Fate of nitrogen in subsurface in filtration system for treating secondary ef fluent

Ying-hu Li,Hi-o Li,*,Xin-yng Xu,Si-yo Xio,Si-qi Wng,Shu-cong Xu

aSchool of Resources and Civil Engineering,Northeastern University,Shenyang 110004,China bSchool of Material Science and Engineering,Shandong University,Ji'nan 250002,China

Received 28 December 2016;accepted 23 March 2017 Available online 13 October 2017

Fate of nitrogen in subsurface in filtration system for treating secondary ef fluent

Ying-hua Lia,Hai-bo Lia,*,Xin-yang Xua,Si-yao Xiaoa,Si-qi Wanga,Shu-cong Xub

aSchool of Resources and Civil Engineering,Northeastern University,Shenyang 110004,ChinabSchool of Material Science and Engineering,Shandong University,Ji'nan 250002,China

Received 28 December 2016;accepted 23 March 2017 Available online 13 October 2017

The concentration of total nitrogen(TN)is reported to vary between 20 and 35 mg/L in domestic wastewater.In raw wastewater,ammonia nitrogenis the main nitrogen form,accounting for 70%-82%of the TN concentration.Organic nitrogen,nitrite nitrogenand nitrate nitrogenare present as well.For years,due to the lack of regulatory limits on nitrogen concentration in surface waters,nitrogen from secondary ef fluent has posed a signi ficant threat to the health of aquatic ecosystems.Researchers have made substantial efforts to reduce the nitrogen concentration in secondary ef fluent.As a kind of advanced wastewater treatment technology,the subsurface in filtration(SI)system has been widely used,owing to its advantages,which include low operation cost,easy maintenance,and low energy consumption.This review discusses the fate of various forms of nitrogen in SI treatment,including organic nitrogen,Major biological processes involved in nitrogen removal and the main factors in fluencing its transformation are suggested.Finally,it is shown that ammoni fication followed by nitri fication-denitri fication plays a major role in nitrogen removal.Further research needs to focus on the emission characteristics of gaseous nitrogen(generated from the nitri fication,denitri fication,and completely autotrophic nitrogen-removal over nitrite(CANON)processes)with respect to their greenhouse effects.

©2017 Hohai University.Production and hosting by Elsevier B.V.This is an open access article under the CC BY-NC-ND license(http://creativecommons.org/licenses/by-nc-nd/4.0/).

Wastewater treatment;Natural treatment technology;Subsurface in filtration;Nitrogen;Biological process

1.Introduction

In China,thousands of scientists and engineers have made substantial efforts in water pollution control.A number of technologies and procedures for municipal wastewater treatment have been developed and applied(Yuan and He,2015).Of those,the activated sludge technology and bio film treatment method are most commonly used for centralized wastewater treatment in big cities.

However,due to the lack of legal restraints,conventional sewage treatment processes,such as activated sludge and trickling filters,are known to be less ef ficient for nitrogen removal,compared with the removal ef ficiency of organic matter.Therefore,after a secondary treatment,wastewater discharge can be a major source of nutrients that cause eutrophication in lakes and rivers.Various physio-chemical processes such as filtration or adsorption have been employed for nutrient removal,but such processes are often costly and dif ficult to maintain(Bali et al.,2010;Curia et al.,2011).In an increasingly restrictive economic climate,especially in rural areas,capital resources for implementation and operation of such wastewater treatment infrastructure and facilities are limited.Therefore,natural treatment systems,such as wetlands,oxidation ponds,and subsurface in filtration(SI)systems,have emerged as appropriate alternatives.Compared with conventional systems,natural systems require less energy and less skilled labor(Sun et al.,2006).In SI,a kind of natural treatment system,wastewater is first treated by conventional physicochemical and/or biological treatment and then allowed to in filtrate through the aerated unsaturated zone,where it gets puri fied through processes such as filtration,adsorption,chemical processes,and biodegradation(Ji et al.,2012).SI systems have the advantages of low construction and operation costs,no aeration,low energy consumption,the ability to utilize fertilizer resources in wastewater,and simultaneous wastewater treatment and ecological service.Since 1990,large-scale SI systems have been developed in many countries to treat municipal wastewater(Phuong et al.,2014).Some demonstration treatment systems have also been established.Table 1 summarizes the treatment ef ficiency of SI systems.

