Chemosphere 204 (2018) 51e62

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Chemosphere
journal homepage: www.elsevier.com/locate/chemosphere

Review

Role and application of iron in water treatment for nitrogen removal:

A review
Hongbo Liu a, *, Zihua Chen a, Yongnian Guan b, Suyun Xu a
a
School of Environment and Architecture, University of Shanghai for Science and Technology, 516 Jungong Road, 200093, Shanghai, China
b
Qingyuan-Hong Kong & China Water Co. LTD, 215000, Suzhou China

h i g h l i g h t s g r a p h i c a l a b s t r a c t

State of arts regarding to iron applied

in nitrogen removal was reviewed.

Pathways of nitrogen removal with
the coupling of iron were proposed.

Challenges and role of iron in water
treatment for nitrogen removal were
discussed.

Solutions to pertinent practical engi-
neering application of iron were
proposed.

a r t i c l e i n f o a b s t r a c t

Article history: It is crucial to have a review on the role of iron in water treatment for the guidance towards the selection
Received 2 November 2017 of appropriate processes, content of iron, and application conditions, as there are few reviews available at
Received in revised form present and the systematic information is lacking for both researchers and engineers. The objectives of
3 April 2018
this review are to summarize the state of arts with respect to iron applied in nitrogen removal, discuss
Accepted 3 April 2018
chemical and biological or bio-chemical combined nitrogen removal pathways and processes coupled
Available online 6 April 2018
with iron, and to reveal reaction mechanisms as well as providing references or even solutions to
Handling Editor: Hyunook Kim pertinent the practical engineering application of nitrate removal coupling with iron. The following
information have been summarized and discussed in details: (1) iron based materials with varieties of
Keywords: preparations and forms, (2) major coupling ways of nitrogen removal methods or processes with iron
Iron application, (3) chemical reaction equations about a variety of chemical and biological or bio-chemical
Water treatment combined processes and the main mechanisms. In addition, challenges and/or drawbacks during the
Nitrogen removal nitrogen removal processes will also be discussed in this paper, which is aimed to seek better practical
Existing states
engineering applications of nitrate removal coupling with iron.
Mechanism
© 2018 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license
(http://creativecommons.org/licenses/by/4.0/).

Contents

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 52
2. Existence forms and preparation methods of iron applied in nitrogen removal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53
2.1. The existence forms of iron in nitrogen removal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53
2.2. Preparation methods of iron applied in nitrogen removal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53

* Corresponding author.
E-mail addresses: Liuhb@usst.edu.cn (H. Liu), xusuyun1127@126.com (S. Xu).

https://doi.org/10.1016/j.chemosphere.2018.04.019
0045-6535/© 2018 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).
52 H. Liu et al. / Chemosphere 204 (2018) 51e62

2.2.1. Modified by physical or chemical methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53

2.2.2. Iron associated, loaded or loading with other matters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53
2.2.3. Joint use of diverse valent iron . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 54
2.2.4. Fe0 in different sizes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 54
2.2.5. Fe0 combined with transition metal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 54
3. Iron applied in the chemical removal of nitrogen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55
3.1. Iron applied in the chemical removal of nitrogen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55
3.2. Mechanisms involved in chemical removal of nitrogen with iron . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55
3.2.1. Pertinent chemical reactions equations in chemical removal of nitrogen coupling with iron . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55
3.2.2. Electron transfer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55
3.2.3. Iron as catalysts . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 56
4. Iron applied in biological or bio-chemical coupled nitrogen removal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 56
4.1. Pathways and processes of biological or bio-chemical combined nitrogen removal processes coupling with iron . . . . . . . . . . . . . . . . . . . . . . . . . 56
4.2. Mechanisms of nitrogen removal in biological or bio-chemical combined processes coupled with iron . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58
4.2.1. Chemical reaction equations in biological or bio-chemical coupled nitrogen removal processes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58
4.2.2. Electron transfer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58
4.2.3. Effect on activity of enzymes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59
4.2.4. Effect on microorganism and microbial community . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59
5. Conclusions, challenges, and future prospects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60
5.1. Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60
5.2. Challenges and future prospects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 60
Acknowledgements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61
Supplementary data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61

1. Introduction 1996), methemoglobinemia (Huang et al., 2012) and bladder,

ovarian (Weyer et al., 2001) or gastric (Liu et al., 2016) cancers, etc.
Nitrogen is an essential element for living creatures, while to humans. The nitrate load in surface water is commonly consid-
excessive release of both organic (Camargo and Alonso, 2006) and ered as the cause of water quality deterioration and eutrophication
inorganic (Sharma, 2010b) nitrogen species can bring about severe (Arnon et al., 2007). High loads of nitrogenous compounds in sur-
environmental problems. As a major contributor to oxygen demand face water can also affect the ground water because of the inter-
of receiving waters, ammonia has been considered as critical action between the ground and surface water resources (Novotny
pollutant causing eutrophication in water environment (Sharma et al., 2010). Nitrogen pollution mainly results from the use of
et al., 1998; Park et al., 2009). Nitrate pollution in drinking water nitrogen-containing fertilizers, animal waste, septic system, in-
might increase the risk of non-Hodgkin's lymphoma (Ward et al., dustrial processes atmospheric deposition from nitrogen oxide

