Removal of Nitrogen and Phosphorus from Wastewater by Modified Pyrite in a Sequencing Batch Reactor (SBR)

: The removal efficiency of nitrogen and phosphorus is challenging in the conventional biological nitrogen and phosphorus removal processes. In this study, the modified pyrite was used as the fillings of sequencing batch reactor (SBR) in order to improve the efficiencies of nitrogen and phosphorus removal from wastewater. The results showed that SBR with the modified pyrite could significantly improve the removal efficiencies of nitrogen and phosphorus when compared with that in SBR without fillings (control SBR). The average influent ammonia nitrogen (NH 4 + -N) and total phosphorus (TP) were 6.96±0.17 mg L -1 and 6.94±0.01 mg L -1 , respectively. The average NH 4+ -N and TP removals of modified pyrite constructed SBR were 49.65±19.49% with 3.54±1.31 mg L -1 of average effluent NH 4+ -N and 76.20±6.55% with 1.84±0.46 mg L -1 of average effluent TP, respectively. While the average NH 4+ -N and TP removal efficiencies of control SBR were only 34.76±11.28% and 56.28±0.11%. The mechanisms of the SBR with enhanced simultaneous nitrogen and phosphorus removals might be anaerobic and aerobic oxidations of modified pyrite, and phosphorus retained in the SBR of modified pyrite was mostly in the form of Fe-bound-P.


Introduction
Nitrogen (N) and phosphorus (P) discharge from wastewater to natural water are the major causes of eutrophication [1]. It is critical for the removals of N and P before wastewater discharge in order to reduce eutrophication [2]. Conventional biological N and P removal processes have been proven to be feasible, while the conventional biological techniques have some design and operational drawbacks, such as high costs and low removal rate and efficiency [3]. Therefore, it is necessary to develop an efficient and cost-effective process that can remove N and P from wastewater.
To date, various configurations of reactors have been developed to remove the N and P from wastewater, including sequencing batch reactors (SBRs), attached growth biofilm reactors, moving bed biofilm reactors (MBBRs) and upflow granular sludge reactors [4]. Among them, SBR is widely used for the treatment of N and P-rich wastewater. However, the operating strategies are crucial to maintaining the stable performance of SBR process and the removal efficiency of N and P in conventional SBR need to be further improvement [5]. Therefore, in order to enhance the N and P removal in SBR, the fillings promoting the removal of N and P should be selected.
Pyrite is the most abundant and widespread mineral of authigenic iron minerals and widely applied for wastewater treatment [6]. Autotrophic denitrifiers also can use pyrite as the electron donor to reduce NO3to N2 and metabolites are ferrous ion (Fe 2+ ), ferric ion (Fe 3+ ) and ferric hydroxide, which are widely used to remove phosphate from wastewater [7]. Therefore, pyrite can be used as the fillings in SBR to simultaneously enhance simultaneous N and P removal from wastewater. Ge et al. (2019) reported that natural pyrite in the wetland could enhance simultaneous nitrogen and phosphorus removal [8]. Modified pyrite, which is called colloidal pyrite, is one of cryptocrystalline pyrite with nanometer and submicrometer-sized crystals, and has a higher chemical activity than natural pyrite due to its special structure [9]. According to the best of our knowledge, few SBRs with modified pyrite as the fillings have been studied.
SBRs were constructed with the modified pyrite as the fillings in this study. The aims of this study are to evaluate the feasibility and performance of using modified pyrite as the fillings in the SBR for wastewater treatment on the simultaneous N and P removal.

Experimental set-up
Two SBRs were constructed with an internal diameter of 14 cm and a height of 50 cm, forming 4 L working volume. The SBR was operated in a 8-h cycle under anoxic and aerobic condition sequentially. Each cycle was consisted of 1-h anoxic period and 5-h aerobic period, following 40-min settling, 10-min effluent withdrawal and 1-h idle period. A total of 2 L synthetic wastewater was pumped into the SBR in the first 10 mins of a cycle. Then the reactor was completely mixed at about 220 rpm by a mechanical stirrer. Air was supplied with a gas flow rotameter during the aerobic period. One of reactors was added about 6 g modified pyrite and the other was as the control group. The two reactors were inoculated with aerobic sludge of a domestic wastewater treatment plant of Hefei, China at an initial concentration of mixed liquor suspended solids (MLSS) of about 4.5 g L -1 .