In SI systems,the wastewater moves upwards slowly under capillary action,passing through the aerobic distribution layer,then the unsaturated and saturated substrate layers.During this movement,it is generally accepted that organic molecules are broken down by the microbiota through fermentation and/or respiration,and mineralized as a source of energy or assimilated into biomass(Qin et al.,2014),while the main removal mechanisms of phosphorus are short-term or long-term storage in soil/sediments and(to a lesser extent)plant uptake.More controversially,nitrogen can be removed through ammonia volatilization,soil adsorption,nitri fication-denitri fication,and anaerobic ammonium oxidation(Anammox).This paper reviews the forms and fates of nitrogen in secondary ef fluent,as well as the biological processes involved in nitrogen removal via SI treatment.

2.Nitrogen forms and levels in secondary ef fluent

Nitrogen levels in secondary ef fluent change with local lifestyle,local economic level,secondary treatment methods,and other factors.Generally,the secondary ef fluent from a municipal wastewater treatment plant is characterized by a low carbon-to-nitrogen ratio in China(Wang et al.,2015).Kim et al.(2011)suggested that 2mg/L is the lowest concentration for total nitrogen(TN),which can be removed from a treated wetland without additional chemical and mechanical treatments.If TN concentration in the in flow does not exceed this limit,the treatment system(e.g.,wetland and SI)may not remove TN effectively.Li et al.(2013)reported that ammonia nitrogenwas a dominating form of nitrogen in secondary ef fluent,accounting for 85.1%±1.2%of TN.The concentration of nitrate nitrogenwas only 0.1-0.2 mg/L,less than 1%of TN.Meanwhile,in the ef fluent of the SI system,theconcentration increased to 1.8-3.2 mg/L,accounting for 33.5%±2.3%of TN.On the other hand, theconcentration decreased to 2.0-3.7 mg/L,accounting for 64.4%±1.9%of TN.In the research presented by Boonchai and Seo(2015),TN mostly occurred in the form ofin secondary ef fluent from the Jinhae Sewage Treatment Plant in Changwon City,South Korea,with a concentration of 18.8 mg/L.Consistent with this conclusion,Jin et al.(2015)suggested that,in secondary ef fluent,nitrogen is mainly composed of inorganic nitrogen,i.e.,accounted for 65%of the TN in their study.The concentration of dissolved organic nitrogen changed little via secondary treatment,accounting for 15%-20%of TN.In raw water,the proportions of dissolved nitrogen and particulate nitrogen existed almost at the same level,with concentrations of 39.2 and 41.2 mg/L,respectively.Dissolved nitrogen was mainly composed of inorganic nitrogen,at a proportion of 93.2%(36.6 mg/L),but the concentration of dissolved organic nitrogen was relatively low(2.7mg/L).For particulate nitrogen,organic nitrogen was the main form(96.2%).Dissolved organic nitrogen was signi ficantly removed by anaerobic microbial degradation in the anaerobic and anoxic compartments of the secondary treatment,whereas a slight increase of dissolved organic nitrogen concentrations was observed in the aerobic compartment.

3.Nitrogen transformation process in SI systems

Generally speaking,nitrogen in secondary ef fluent goes into SI systems in the form ofand organic nitrogen(ON),along with a small amount of nitrate and nitrite.After complex reactions in the system,as shown in Fig.1,there may emerge several gaseous forms of nitrogen,e.g.,N2,NO,and N2O.

Table 1 Treatment ef ficiency of SI systems.

Fig.1.Nitrogen transformation in SI system.

3.1.Organic nitrogen removal

In SI systems,organic nitrogen is mainly removed by reactions of sedimentation, filtration,and adsorption of soil.The remaining organic nitrogen hydrolyzes into soluble amino acids,and the assimilation of animals,plants,and microorganisms may utilize some of the solid organic nitrogen directly.Water-soluble organic nitrogen is transformed into ammonium by ammoni fiers.It then enters the nitrogen cycle,or is partly adsorbed by microorganisms(Heil et al.,2015).