List of abbreviations NADH Nicotinamide adenine dinucleotide

NAFO Nitrate-dependent anaerobic ferrous oxidizing
AHLs Acylhomoserine lactones NAR Nitrate reductase
ANAMMOX Anaerobic ammonium oxidation NDAMO Nitrite dependent anaerobic methane oxidation
BES Bioelectrochemical system NH3-N Ammonia
Cyt. b, c, c1 Cytochrome b, c, c1 NIR Nitrite reductase
DO Dissolved oxygen NOR NO reductase
EIR Electrically induced reduction nZVI (nFe0) Nanoscale zero valent iron
EK/Fe0 PRB Electrokinetic/Fe0 permeable reactive barrier pHin Initial pH
Fe0-C MEBD Fe(0) -carbon micro-electrolysis and biological PCA Principal component analysis
denitrification PNP p-nitrophenol
Fe2þaq Dissolved Fe2þ PPAD termed pyrite autotrophic denitrification
Fe(II)-MAD Fe(II)-mediated autotrophic denitrification PRB Permeable reactive barrier
Fe-CAD Ferrous iron-based chemo autotrophic denitrification OD422 Optical density at 422 nm
FHC Ferrous hydroxyl complex ORP Oxidationereduction potential
GRs Green rust system Q Quinone pool
HDH Hydrazine dehydrogenase SDH Succinate dehydrogenase
HFO Hydrous ferric oxide SNDPR Simultaneous nitrification, denitrification and
HIDs Hydrogenotrophic integrated denitrification system phosphorus removal process
HRT Hydraulic retention time TN Total nitrogen
HZS Hydrazine synthesis VLR Volumetric loading rate
kobs Degradation rate constant VRR Volumetric removal rate
mZVI (mFe0) Micro-size zero valent iron ZVI (Fe0) Zero valent iron
N2OR N2O reductase
 H. Liu et al. / Chemosphere 204 (2018) 51e62
emission (Xu et al., 2012), irrigation and storm runoffs from farm-
lands (Huang et al., 1998) etc., which has been an increasingly
ubiquitous issues worldwide, affecting the quality of drinking wa-
ter, the ecology, and the value of the aquatic ecosystems. For
instance, two-thirds of coastal rivers and bays in the United States
have been moderately or even severely degraded by nitrogenous
pollution (Kevin, 2001); More than 85% of the lakes and about 82%
of the 532 major rivers in China have experienced the deterioration
of their water environment and eutrophication due to heavy N
pollution (Novotny et al., 2010). Due to the tremendous perni-
ciousness caused by the excessive release of nitrogen, more strin-
gent emission standards of nitrogen-containing pollutants have
been issued (Tong et al., 2013; Zhang et al., 2014a). The World
Health Organization (WHO) has set the maximum contaminant
level for nitrate of 10 mg N L1 in drinking water (Shi et al., 2016).
The US environmental protection agency (USEPA) has set
maximum contaminant levels of 10 mg L1 and 1 mg L1 for NO 3 -N
and NO 2 -N, respectively (USEPA, 2016). The draft of China's new
discharge standard of pollutants for municipal wastewater treat-
ment plants (GB18918-2016) has decreased its total nitrate level to
10 mg L1 from previous 15 mg L1 (MEPPRC, 2015).
For a long time, due to iron's various advantages, a great deal of
interest has been developed to study using iron in the treatment of
polluted water (Marsalek et al., 2012), sludge disposal (Kim and
Sharma, 2004; Jiang and Sharma, 2008), air pollution control
(Jabłon
ska et al., 2013), the remediation of contaminated soil
(Suzuki et al., 2012b), groundwater (Shi et al., 2011), and wetlands
(Song et al., 2016). To be more specific, miscellaneous morphologies
of iron have been extensively used in various ways to degrade ni-
trogen containing inorganic (Sharma, 2010a, 2011) and organic
pollutants, including atrazine (Wang et al., 2014), nitro aromatic
compounds (Agrawal and Tratnyek, 1995), nitrobenzene (Wu et al.,
2013) etc. It is momentous to emphasize that iron plays a crucial
role in nitrogen removal. Especially, many studies have been
concentrated on denitrification by combining iron with various
processes, including abiotic (Tai and Dempsey, 2009) and biotic
processes (Zhou et al., 2016). On one hand, mono, bimetallic and
even polymetallic iron materials were used in nitrate reduction (An
et al., 2010c; Lai et al., 2013b). Meanwhile, various iron associating,
coating or being coated with other materials or substances were
applied in the treatment of nitrate containing wastewater (Lai et al.,
2013a; Lee et al., 2016). Iron mines such as FeS2 (Tong et al., 2017b)
and goethite (Jung et al., 2014) have also been explored as options
for nitrogen removal. On the other hand, pathways or processes of
nitrogen removal have been combined with iron during denitrifi-
cation processes, such as anaerobic ammonium oxidation (ANAM-
MOX) (Zhou et al., 2016), Fe0-carbon micro-electrolysis (Deng et al.,
2016), bioelectrochemical system (BES) (Zhu et al., 2015) and
permeable reactive barrier (PRB) (Suzuki et al., 2012b).
The removal of nitrogen containing pollutants and the applica-
tion of iron in water treatment for denitrification have raised wide
spread concerns. Although reviews referring to the application of
iron in the environmental field as well as overviews about nitrogen
removal technologies in water treatment have been extensively
published in recent years, most reviews are focusing on iron or
denitrification. Few reviews have paid attention to the specific use
of iron in nitrogen removal. Above all, during the water treatment,
the selection of processes depends on the type of influent waste-
water and it's water quality (Zhu et al., 2015; Deng et al., 2016); the
inappropriate amount of iron added in denitrification processes
may result in undesired efficiency, such as the Fe(III)'s effect on the
N2O yield (Jia et al., 2016) or the ANAMMOX activity (Chen et al.,
2014) and the effect of Fe0 contents on biomass concentrations
(An et al., 2009). The difference in application conditions may also
bring about undesired products (Sharma et al., 1998; Wu et al.,
53
2015). In addition, the catalytic effects may decrease with a slight
difference in operation condition like temperature (Lubphoo et al.,
2015), pH (Liou et al., 2009) and the type or ratio of metal (Lubphoo
et al., 2016) when iron was considered as the important part of the
catalyst in water treatment for nitrogen removal. Therefore, it is
vital and meaningful to write a review on the application of iron
nitrogen removal of wastewater.
The objectives of this review are to (1) summarize the existing
states and preparation methods of iron applied in nitrogen removal,
(2) sum up the chemical methods and biological or bio-chemical
combined pathways or processes used in nitrogen removal
coupling with iron, (3) make a summary of mechanisms, and (4)
provide references or solutions to the pertinent practical engi-
neering application in nitrate removal of water treatment coupled
with iron.
2. Existence forms and preparation methods of iron applied
in nitrogen removal
2.1. The existence forms of iron in nitrogen removal
Various categories and valences of iron are widely applied in
nitrogen removal processes, including Fe0, ferric oxides, iron hy-
droxides, ferric salts and ferrous mineral. Table S1 gives a summary
of the iron-based materials that have been extensively used in ni-
trogen containing pollutants removal through chemical or biolog-
ical pathways.
2.2. Preparation methods of iron applied in nitrogen removal
Iron in various valences has been increasingly applied to nitrate
removal. However, generally it is not directly used for removal of
nitrogen, but need to be improved by different preparation
methods and associating, coating or being coated with other ma-
terials and substances in most cases. A large number of researchers
paid their attention to adopting different preparation methods of
iron to get appropriate properties.
2.2.1. Modified by physical or chemical methods
Many physical or chemical methods have been adopted to
modify iron applied in chemical removal of nitrogen. The primary
purposes of these methods are to prevent passivation (Luo et al.,
2010), keep continuous activation of passive layers (Guo et al.,
2015), provide an accessible surface area (Lee et al., 2016) so as to
ultimately improve mass and electron transfer efficiency.
Premagnetization has been utilized to improve the reduction of
nitrate and selectivity by enhancing mass transfer (Fe2þ, Hþ etc.)
(Ren et al., 2017). Common oxidants (O2, H2O2, KMnO4 and CrO4 2)
have been utilized to activate the reactivity of Fe0 in facilitating
nitrate reduction (Guo et al., 2015). Macroporous alginate substrate
based Fe0 nanoparticles has been synthetized by chemical methods
to avoid the formation agglomerates by Luo et al. (2010). Meso-
porous spheres of copper on iron/aluminum oxide with various
iron content were synthesized and applied in denitrification reac-
tion showing that iron-based spheres were active in the reaction,
with minor ammonia formation (de Sousa et al., 2014). Another
case where electrochemical and ultrasonic methods were com-
bined to produce nFe0 verified that the nitrate decline was faster in
terms of g NO 0
3 -N/g nFe at lower pH (Chen et al., 2004).
2.2.2. Iron associated, loaded or loading with other matters
Iron is broadly associated, loaded or loading with other matters.
nFe0 particles coated with chitosan and sodium oleate were
compared to support autotrophic denitrification and a conclusion
was drawn that sodium oleate might be more appropriate as the
54 H. Liu et al. / Chemosphere 204 (2018) 51e62