Fillings
Natural pyrite was obtained from Tongling, Anhui province, China. Before use, the natural pyrite was washed in 10% HCl for 2 h to remove any potentially foreign compound from the pyrite and then was washed 3-4 times with ultra pure water. After that, the dried pyrite was placed in a muffle furnace ini-tially at room temperature and was heated to 650°C in flowing nitrogen (N2). The final heating temperature was maintained for 5 mins. After cooling to room temperature, the modified pyrite was immediately sealed and preserved. X-ray diffraction (XRD) spectra showed that it was one of cryptocrystallinee pyrite.

Analytical methods
Samples from the two SBR were filtered through a 0.45 µm membrane and stored at 4°C before analysis.

Nitrogen removal
The variations of N concentration in the two SBRs during the experiment period were shown in Fig 1. The results showed that the NH4 + -N concentration reduced from 6.84±0.27 mg L -1 in influent to 4.48±0.87 mg L -1 in effluent and 34.76±11.28% of NH4 + -N removal efficiency was obtained in control SBR (Fig. 1A). However, in the SBR with the fillings of modified pyrite, the NH4 + -N concentration reduced from 6.96±0.17 mg L -1 in influent to 3.54±1.31 mg L -1 in effluent (Fig. 1B). The NH4 + -N removal efficiency was up to 49.65±19.49%, which was higher than that in the control SBR ( Table 1). Kong et al. (2015) reported that the integrated twostage soil infiltration bioreactor incorporated with pyrite-based denitrification demonstrated good removal performance of NH4 + -N [10]. Ge et al. (2019) suggested that the natural pyrite had no negative effects in the removal of NH4 + -N, but enhanced total nitrogen in constructed wetland [8]. Meanwhile, the nitrate (NO3 --N) and nitrite (NO2 --N) were also detected in two reactors (Fig. 1). The results concluded that the concentrations of NO3 --N and NO2 --N were varies slightly with reaction time and the concentration of NO3 --N and NO2 --N in two SBR were also similar. Therefore, the mechanism of the N removal in SBR with modified pyrite was highly related to anaerobic and aerobic oxidation of pyrite in the SBR [8].

Phosphorus removal
As shown in Fig. 2, TP concentration decreased from 6.93±0.14 mg L -1 in effluent of SBR with the fillings and 6.94±0.01 mg L -1 in effluent of control SBR to 3.03±0.81 mg L -1 and 1.84±0.46 mg L -1 , respectively. The TP removal efficiency in SBR with the fillings was 76.20±6.55%, which was higher than the removal efficiency of 56.28±11.47% in the control SBR (  [8,10]. According to the previous studies, the phosphorus could be used as nutrient for bacteria growing or enzyme composing and be essential in autotrophic denitrification [11,12]. And Zhu et al. (2011) reported that phosphate-accumulating organisms (PAOs) would uptake PO4 3--P when the carbon source was sufficient [13]. Moreover, PO4 3--P could be adsorbed by Fe 3+ or Fe(OH)3 produced by modified pyrite in SBR [14,15]. While the latest study suggested that phosphorus removal was mostly in form of Febound-P [8].

Conclusions
SBR was constructed with modified pyrite as the fillings, which concluded that modified pyrite had positive effects on the removal of nitrogen and phosphorus from wastewater. The average NH4 + -N and TP removal efficiencies in SBR with modified pyrite were 49.65±19.49% and 76.20±6.55%, respectively. The main mechanisms of simultaneous nitrogen and phosphorus removal may be anaerobic and aerobic oxidations of modified pyrite, and phosphorus reserved in the SBR was mainly in form of Fe-bound-P. Future studies should focus on confirming the role of modified pyrite on the removal of N and P and optimizing the key factors such as the particle size.