3.2.Ammonia nitrogen removal

There are two methods of ammonia nitrogen removal:(1)Adsorption followed by biological oxidation:Because the soil comprises electronegative colloids,that exists in the form of a positive charge is first adsorbed by soil colloids and soil particles.Then,absorbed by soil particles is dif ficult to move(Pan and Yu,2015).The cation exchange capacity(CEC)of soil determines the adsorption capacity of.At the same time,nitri fication,temperature,humidity,and pH can also affect the adsorption capacity ofAfter absorption by soil colloidal particles,ammonia nitrogen is removed by microbial nitri fication-denitri fication.In an SI system,nitri fication is completed by nitrite bacteria and nitrate bacteria.Nitrite bacteria oxidizeobtaining the energy,thenis ready to be oxidized intoby nitrate bacteria.Because the organic nitrogen in in fluent merely constitutes 15%-25%of TN,the ammoni fication process of organics does not limit nitri fication signi ficantly.Nitrate bacteria can use CO2as a carbon source.Thus,the carbon source is not the limiting factor of nitri fication,either.In addition,pH,temperature,dissolved oxygen(DO),and humidity are factors in fluencing the nitri fication process.The most important factor among them in the control of the nitri fication rate is DO.Research by Llor■ens et al.(2011)showed that,in SI systems,most of the nitrogen is nitri fied at 0.3 m below the water distributor,but,owing to respiration or saturation causing the oxygen level to drop,nitri fication may be slowed and even stopped.Their study showed that,when the system holds water for too long,such that it becomes saturated,anaerobics will emerge at the bottom of percolation ditch,which will reduce oxygen exchange and the nitri fication capacity of soil.This can also explain why the nitri fication performance of the SI system is not very strong when permeability is poor;is barely absorbed by soil,except for when it is used by organisms;and it is therefore free to move in the soil.Whenis transformed intoby nitrate bacteria and nitrite bacteria,they are eluted.The negative charge of soil particles is released again,and theadsorption capacity is recovered.(2)Volatilization:Ammonia nitrogen can escape from the SI system in the same way as NH3,and this is particularly signi ficant in dry climatic regions.The main factors affecting this process are the pH of the soil solution,the escape velocity of ammonia,and the temperature.In general,there is less volatilization of ammonia with greater depth of the soil in the SI system.The removal ef ficiency decreases at the same time.There are also some results showing that,when the pH value of soil is lower than 8.0,the volatilization of ammonia can be neglected(Kumar et al.,2015;Oksana et al.,2015).

3.3.Nitrate and nitrite removal

InSIsystems,themethodsofnitrateandnitriteremovalfrom sewage include the following:(1)Biological effects:Denitrifying bacteria use nitrate and nitrite as electron acceptors,converting them into gas under anaerobic conditions.Even under aerobic conditions,the denitri fication process in SI systems can also occur.This may be because of the anaerobic environment that is created by the respiration of microorganisms.Kong et al.(2002)observed that,after wastewater enters soil,with the increase of soil saturation,the release rate of N2O increases.The reason is that the contact between nitrate and inorganics affects the distribution of oxygen and microbial activity.In addition,temperature,pH,nitrate content,and organic carbon content can affect the denitri fication.(2)Chemical reaction:When theof the soil accumulates to a certain extent,it can react with organics in the SI system,and produce N2,NO2,and N2O.Generally,the process of nitrite nitrogen being converted into nitrate nitrogen occurs much faster than that of ammonia being converted into nitrate.When nitrogen in thegaseousformincreaseintheSIsystem,itcausestheincrease of pH and restrains the activity of nitrate bacteria.However,it does not restrain the activity of nitrite bacteria signi ficantly;nitrite nitrogen may accumulate.In the denitri fication process,if the content of nitrate is relatively high,microorganisms will first use nitrite nitrogen as electron acceptors,causing the accumulation of nitrite nitrogen.Foran SI system,the chemical reaction process of gaseous nitrogen generation is always ignored,but when the denitri fication reaction is signi ficant,it cannot be ignored.(3)Absorption by plants:Whenandmove with the water,they can be absorbed and used by plant roots,and they become the necessary elements for plant growth.Stewart et al.(2010)analyzed the relationship between TN removal and the nitrogen intake by plants.The results show that,with the decrease of soil permeability,the intake of nitrogen increases.When the pollution load of BOD is 1735 kg/(hm2.year),about 46%of TN is absorbed by plants.