intermediate product ammonium increased when nFe0 coated with considerable scholars concentrate on applying iron particles of
chitosan was applied (An et al., 2010b). There are far more examples different sizes, including cast iron filings (Laskowski et al., 2015),
that could be discussed, such as, nitrite reduction through Fe2þ iron craps or powder (Liu et al., 2013; Deng et al., 2016), micro-size
associated with kaolinite (Rakshit et al., 2016). zero valent iron (mFe0) (Choi et al., 2009; Suzuki et al., 2012a; Xiong
et al., 2016), nano-size nFe0 (An et al., 2009; Hamid et al., 2015; Lee
et al., 2016). In general, nano-sized metal particles are more reac-
2.2.3. Joint use of diverse valent iron
tive than micron-scale powders, most probably attributing to the
Quite a few researchers gave their insight into the joint use of
high specific surface area and increasing surface reactivity (Sun
diverse valent iron in nitrate removal. Fe0 coexisting with dissolved
et al., 2016). However, some studies also revealed that there was
Fe2þ was used for the reduction of NO 
3 , in which the NO3 reduction
no particular correlation between the Fe0 performance and specific
and the decreased Fe2þaq concentration are promoted by each
surface area (Velimirovic et al., 2013).
other, thus sustaining reactivity of Fe0 for nitrate reduction can be
acquired (Han et al., 2016). In 2003, the “ferrous wheel hypothesis”
was proposed by Davidson et al. (2003) with regard to the reduc-
2.2.5. Fe0 combined with transition metal
tion of Fe3þ (hydr)oxide by the carbonaceous compounds followed
It was often seen that mono, bimetallic even polymetallic iron
by the NO 
3 to NO2 reduction and Fe
2þ
to Fe3þ oxidation. Although
containing substances were greatly used for denitrification in
the dispute upon this is still on going, it is obvious that there are
abundant studies. The performance of nitrate removal and N2
some inevitable relations between the system of NO 3 to NO2

selectivity were investigated based on nFe0 supported Cu-Pd
reduction and Fe2þ to Fe3þ oxidation. Fe(II)/Fe(III) oxide was usually
bimetallic catalyst (Cu-Pd/nFe0) in a continuous reactor system
used as an important redox couple in environmental systems,
(Hamid et al., 2015). Bimetallic Fe/Ni nanoparticles supported by
including nitrite reduction (Tai and Dempsey, 2009; Bao and Li,
kaolinite (K-Fe/Ni) were used to simultaneously remove Cu(II) and
2017).
nitrate (Cai et al., 2014). Other examples can be also found in a large
number of publications, such as Fe-Pd bimetallic nanoparticle
2.2.4. Fe0 in different sizes supported on chelating resin for the selective reduction of nitrate
When it comes to Fe0 that has been applied in nitrate removal, into nitrogen (Shi et al., 2016), highly-efficient reduction of nitrate

Table 1
Chemical methods adopted to remove nitrogen-containing pollutants coupling with iron.

Target Method Conditions Treatment efficiency Ref.

