 2014). In addition, the biofilter required the existence of

306

iron and manganese with As in the groundwater. The change in As forms during the

307

wastewater treatment process, however, needs to be studied due to the effects of

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different forms of As on the natural environment. Furthermore, operational conditions

309

and parameters should be optimized for long-term operation.

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3.3 Mechanism of As(III) removal by periphytic biofilm

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The XRD spectra of periphytic biofilm in the presence and absence of As(III) were

312

investigated. Two diffraction peaks were observed at the biofilm surfaces in the absence

313

of As(III) at 2θ = 29.405 and 39.401. These peaks correspond to the 104 and 113 planes,

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which indicate calcite structures (Wang et al., 2014). There were no peaks observed at

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the periphytic biofilm surfaces in the presence of As(III). These results imply that

316

calcite might combine with As(III), leading to the entrapment of As(III) by periphytic

317

biofilm.

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The functional groups on the surfaces of periphytic biofilms in the presence and

319

absence of As(III) were also investigated. The broad and strong bands at 3439 cm-1 and

320

3417 cm-1 represent the vibration of hydroxyl groups (-OH) (Cebi et al., 2017). The

321

bands at 1413 and 1386 cm-1 represent symmetric and asymmetric stretching vibration

322

of carboxyl (-C=O) groups (Zhao et al., 2013). The peaks at 531, 550, 676 and 707 cm-1

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represent As-OH or As-O stretching vibration (Lata & Samadder, 2016; Liu et al., 2015).

324

The peaks at 1060 and 1030 cm-1 can be attributed to the existence of CO32- (Kim et al.,

325

2017).

326

Two wavenumbers of the periphytic biofilm surfaces were studied further,

327

4000-1300 cm-1 and 1300-400 cm-1. In the wavenumber from 4000-1300 cm-1 there

328

were no new peaks on the biofilm surface after the presence of As(III). The peak

329

intensity and positions shifted, from 3439 cm-1 to 3419 cm-1 for -OH groups and from

330

1413 to 1386 cm-1 for -C=O groups. Accordingly, the -OH and -C=O groups might be

331

involved in the biosorption of As(III) by periphytic biofilm.

332

The peaks changed dramatically in wavenumbers ranging from 1300-400 cm-1. The

333

new peaks at 531, 550, 676, 707 and 1060 cm-1 were detected after the As(III) treatment,

334

and represent As-OH or As-O stretching vibrations (Lata & Samadder, 2016; Liu et al.,

335

2015) and CO32- (Kim et al., 2017). These results further imply that calcite on the

336

periphytic biofilm surfaces played an important role in entrapping As(III), and further

337

indicates that both -OH and -C=O groups were associated with the biosorption of As(III)

338

by periphytic biofilm.

339

3.4 Practical implications

340

As pollution affects millions of people globally, especially in developing countries

341

such as Bangladesh, India, North Africa and China (Mirza et al., 2010; Sun et al., 2015).

342

This work provided an innovative and effective As(III) treatment method by combining

343

physical adsorption and biosorption processes. Many modifications are needed to

344

improve the BPS and produce an actual realistic filtration system. Firstly, the scale of

345

the BPS should be increased. Secondly, the total As removal efficiency must be

346

improved to meet the requirements of the World Health Organization drinking water

15

347

guidelines of less than 10 μg L-1. Third, the regeneration of bismuth impregnated

348

biochar should be further tested. Then, the periphytic biofilm should be properly treated

349

and/or disposed of after As treatment. Finally, previous studies report that As toxicity

350

inhibited the absorption of phosphate nutrients by periphyton biofilm while phosphate

351

concentration could impact on As toxicity (Rodriguez Castro et al., 2015). The

352

consumption of phosphate and uptake of As by the biofilm matrix, led to As stress

353

becoming more and more serious to the periphyton biofilm. Thus, maintaining a high

354

ratio of phosphorus to As in wastewater may contribute to the As removal process.

355

4. Conclusion

356

The biochar and periphyton-based system (BPS) resisted As(III) toxicity when the

357

As(III) concentration was lower than 5.0 mg L-1 and was able to remove As(III) from

358

wastewater. In the initial stage, periphytic biofilm presented a high ability to entrap

359

As(III) by biosorption. This process fit the pseudo-second kinetic model well. The BPS

360

was capable of effectively removing As(III) at initial As concentrations of 2.0 and 5.0

361

mg L-1. The calcite, -OH and -C=O groups on the periphytic biofilm surfaces played an

362

important role in As(III) entrapment by periphytic biofilm. Results indicate that the use

363

of BPS is feasible for As(III) removal and avoiding microorganism poisoning.

364

Acknowledgments

365

This work was supported by the National Natural Science Foundation of China

366

(41422111), the State Key Development Program for Basic Research of China

367

(2015CB158200) and the Natural Science Foundation of Jiangsu Province, China

368

(BK20150066). This work was also supported by the Youth Innovation Promotion

369

Association, Chinese Academy of Sciences (2014269).

370 371 16

372 373 374 375 376 377 378 379 380 381 382 383 384 385 386 387 388 389 390 391 392 393 394 395 396 397 398 399 400 401 402 403 404 405 406 407 408 409 410 411 412 413 414

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530 531 532 533

Figure 1 Experimental apparatus (stock tank (1), via pump (2), biochar column (3), and periphytic biofilm reactor (4)). Figure 2 (a) The adsorption kinetic of As(III) with different initial concentrations by periphytic biofilm and (b) the As(III) removal rates by periphytic biofilm.

534

Figure 3 (a) The effects of initial concentrations of As(III) on As(III) removal, and

535

(b) the effects of flow rates on As(III) removal. Here, Ct was the effluent concentration

536

of As(III) at time t and C0 was the initial concentration of As(III).

537 538

Figure 4 The As(III) removal efficiency of the biochar and periphyton-based system (BPS).

539

21

540 541 542

Figure. 1

22

543 544 545 546

Figure. 2

23

547

548 549 550 551

Figure. 3

24

552 553 554

Figure. 4

25

555

Table 1 The operational parameters of the biochar and periphyton-based system (BPS). Operating

Stage 1

Stage 2

Stage 3

Stage 4

Stage

dates

(1-35 d)

(36-51 d)

(51-54 d)

(55-60 d)

(60-64 d)

(65-77 d)

0

2.0

5.0

2.0

2.0

2.0

1.0

1.0

1.0

0.5

2.0

1.0

-1

As(III) (mg L ) -1

Flow rate (ml min )

556 557

26

5

Stage 6

558 559

27

Highlights

560 561



Periphytic biofilm could remove As(III) from wastewater.

562



Calcite and the -OH and -C=O groups in biofilm facilitate As(III) removal.

563



The BPS achieved stable and effective removal of As(III).

564



The As(III) adsorption process by biofilm obeys pseudo-second-kinetic model.

565 566

28