4.Major biological processes involved in nitrogen removal

As shown in Fig.1,many mechanisms are involved in the fate of nitrogen:uptake of inorganic nitrogen by bacteria,mineralization,sedimentation,volatilization of ammonia,and nitri fication coupled with denitri fication.

For a long time,contradictory assumptions about the main mechanisms responsible for nitrogen removal have been proposed.Some researchers have argued that ammonia volatilization largely explains nitrogen removal by SI systems(O'Reilly et al.,2012).Other authors have reported that ammonia volatilization accounts for only a small fraction of nitrogen removal,even with a pH value as high as 8.5(Oksana et al.,2015).Most investigations have suggested that sedimentation and adsorption of ammonia are the major processes responsible for nitrogen removal in the start-up periods(Chen et al.,2014).For an SI system in stable performance periods,nitri fication coupled with denitri fication can be the major process of nitrogen removal,especially on warm days(Sun et al.,2012).Generally speaking,of the several methods of nitrogen removal by SI systems,nitri fication-denitri fication constitutes 69%-77%,the adsorption ofplantsconstitutes 14%-17%,and volatilization and other methods constitute 6%-17%(Bernadette et al.,2009).Biological effects are therefore the main form of nitrogen removal.During soil treatment of wastewater,organicnitrogeniseasilyconvertedintoammonifying bacteria in SI systems,can be adsorbed onto the soil because the zeta potential of the soil particles is negative,and the adsorption capacity of the soil can quickly return to the initial state due to nitri fication of intowhich is subsequently denitri fied to N2orN2O by the denitrifying biomass under the anoxic conditions.This is the circulation mode of nitrogen removal in SI systems.

4.1.Ammoni fication

The process of microorganisms decomposing organic nitrogen compounds into ammonia is called ammoni fication.It can also be called mineralization.In SI systems,the majority of ammonia produced through ammoni fication is converted into nitrate and nitrite,and the rest is absorbed by plants or escapes from the system.A lot of bacteria,fungi,and actinomycetes can secrete proteases,which can break down proteins into polypeptides,amino acids,and NH3.Of the microorganisms,those with the ability to decompose organic nitrogen are called ammonifying microorganisms,and are widely distributed in nature,including in aerobic,facultative,and anaerobic conditions.

In SI systems,the factors affecting ammoni fication mainly include the following:(1)Temperature:Within the range of 25°C-35°C,the velocity of ammoni fication grows with the increase of temperature,but the absorption of plants rises at the same time.(2)Moisture content of the substrate:Ammoni fication is enhanced with the rise of water content of soil,but when the water content rises to some certain extent,the velocity of ammoni fication drops quickly.(3)Organic matter:Ammoni fication velocity is positively correlated with the content of organic matter.(4)Soil depth:In general,the velocity of ammoni fication drops with the rise of soil depth.The amount of organic matter at different depths of soil is also the main factor affecting ammoni fication.Within the range of 0-120 cm,ammoni fication is different at different soil depths.This is because,with the increase of soil depth,the breathability of microorganisms and mineralization extent of organic matter always change.With the breathability of soil dropping gradually,less organic matter can be degraded,the amount of microorganisms drops quickly,and the ammoni fication rate decreases.(5)Other factors:The physical state of soil,like the water content in the soil and the degree of compaction,affects the condition of soil breathability.Any changes in these factors affect the ammoni fication performance of SI systems.

4.2.Nitri fication

Nitri fication is completed by autotrophs in stages(Pan et al.,2013).The first stage is nitrosation,which is the stage of converting ammonia into nitrite.There are mainly five species of nitrite bacteria taking part in this stage:Nitrosomonas,Nitrosocytis,Nitrosococcus,Nitrosospira,and Nitrosogloea.The second stage is nitri fication,during which nitrite is oxidized into nitrate.There are mainly three species taking part in this stage:Nitrobacter,Nitrospina,and Nitrococcus.As shown in Fig.2,N2O and NO are produced as reaction byproducts during the process in whichis oxidized intowhere N is an electron acceptor(Kong et al.,2002):

Fig.2.Nitri fication process for gaseous nitrogen production.