Groundwater NO 3, Fe2þ and Fe3O4 T ¼ 25  C, initial pH (pHin) ¼ 6.7, The nitrate removal efficiency reached 93% under the (Xu et al., 2012)
C0 ¼ 50 mg L1 promoted Fe0 CFe(0) ¼ 10 g L1,CFe(II) ¼ 50 mg L1 optimal conditions
system
Nitrate-containing Premagnetization T ¼ 25  C, pHin ¼ 7.0, intensity of magnetic A high rate (kobs for nitrate reduction of (Ren et al.,
secondary NO3, Fe0/(Fe/Cu) system field ¼ 20 mT, premagnetization time ¼ 30 min, 0.732 min1), good selectivity of [TN removal]/[ 2017)
C0 ¼ 20 mg L1 mass percentage of Fe0 ¼ 40%, total mass of medium NO3 -N removal]
materials ¼ 20 g and hydraulic retention
time ¼ 4.2 min
Nitrate contaminated Fe0 þ Fe/Cu system pHin ¼ 7.0, the hydraulic retention time and M(Fe0)/ High rate (kobs for nitrate reduction of 0.403 min1), (Ren et al.,
water NO 3, M(Fe/Cu) ratio was 16 min and 0.67 respectively. 99.0% NO þ
3 -N removal and low NH4 -N generation 2016)
C0 ¼ 50 mg L1 rate (61.1%)
Nitrate-polluted Cu/Fe0 bimetallic The best weight ratio of Fe0/NO
3 was 15. Comparing with the tests without copper, complete (Muradova
groundwater with nanoparticles removal time of nitrate was reduced by 1.5 times et al., 2016;
initial nitrate system when copper was added. And the nitrate removal Vilardi and Di
concentration of process contained both reduction and adsorption. Palma, 2017)
57.5 mg L1
Groundwater NO 3, Fe2þaq enhanced Fe2þaq dosage of 1.0 mM, pHin from 5 to 9 97% reduced NO þ
3 was transformed into NH4 under (Han et al.,
C0 ¼ 2 mM Fe0 nitrate best conditions 2016)
reduction system
Nitrate contaminated nFe0/MAS system nFe0 was 2.0 g L1 More than 96.5% of nitrates were removed from an (Lee et al.,
water aqueous solution in 30 min, while the control was 2016)
only 44.7% after longer time.
Groundwater NO 3, Electrically Low-voltage direct current electric field of 6 V/m Nitrate reduction was 47.8% faster than in the ZVI (Luo et al.,
C0 ¼ 5000 mg L1 induced reduction reactor without supplemented current. 2010)
(EIR) PRB system
Nitrite contaminated Fe(II)/HFO/oxidant pH 6.8, 26e28  C and strictly anaerobic The reaction was fast in the presence of hydrous (Tai and
water systems Fe(II)/Fe(III) molar ratio less than 3.0 ferric oxide (HFO). Dempsey,
NO2 , C0 ¼ 500 mM 2009)
Nitrite contaminated Ferrous hydroxyl pHin ¼ 8, Fe2þ/OH molar ratio ¼ 7:12 Nitrite and total nitrogen removal efficiency can (Wu et al.,
groundwater and complex (FHC) approach 95% and 75% respectively. 2015)
denitrification in nitrite Ammonium decreased when Fe2þ/OH molar ratio
water, NO 2, denitrification range from 8:12 to 12:12.
C0 ¼ 10 mM system
Industrial and The NO 2
2 GR(CO3 ) pH and initial Eh was 9.5 and - 473 mV, respectively. 97.1% NO
2 was degraded and the total nitrogen (Lu et al., 2016)
agricultural system (GRs) removal efficiency 89.6% under optimal conditions.
wastewater, NO 2,
C0 ¼ 10.71 mM
Ammonia-containing Ferrate(VI) T ¼ 25  C, pH from 7.5 to 11 The highest ammonia removal efficiency approaches (Sharma et al.,
wastewater, NHþ 4, oxidation system 22% at best pH value. 1998)
C0 ¼ 0.05 M
Refractory wastewater mFe0/O3 process pHin ¼ 5.3, O3 flow rate 1.5 L min1, Fe0 dosage COD removal efficiency was 89.5% under the optimal (Xiong et al.,
PNP, C0 ¼ 500 mg L1 40.0 g L1 conditions 2016)
 H. Liu et al. / Chemosphere 204 (2018) 51e62
to nitrogen by metals/NaY (Soares et al., 2015) or iron deposited
bimetals (Liou et al., 2009).
3. Iron applied in the chemical removal of nitrogen
3.1. Iron applied in the chemical removal of nitrogen
It is prevalent that various iron-based materials have been
applied in the chemical removal of nitrogen-containing pollutants
in groundwater (Xu et al., 2012), sewage (Ren et al., 2017) or in-
dustrial wastewater (Lu et al., 2016). The characteristics of waste-
waters were distinct and the valence of iron varied from Fe0, Fe(II)
and Fe(III) in oxide, to ferrates (IV, V and VI). Table 1 summarizes
the chemical methods that have been adopted to remove nitrogen-
containing pollutants coupling with iron. At the same time, the
targets of the processes, the conditions of use, and the achieved
efficiencies are also listed.
3.2. Mechanisms involved in chemical removal of nitrogen with iron
3.2.1. Pertinent chemical reactions equations in chemical removal of
nitrogen coupling with iron
Various oxidation-reduction reactions are involved in the
chemical removal of nitrogen using iron of different valences.
Table S2 provides a list of reaction equations that may appear
during chemical nitrogen removal processes. As illustrated in
Table S2, low valent iron usually acts as reductants while high
valent iron often serves as oxidants. For example, Fe0 (Lee et al.,
2016) and Fe2þ (Tai and Dempsey, 2009) are used for the reduc-
tion of NO 
3 and NO2 and their end products includes N2, N2O, NH4
þ
Fig. 1. The schematic model of the electron transfer in the denitrification of water treatment referring to redox reactions among five main N-containing substances and iron in
various valences.
55
and NO; NO can be gradually oxidized to NO 3 by ferrates(VI, V,
and IV) (Sharma, 2011) while ammonia can be directly oxidized to
N2 or NO3 by ferrate(VI) (Sharma et al., 1998).
3.2.2. Electron transfer
First of all, electron transfer can be accelerated by changing the
physical or chemical properties and structures of iron-based ma-
terial. One study indicated that in the presence of Fe2þ, the elec-
trical resistance (103 Um) of generated Fe3O4 is so much lower
than Fe(III) oxides (101-104 Um) that it can function as a semi-
conductor and do good to the electron transfer from Fe0 to nitrate,
contributing to the efficiency improvements of the electron trans-
fer (Xu et al., 2012). Researchers also confirmed that galvanic cells
(Fe-Cu cells) formed was capable of accelerating the electron
transfer (Ren et al., 2016). Research results showed that the inter-
facial electron transfer between the species and NO 3 can be
accelerated through the spontaneous electron transfer between the
Fe2þaq and surface iron oxides (Han et al., 2016). Surprisingly, the
Fe0 þ Cu0 system could change the rate and path of electron
transfer (Ren et al., 2016).
In the second place, the nitrogen removal is closely related to
chemical reactions, especially oxidation/reduction reactions, thus
electron transfer makes a great difference in this process. Fig. 1
shows the schematic model of the electron transfer in the deni-
trification of water treatment referring to redox reactions among
five main N-containing substances and iron in various valences.
Iron acts as an electron donor in most cases, while iron also plays
the role of electron acceptor in some situations. Significantly, a