The factors affecting nitri fication in SI systems include temperature,DO,pH,carbon-to-nitrogen ratio(C/N ratio),and toxic substances(Zhao et al.,2013):(1)Temperature:Temperature affects nitri fication mainly by affecting the fluidity of microbial cells and the activity of biological macromolecules.On one hand,the reactive velocity of microbial cells accelerates,and the metabolism and growth are accordingly accelerated.On the other hand,with the rise of temperature,biologically active substances change,the functioning of cells drops,and the microorganisms may even die.Biological nitri fication can occur within the range of 4°C-45°C,the most suitable temperature for nitrate bacteria is 35°C,and the most suitable temperature range for nitrite bacteria is 35°C-42°C.Temperature not only affects the growth rate of nitrate bacteria,but also the activity of bacteria.Jin et al.(2014)showed that nitri fication is limited at temperatures below 10°C,and stops below 6°C.Thus,low temperature limits nitri fication.When the temperature rises from 5°C to 30°C,the velocity of nitri fication also rises.(2)DO:Nitri fication is generally carried out under aerobic conditions.The concentration of DO can also affect the velocity of nitri fication,which generally should be above 2 mg/L.For SI systems,DO is not as sensitive as it is to activated sludge and bio film systems.However,in order to ensure adequate nitri fication,the DO level must be high,generally greater than 3mg/L,because 4.6 mg/L of DO is needed to oxidize 1.0 mg ammonia.(3)pH:Nitrate bacteria are generally thought to be pH-sensitive.The most suitable pH range is from 6.6 to 8.0,and the best value is 8.0.When the pH value is less than 6.0,the nitri fication rate will drop,and when the pH value is below 4.5,nitri fication will stop.Fan et al.(2013)argued that the most suitable range of pH values for nitrate bacteria is 7.2-8.0 when the system is under acidic conditions,and the activity of nitrite bacteria as well as nitrate bacteria is limited signi ficantly.(4)C/N ratio:The C/N ratio is an important factor in the velocity and process of nitri fication.If the BOD concentration is high in sewage,heterotrophic bacteria will compete with nitrate bacteria for substrate and oxygen,restricting the growth of nitrate bacteria.(5)Toxic substances:High levels of NH3-N,heavy metals,toxic substances,and organics will suppress nitri fication.There are two main interventions that suppress nitri fication:interfering with the metabolism of cells,and destroying the original oxidation ability of bacteria.Generally speaking,for a toxic substance,the effect of nitrite bacteria is more signi ficant than that of nitrate bacteria.There are two reasons that organics suppress nitri fication(Rodriguez-Caballeroa et al.,2014).The first is that,when the organic concentration is high,the amount of heterotrophic bacteria will be much greater than that of nitrite bacteria,and get in the way of ammonia passing to nitrite bacteria.The number of nitrate bacteria is also reduced because of the existence of heterotrophic bacteria.Another reason is that some organics are toxic or suppressive to nitrite bacteria.

The nitri fication potential,an indicator of nitrifying bacteria activity,was measured as in Kadlec et al.(2010).20 g soil samples were added to 100 mL of NH4Cl solution with a nitrogen concentration of 500mg/L.The mixture was incubated at 25°C for 24 h.The amount of nitri fied ammonium nitrogen was determined by Eq.(1):

where ω is the rate of nitrate nitrogen production(mg/(kg.h)),ρiis the initialconcentration of substrate solution(mg/L),ρtis theconcentration of the solution after incubation time t(mg/L),ρbis theconcentration of the blank sample after incubation time t(mg/L),V is the volume of substrate solution(L),M is the mass of the soil sample(kg),w is the water content of the soil sample(fraction),and ρ is the density of water at 25°C(0.997 kg/L).

In nitri fication depths,the Pearson correlation coef ficients of functional genes amoA-nxrA,amoA-napA,nxrA-napA,amoA-nirK,amoA-nosZ,nirK-nosZ,nxrA-nosZ,napA-nosZ,nxrA-nirK,and napA-nirK were all greater than 0.6(P＜0.05).Thus,associated enrichments between amoA,nxrA,napA,nirK,and nosZ existed.Based on this in-depth analysis,it was found that amoA,nxrA,napA,nirK,and nosZ genes could be expressed under aerobic conditions and had similar ecological adaptability during the nitri fication process(Ji et al.,2012).

4.3.Denitri fication

Denitri fication is the process,in which denitrifying bacteria deoxygenate nitrite,with N2or N2O released under hypoxic conditions.Most denitrifying bacteria are facultative anaerobes,such as Achromobacter,Aerobacter,Alcaligenes,and Pseudomonas.These denitrifying bacteria use many kinds of organic substrate as electron donors,deoxygenatinginto N2by stages.According to Jacobs and Harrison(2014),N2O is produced as an intermediate product during the denitri fication process in whichis converted into N2(Fig.3).

In SI systems,the main factors that affect denitri fication are the following:(1)Temperature:The most suitable temperature range for denitri fication is 20°C-40°C.When the temperature is below 15°C,the velocity of denitri fication will drop dramatically,and when the temperature is below 3°C,the denitri fication process will stop.(2)DO:Denitri fication generally occurs under conditions of hypoxia.DO will restrain the denitri fication process,because DO will compete with nitrate for electron donors,which can also restrain the mixture and the activity of nitrate reductase.(3)pH:The most suitable pH values for denitri fication are 6.5-7.5.Unsuitable pH values will affect the growth of denitrifying bacteria and the activity of enzymes.When the pH value is below 6.0 or above 8.0,it will restrain the denitri fication process signi ficantly.The denitri fication process will produce alkalinity,which is helpful to maintaining the pH value within a suitable range.(4)Carbon source:Generally,when the BOD/total Kjeldahl nitrogen(TKN)ratio is above 3 to 5 in SI systems,the carbon sources are suf ficient.If organics are in shortage in wastewater,carbon sources,like methanol,ethanol,citric acid,and molasses,need to be added.Municipal wastewater and some industrial wastewater can also serve as the carbon sources of denitri fication.The term solid-phase denitri fication came into existence in recent years,where solid substrates were used as a constant carbon source for denitri fication and also to provide a platform for microbial biomass development.Materials such as starch,polycaprolactone,and polybutylene succinate have been widely used due to their excellent denitri fication ef ficiencies in short time periods(Ashok and Hait,2015).The donor species or technologies can be adopted,depending on environmental,physical,and biological conditions of the environment,and should not lead to pollution swapping.

Fig.3.Denitri fication process for gaseous nitrogen production.

The method of determining the denitri fication potential of the soil medium developed by Kadlec et al.(2010)was used with slight modi fications.A 10 g soil sample was incubated with a 100 mL KNO3substrate solution in each 250 mL flask,and the flasks were shaken at 180 rpm for 48 h at 37°C.After the incubation,soil suspension was filtered through a 0.45 μm-membrane filter for further analysis ofandconcentrations.The denitri fication potentialwas calculated by Eq.(2).

where ω′isthe rate ofgaseousnitrogen production(mg/(kg.h)),is the initialconcentration of the substrate solution(mg/L),is theconcentration of the solution after incubation time t(mg/L),is theconcentration of the solution after incubation time t(mg/L),represents theconcentrations of blank samples after incubation time t(mg/L),V1is the volume of the substrate solution(L),V2is the water content of the soil sample(L),m is the mass of the soil sample(kg),and k is the moisture coef ficient of the matrix at 25°C.

Li et al.(2013)reported that denitri fiers increased with soil depth.In contrast,nitri fiers decreased with the increase in soil depth.Ji et al.(2012)found that the associated enrichments between narG and qnorB genes existed in anaerobic areas.Based on the depth analysis,they insisted that narG and qnorB could be expressed under anaerobic conditions and they had similarecologicaladaptability during the denitri ficaiton process.