direct reduction in which NO 3 receives electrons directly from an
iron corrosion product layer (magnetite), rather than indirectly via
56 H. Liu et al. / Chemosphere 204 (2018) 51e62
H2 gas produced by the reaction between Fe0 and an acid was
confirmed (Suzuki et al., 2012a).
3.2.3. Iron as catalysts
For one thing, iron itself can play the role of catalyst. HFO has
been proven to be a catalyst during the nitrite reduction with the
co-existence of dissolved and solid-bound Fe(II) (Tai and Dempsey,
2009). nFe0/MAS was synthetized to improve the effectiveness of
nFe0 to prevent their agglomeration so as to provide a larger
accessible surface area and then elevate the catalytic performance
(Lee et al., 2016).
The removal of contaminates containing nitrogen is an intricate
process, during which many kinds of chemical reactions happen
and varieties of products were yield with undesired products
included. Take nitrate reduction by nFe0 as an example, almost all
the nitrate is converted to the undesired ammonia (NH3-N) rather
than the nontoxic gas N2 (Shi et al., 2016). Fe can act as the potential
support materials to provide appropriate sites for the deposition of
bi- or tri-metals (Jung et al., 2014). The nitrogen removal using nFe0
bimetallic system (Hamid et al., 2015) or coated nanoparticles
system (An et al., 2010b) captured much attention because of its
selectivity toward nontoxic products. Interestingly, in the pre-
magnetization Fe0/(Fe/Cu) system, the higher nitrate concentration
at the surface of Fe0 or catalysts attracted by the gradient force
benefits the generation of N-N recombination with the existence of
a gradient force and a Lorentz force (Prüsse and Vorlop, 2001; Jiang
et al., 2015), leading to better selectivity towards nitrogen gas (Ren
et al., 2017). Table S3 shows the catalysts with Fe-containing ma-
terials as support and their selective efficiencies. It can be easily
seen from Table S3 that the selectivity has much to do with reaction
conditions, such as pH, time and the constituents of the catalyst.
Simultaneously, the catalysts are mostly transition metals,
including bimetal and trimetal system.
4. Iron applied in biological or bio-chemical coupled nitrogen
removal
4.1. Pathways and processes of biological or bio-chemical combined
nitrogen removal processes coupling with iron
Many kinds of biological or bio-chemical coupled pathways and
processes of nitrogen removal combining with iron have been used
in considerable studies, including both traditional denitrification
process and emerging sewage denitrification processes. The path-
ways and processes that have been widely used by researchers are
as follows:
AnFe0 and hydrogenotrophic integrated denitrification system
(nFe0-HIDs)
In the integrated denitrification system, it has been verified that
the competition exists between the iron nanoparticles and the
bacteria during the first step of the denitrification process. Also Fe0
was proved to have both the biostimulatory effect associated with
H2 produced in the anaerobic corrosion process and the inhibitory
effect upon bacteria as a result of the competition for nitrate (An
et al., 2010a). Besides, hydrogenotrophic denitrifying bacteria can
also be used to decrease the ammonium generation and remove the
nitrate completely (An et al., 2009).
AAnaerobic ammonium oxidation (ANAMMOX) coupling
with iron
In recent years, a unique pathway of ammonia oxidation per-
formed by ANAMMOX bacteria under the condition of limited
oxygen has been reported, during which the high concentration of
ammonia was oxidized by using nitrite as the electron acceptor
(Park et al., 2009). Furthermore, ANAMMOX coupled with ferric
iron reduction, termed Feammox, was a newly discovered nitrogen
cycling process (Li et al., 2015; Bao and Li, 2017). The mechanisms
involved in the dissimilatory Fe (III) reduction were revealed by
adopting isotope tracing and 16S rRNA gene based amplicon
sequencing techniques (Zhou et al., 2016). In addition, ferrous iron
has been applied in accelerating the start-up of the AANMMOX
process (Bi et al., 2014).
ASimultaneous nitrification, denitrification and phos-
phorus removal process (SNDPR)
During the SNDPR process, a transitory decrease of the nitrogen
removal would appear at the beginning of each operating cycle.
After serval cycles, the removal of nutrients would no longer be
inhibited by Fe3þ. As iron was continuously added in the reactor,
the characteristics of sludge as well as the effect of nitrogen
removal would be improved especially when the Fe load was over
40 mg L1 (Jia et al., 2016).
AFe(II)-mediated autotrophic denitrification (Fe(II)-MAD)
The Fe(II)-MAD process is an alternative biotechnology that can
remove nitrate and recover iron simultaneously through the for-
mation of Fe(III) precipitates. Recently, Fe(II)-MAD has been
obtaining increasing scientific interest over classical heterotrophic
denitrification especially for the treatment of industrial wastewater
that is usually poor of carbon (Kiskira et al., 2016, 2017). In 2017, a
new type of Fe(II)-MAD process that was termed pyrite autotrophic
denitrification (PPAD) process was developed and alkalinity can be
consumed in the process (Tong et al., 2017a).
AFerrous iron-based chemo autotrophic denitrification (Fe-
CAD)
In the Fe-CAD reactor, sludge was found to be rich in ferrous
iron-oxidizing nitrate-reducing bacteria which include Rhodano-
bacter, Mizugakiibacter, Sulfuricella, Comamonas and Gallionella.
Moreover, in order to improve the performance of the Fe-CAD
reactor, the iron encrustation around microbial cells should be
removed or prevented (Wang et al., 2017) and pH is also a key factor
(Benz et al., 1998).
ANitrite dependent anaerobic methane oxidation (NDAMO)
This process uses methane as an external electron donor. And it
is indicated that the medium containing Fe2þ (10 mM) significantly
stimulate the activity and the growth of NDAMO bacteria, resulting
in large aggregates of the NDAMO bacteria (He et al., 2015).
AFe0 permeable reactive barrier (Fe0 PRB)
Fe0 is used as the reactive media in PRB (Fe0 PRB), which is a
cost-effective in situ technology for the remediation of contami-
nated subsurface regions, referring to organic compounds and
heavy metals. Whereas, a challenging task occurs when the tradi-
tional Fe0 PRB system is used as an effective long-term nitrate
removal pathway, which results from the corrosion product foamed
on Fe0 hindering the effective electron transfer from Fe0 to surface-
bound nitrate. Accordingly, EK/Fe0 PRB were developed by
Thiruvenkatachari et al. (2008) and Suzuki et al. (2012b).
AGreen rust system (GRs)
 H. Liu et al. / Chemosphere 204 (2018) 51e62 57

Fig. 2. The schematic model of nitrogen removal pathways or processes coupled with iron.

GRs has been defined as a mixed ferrous/ferric hydroxides in (Choi et al., 2012). Fe(II) in GRs was known to oxidize both nitrate
which there are assumed structures consisting of alternating and nitrite. An early research by Ruby et al. (2006) confirmed that
positively charged hydroxide layers and hydrated anion layers fougerite was helpful for the bacterial activity and acted as the

Table 2
Summary of processes or pathways applied in nitrogen removal coupled with iron and their treatment goal or corresponding type of wastewater.

Processes Type of wastewater or the characteristics of processes Pollutants load and conditions Treatment efficiency Ref.
or pathway

nFe0-HIDs Commonly used for treating groundwater and surface The average concentration of NO 3 Bacteria's growth rate increased (OD422 (An et al., 2010a)
water. was 50 mg L1. increased from 0.0016 to 0.0067).

T ¼ 30 C, pHin was 8.0e9.0,
ZVI ¼ 0.5 g L1.
ANAMMOX Wastewater containing high concentration of T ¼ 35  C, average pH ¼ 6.8 mixed The molar ratio of oxidized NHþ
4 and (Park et al., 2009)
ammonium liquor volatile suspended solid reduced Fe3þ was 0.33 M NH4þ/M Fe3þ.
(MLVSS) was 1120 mg L1.
SNDPR Applied in treatment of municipal wastewater to 350 mg L1 COD, 50 mg L1 NHþ 4 eN The yield of N2O can be decreased (Jia et al., 2016)
decrease the emission of N2O. comparing to the control.
Fe(II)-MAD Designed to treat industrial wastewater containing pH ¼ 7.0, nitrate and Fe(II) was 2, Nitrate removal rate was above 60%. (Kiskira et al., 2016,
poor of carbon 10 mM, respectively. And EDTA/Fe(II) 2017)
ratio of 2.0.
Fe-CAD Used for treatment of low C/N ratio wastewater. T ¼ 35  C, HRT ¼ 16 h, the volumetric Volumetric removal rate (VRR) (Wang et al., 2017)
loading rate of (VLR) NO 3 and Fe
2þ
increased more than 20 times and the
was 0.26 kg-N/(m3/d) and 1.67 kg-Fe/ bacteria have been enriched.
(m3 d1).
NDAMO Bacteria grow slowly and the activity of cell is low. 20 mM of Fe(II) and 10 mM of Cu(II) The activity and the growth of NDAMO (He et al., 2015)
bacteria can be significantly stimulated

GRs Usually being used to reduce nitrate by limiting the T ¼ 30 C, initial nitrate Selectivity of ammonium has a close (Etique et al., 2014)
formation of ammonium. concentrations varied from 0.81 to relationship with pH, Cl, F, and SO2
4
3.23 mM, NO 3 /Fe(II) was between anions.
0.48 and 7.7.
Fe0 PRB Suitable for remediating contaminated groundwater; NO 3 -N ¼ 40 mg L
1
, pH ¼ 7.0, Nitrate could be removed more than (Thiruvenkatachari
converting toxic nitrate into less toxic substances and DO ¼ 6 mg L1, flow rate of 94%. et al., 2008; Suzuki
1
recovering the dissolved reaction products. 235 mL d . et al., 2012b; Liu
et al., 2013)
1 þ þ
Fe0-C Developed for high nitrogen load and low carbon COD ¼ 15 mg L , NH4 NH4 -N and total nitrogen (TN) removal (Deng et al., 2016)
MEBD source wastewater treatment eN ¼ 10 mg L1, NO 3 ¼ 15 mg L
1
efficiencies of 92.6% and 95.3%,
DO ¼ 1.0 ± 0.1 mg L1, HRT ¼ 3 h, and respectively. And functional bacteria
the optimal pH from 7.6 to 8.4. increased significantly.
BES More efficient for utilizing limited carbon sources in T ¼ 35  C, HRT ¼ 24 h, Exoelectrogen species was enriched, (Zhu et al., 2015)
1 
low C/N wastewater COD ¼ 2000 mg L , NO3 -N from 200 enhancing the denitrification.
1
to 375 mg L to achieve C/N ratio
from 3.75 to 2.
Fe-graphite plate electrodes, applying
direct current of 0.5 V.
NAFO Organic affects little; the pH of effluent showed strong T ¼ 30  C, anaerobic condition, initial 30% of nitrate was reduced and 85% of (Zhang et al., 2014b,
correlation with nitrogen removal and nitrate Fe(II) concentration of 3.4 mM and Fe(II) was oxidized over 72 h. 2015; Su et al., 2015)
concentration was preferred to be less than 130 mg-N nitrate concentration of 9.5 mM. The maximum nitrate conversion rates
1 1
L ; there were optimal values of Fe/N molar ratio, Optimal values of Fe/N ratio, (qmax) reached 57.89 mg (g VSS d) .
temperature and C/N molar ratio. temperature and pH were 2.0, Nitrate removal efficiency approached
30.15  C and 8.0, respectively. 100%, and capacity of 16.57 mg P/g VS
T ¼ 30  C, pHin was around 7.0, and removal rate of 94.77 ± 2.97%
concentration of effluent was varied
from 32.13 ± 0.43 to
173.54 ± 1.96 mg L1.
58 H. Liu et al. / Chemosphere 204 (2018) 51e62

electron shuttle for nitrate reduction in GRs. Microorganisms were
identified to be able to couple Fe(II) oxidation with nitrite reduction
directly (Pantke et al., 2012).
AFe (0)-carbon micro-electrolysis and biological denitrifi-
cation (Fe0-C MEBD)
With the enhanced denitrification by micro-electrolysis-based
electron donor production, an efficient simultaneous nitrification
and autotrophic denitrification was obtained and high nitrogen
removal load was maintained for low organic carbon water treat-
ment in the Fe0-C MEBD system (Deng et al., 2016).
ABioelectrochemical system (BES)
In BES, the electrode could significantly improve the denitrifi-
cation at low C/N, and Fe further enhanced its performance because
of the combination of anodic oxidation and cathodic denitrification.
What's more, a better effect of nitrate removal was achieved by
means of Fe-C electrodes because more exelectrogenic bacteria
were enriched to enhance the anodic and better drive the occur-
rence of cathodic denitrification the presence of Fe (Zhu et al.,
2015).
ANitrate-dependent anaerobic ferrous oxidizing (NAFO)
process
Although various pathways or processes have been applied
coupling with diverse iron, the essential of them are either bio-
processes or bio-chemical coupled processes. Fig. 2 shows the
schematic model of nitrogen removal pathways or processes
coupled with iron.
Generally speaking, the selection of pathways or processes
usually varies greatly from the type of wastewater. Each kind of
process was selected to treat a certain wastewater or to achieve a
certain treatment goal. Table 2 sums up the foregoing processes or
pathways applied in the denitrification processes coupled with
iron, including their treatment goal or corresponding type of
wastewater, pollutants loads, using conditions and treatment
efficiency.
4.2. Mechanisms of nitrogen removal in biological or bio-chemical
combined processes coupled with iron
4.2.1. Chemical reaction equations in biological or bio-chemical
coupled nitrogen removal processes
Chemical reaction equations in biological or bio-chemical
coupled nitrogen removal processes are provided in Table 3.
4.2.2. Electron transfer
On the one hand, iron directly plays a key role in the electron
transfer processes. A research made by (Jean-ChristopheCle
ment
et al. (2005) showed that NHþ 4 was anaerobically nitrified into