4.4.CANON and Anammox

Nitrogen removal from municipal wastewater via the traditional nitri fication-denitri fication has become a key process in biological treatment over recent decades.However,the implementation of conventional nitrogen removal,i.e.,the aerobic conversion of ammonia to nitrate combined with the anaerobic conversion of nitrate to nitrogen gas,is energyintensive,mainly because of aeration costs(Susanne et al.,2014).In recent years,a new pathway for ammonium removal has been discovered and widely studied:completely autotrophic nitrogen-removal over nitrite(CANON)and anaerobic ammonium oxidation(Anammox).

CANON is closely related to the Anammox process,in whichis first partially oxidized into nitrite nitrogen before being transformed with the remaining ammonium into dinitrogen gas by planctomycetes-like bacteria.These planctomycetes-like bacteria grow in the anaerobic zones of treatment systems.The stoichiometry of the CANON process is described by Eq.(3):

Because the CANON process generates dinitrogen gas,the loss of TN in the sewage is a natural result.According to the research by Zhang et al.(2015),theconcentration decreases and theconcentration increases with soil depth.In comparison,TN concentration is unceasingly reduced.Above the soil depth of 1.30 m,30.5%of the nitrogen disappeared.Through mass balance analysis,it was observed that the CANON process was responsible for transforming nitrogen into a gaseous form.In addition,47.5%of theparticipated in the CANON process,and the rest played a role in nitration.At depths of 1.30-2.00 m,more than 90%of the TN was composed ofwhich indicated that the removal of TN occurred via denitri fication.

Kuypers et al.(2003),who found Anammox bacteria at the bottom of Black Sea,have shown that the Anammox process is promising for the treatment of low-ammonium content wastewater.The reaction equation for Anammox is Eq.(4):

To date,there are no reports suggesting that Anammox is of high importance in nitrogen removal via SI treatment.With respect to constructed wetlands(CWs),there exist controversial conclusions about whether or not Anammox is as importantasnitri fication-denitri fication fornitrogen removal.Through molecular biological analyses,research by Zhi and Ji(2014)showed that aerobic ammonia oxidation was the dominant ammonium removal path when the C/N ratio was less than or equal to six.However,when the C/N ratio was greater than six,the importance of Anammox was notably enhanced,resulting in another primary ammonium removal pathway,in addition to the aerobic ammonia oxidation.He et al.(2012)examined the effects of pH and seasonal temperature variation on simultaneous partial nitri fication and Anammox in two free-water surface wetlands.Fluorescence in situ hybridization analysis found that aerobic ammonium oxidizing bacteria and Anammox bacteria accounted for 42%-73%of the bacterial populations in the wetlands.A seasonal temperature variation of more than 6°C would affect simultaneous partial nitri fication and Anammox signi ficantly.Signi ficant pH effects were identi fied when the temperatures were below 18.9°C.Anammox was the limiting stage of simultaneous partial nitri fication and Anammox in the wetlands.In comparison,Takaaki et al.(2011)suggested that,although Anammox bacteria were detected by molecular methods,Anammox activity could not be measured and therefore that this process appears to be of low importance in nitrogen transformations in these wastewater eco-treatment systems.Due to the differences between operations of a SI system and CW,e.g., flow-moving paths,hydraulic retention time,substrate composition,and construction,Anammox should be another future research focus relating to the nitrogen removal process via SI systems.

5.Conclusions

(1)During the wastewater treatment process using SI systems,mineralization,volatilization,adsorption,nitri fication,and denitri fication are the main paths of organic nitrogen,ammonia nitrogen,removal.

(2)With respect to TN removal,ammoni fication followed by nitri fication and denitri fication constitutes 69%-77%,adsorption of plants 14%-17%,and volatilization and other methods 6%-17%,showing that biological effects are the main method of nitrogen removal.

(3)The CANON process could be responsible for transforming nitrogen into a gaseous form in upper layers of SI systems.Both denitri fication and the CANON reaction transform in fluent nitrogen into a gaseous form,expressed by N2O,N2,and NO.

(4)Further research needs to focus on the emission characteristics of gaseous nitrogen and its spatial distribution in soil.

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This work was supported by the National Natural Science Foundation of China(Grants No.41571455 and 51578115)and the Basic Science Research Fund of Northeastern University(Grant No.N160104004).

*Corresponding author.

E-mail address:graceli_2003@163.com(Hai-bo Li).

Peer review under responsibility of Hohai University.