With NAFO bacteria firstly being reported by Straub et al. (1996), NO2 with Fe(III) to accept the electrons instead of O2 under

varieties of NAFO bacterium have been isolated from lake or sedi-
ment, such as Strain 2002 (Weber et al., 2006), strain BoFeN1
(Kappler et al., 2005), Citrobacter Freundii strain PXL1 (Li et al.,
2014) and Pseudomonas sp. SZF15 (Su et al., 2015). They also
have been gradually applied in the remediation of nitrate
contaminated groundwater (Li et al., 2014) and the treatment of
wastewater (including low C/N ratio wastewater) (Zhang et al.,
2014b; Wang et al., 2015). As a valuable biological process,
ferrous iron was utilized to convert nitrite into nitrogen gas so as to
achieve the goal of nitrogen removal in NAFO during this process
(Zhang et al., 2015).
anaerobic conditions. It was also found that complete nitrate
removal in the research of conducting autotrophic denitrification
with pseudomonas sp. SZF15 adopting mixed electron donors un-
der the condition of high Mn(II)/Fe(II) molar ratios at 30  C, pH 7.0
and an electron donor/electron acceptor at the molar ratio of 3.00
(Su et al., 2017). In the NAFO process, there was no cell density
increase when iron's role as an electron donor was omitted (Weber
et al., 2006).
On the other hand, iron is indispensable for influencing the
electron transfer indirectly. A study showed that Fe-containing
substances like nicotinamide adenine dinucleotide (NADH), dehy-
drogenase, SDH, quinone pool (Q), b, c1 complex (Cyt. b, Cyt. c1),

Table 3
Pertinent possible chemical reaction equations in biological or bio-chemical coupled nitrogen removal processes.

Pathways or Pertinent possible chemical reaction equations Ref.

processes

nFe0-HIDs (1) 4Fe0 þ NO 3 þ 7H2O / 4Fe

2þ
þ NHþ 4 þ 10OH

(An et al., 2010a)
(2) Fe0 þ 2H2O / Fe2þ þ H2 þ 2OH
(3) 0.33NO 3 þ H2 þ 0.08CO2 / 0.16N2 þ 0.015C2H7O2N þ 1.11H2O
ANAMMOX (1) 3Fe(OH)3 þ 5Hþ þ NHþ 4 / 3Fe
2þ
þ 9H2O þ 0.5N2 (Jean-ChristopheCle
ment et al., 2005; Park et al., 2009;
(2) 6Fe(OH)3 þ 10Hþ þ NHþ 4 / 6Fe 2þ
þ 16H2O þ NO 2 Ding et al., 2014)
(3) 8Fe(OH)3 þ 14Hþ þ NHþ 4 / 8Fe
2þ
þ 21H2O þ NO 3
þ 3þ  2þ þ
(4) NH4 þ 2H2O þ 6Fe / NO2 þ 6Fe þ 8H
(5) NH4 þ 1.32NO2 þ 0.066HCO3 / 1.02N2 þ 0.26NO
þ  
3 þ 0.066CH2O5N0.15 þ 2.03H2O
Fe (II)-MAD (1) 10 Fe2þ (aq) þ 2NO 3 (aq) þ 24H2O / 10Fe(OH)3 (s) þ N2 (g) þ 18H (aq)
þ
(Kiskira et al., 2016; Tong et al., 2017b)
(2) 2FeS2 þ 6NO3 þ 4H / 2Fe(OH)3 þ 4N2(g) þ 4SO4 þ 2Hþ
 þ 2

(3) 0.364FeS2 þ NO 3 þ 0.116CO2 þ 0.821H2O þ 0.023NH4 /

þ
þ
0.364Fe(OH)3 þ 0.50N2 þ 0.729SO2 4 þ 0.480H þ 0.023C 5H7O2N
Fe0 PRB (1) NO 3 þ 5CH2O / N2 þ CO2 þ 4HCO
3
þ 3H2O (Liu et al., 2013)
(2) 5Fe0 þ 2NO 3 þ 6H2O þ 5Fe
2þ
þ N2 þ 12 OH
(3) 4Fe0 þ NO þ
3 þ 7H / 4Fe
2þ
þ NHþ 4 þ 10OH


(4) Fe0 þ NO þ

3 þ H / Fe
2þ
þ NO 2 þ 2OH


GRs (1) FeII4FeIII2(OH)12CO3 þ (3/8)NO þ III

3 þ (7/4)H / 5Fe OOH þ HCO3 þ Fe
 2þ
aq þ (25/8)H2O þ (3/ (Ruby et al., 2006)
8)NHþ 4
Fe0-C MEBD (1) Fe0 þ 2H2O / H2 þ Fe2þ þ 2OH (Suzuki et al., 2012a; Deng et al., 2016)
(2) 2.82Fe0 þ NO 3 þ Fe
2þ
þ 2.25H2O / NHþ 4 þ 1.19Fe3O4 þ 0.50OH


(3) H2 þ 0.35NO þ
3 þ 0.35H þ 0.052CO2 / 0.17N2 þ1.1H2O þ 0.010C5H7O2N
Fe-CAD (1) 2NO 3 þ 10Fe
2þ
þ 24H2O / 10Fe(OH)3 þ N2 þ 18Hþ (Wang et al., 2017)
NAFO (1) Fe2þ þ 2NO 3 þ 24H2O / 10Fe(OH)3 þ N2 þ 8H
þ
(Zhang et al., 2015)
 H. Liu et al. / Chemosphere 204 (2018) 51e62
Fig. 3. The schematic diagrams of Fe-containing enzymes or functional substances in the canonical electron transport system.
and cytochrome c (Cyt. c) are an important part for the canonical
electron transport system (Chen and Strous, 2013; Wan et al., 2016).
From the electron transfer prospective, no iron or an inappropriate
content of iron is obviously detrimental.
4.2.3. Effect on activity of enzymes
Enzymes play a crucial role in biological reaction processes.
Denitrification is carried out by a series of reductases, such as ni-
trate reductase (NAR), nitrite reductase (NIR), NO reductase (NOR),
and N2O reductase (N2OR) (Jia et al., 2016). Moreover, iron exerts a
critical impact on the synthesis and activity of enzymes and tran-
scriptional regulation. Iron has been considered to be helpful for
the synthesis of NOR (Jia et al., 2016). It has already been authen-
ticated that Fe3þ participates in the synthesis of NAR, NIR and
dehydrogenation coenzymes (Fermoso et al., 2009). Owing to the
existence of Fe-containing active site or Fe-S clusters (Berks et al.,
1995), iron does not only play an important part in lots of en-
zymes like above stated NOR, NIR, NOR and particularly those
related to the electron transfer and consumption in denitrification
(Chen and Strous, 2013), but also impacts Fe-containing proteins in
the electron transfer chain, such as flavoprotein and succinate de-
hydrogenase (SDH) (Massey, 1995; Yankovskaya et al., 2003). Take
NOR as an example, it is a kind of iron enzymes with nonheme-iron
as the reaction center (Heiss et al., 1989). Fig. 3 shows the schematic
diagram Fe-containing enzymes or functional substances in the
canonical electron transport system (Chen and Strous, 2013; Wan
et al., 2016).
In ANAMMOX process, heme c-bearing enzyme hydrazine syn-
thesis (HZS) can catalyze hydrazine the production from ammonia
and nitric oxide (Bi et al., 2014). Hydrazine dehydrogenase (HDH)
can contribute to the production of nitrogen gas, provide the
electrons for nitrite reduction and hydrazine synthesis (Kartal et al.,
2012), and convert N2H4 into N2 (Kartal et al., 2011). However, it has
been deemed that chelating ferrous iron is required for the syn-
thesis of heme c to form active regions (Bi et al., 2014). Besides,
appropriate addition of iron can enhance the activity of enzymes,
and an excessive content of iron will inhibit the activity of enzymes
and destroy the structure of microbes. For instance, nitrogen
removal efficiency was improved when Fe3þ load was 40 mg L1,
59
but the adverse result appeared when the Fe3þ load was 60 mg L1
in the simultaneous nitrification, denitrification, and phosphorus
removal process (Jia et al., 2016). However, 0.4 mg L1 has been
considered as the turning point in the ANAMMOX process (Chen
et al., 2014). So it is reasonable for researchers to pay more atten-
tion on mechanisms that how does iron influence the enzymes
activity and what is the amount of iron that should be added in
various biological nitrogen removal processes.
4.2.4. Effect on microorganism and microbial community
It is undisputed that the addition of iron makes a great differ-
ence in multifarious nitrogen removal processes or methods. First
of all, iron has a great impact on microorganisms. As it is described
above, iron is an important electron donor or acceptor. Iron's
function of electron transition can be negligible under certain cir-
cumstances, which however, could enrich the exoelectrogenic
bacteria during the nitrogen removal processes (Zhu et al., 2015). In
addition, it was reported that the activity and the growth of the
NDAMO bacteria can be significantly stimulated in the presence of
an appropriate content of iron and copper (He et al., 2015). How-
ever, precipitation of iron hydroxides or oxides on the microor-
ganism surface leads to cell inactivation and blocks the pore of
substance exchange. It even may damage the structure of cell and
disturbs the material balance and energy exchange between the
intracellular and extracellular, and then affects the activity of en-
zymes and electron transfer (Gao et al., 2014; Wang et al., 2017).
The membrane disruption, DNA damage and protein (enzyme)
inactivation resulting from the toxicity of n Fe0 have also been re-
ported (Zhang et al., 2017). A study proved that the activity of
ANAMMOX can be strengthened in the presence of a reasonable
concentration of Fe2þ and Fe3þ by maintaining high
oxidationereduction potential (ORP) and gathering acylhomoser-
ine lactones (AHLs) which is thought of as a vital substance for cell-
to-cell communication. Iron can promote the growth of ANAMMOX
bacteria by participating in the synthesis of heme c, which is
essential for energy metabolism in ANAMMOX bacteria (Chen et al.,
2014). Nevertheless, an excessive Fe2þ and Fe3þ hindered the mass
transfer process of the ANAMMOX bacteria and even resulted in its
death due to the free radical cytotoxicity induced by Fenton and
60 H. Liu et al. / Chemosphere 204 (2018) 51e62
Like-Fenton reactions (Gao et al., 2014).
Subsequently, the significant role that iron plays in microbial
communities cannot be ignored. According to the analysis of the
average well color development, three different biodiversity indices
(The Shannon index, the Pielou index and the Simpson index) as
well as the principal component analysis (PCA), Fe2þ were
confirmed to be able to enhance the microbial activity and then
increase the nitrogen removal efficiency. It was also found that Fe2þ
change the composition of microbial community and that the effect
of Fe2þ on microbial functional diversity has something to do with
whether to add organic source (Song et al., 2016). In the Fe0-C
MEBD system, Genera Hydrogenophaga and Sphaerotilus became
the dominant communities, taking place of other genera, such as
Blastocatella, Nitrospira, Flavobacterium and so on, and Sphaer-
otilus has been considered to be useful for Fe2þ-based denitrifica-
tion (Deng et al., 2016). Compared with control, functional
microorganisms like nitrate-reducing and iron-oxidizing bacteria
(i.e. Phylum Proteobacteria, Mizugakiibacte, Gallionella, Sulfur-
icella, Rhodanobacter and Comamonas) were extremely enriched
to be dominant genus in the sludge of Fe-CAD process (Wang et al.,
2017). Inexplicably, ANAMMOX bacteria (Candidatus Kuenenia) can
cope with the nFe0 range of 10e200 mg L1 and keep its dominance
even though microbial composition was affected to some extent
(Zhang et al., 2017).
Obviously, the effect of iron on microorganisms and microbial
community shows a great discrepancy among varieties of nitrogen
removal processes, including both positive and negative ones.
5. Conclusions, challenges, and future prospects
5.1. Conclusions
In this review, the conclusions can be made in the following
three points:
(1)As a prevalent and inexpensive material, various valences of
iron and their diverse morphologies have been applied in
Fig. 4. The schematic model of four main mechanisms that iron applies in water treatment for nitrogen removal.
nitrogen removal. Different valences of iron, from Fe0 to Fe(VI),
have been proverbially used in the nitrogen removal of many
kinds of water and wastewater treatment. Iron has also been put
into application by varieties of preparation and forms, modified
by physical or chemical methods, joint use of diverse valent of
iron, Fe0 in different sizes (i.e. iron powder, iron craps, cast iron/
filings, mFe0, nFe0 etc.), loaded or loading with other substances,
supporting mono- bi- or multi-metal, like transition metals and
so on.
(2)The chemical nitrogen removal methods, traditional biolog-
ical or bio-chemical coupled nitrogen removal processes and
emerging ones and their use conditions that have been widely
investigated or used have been summarized. In addition, the
wastewater type, pollutant load and treatment efficiency of
those chemical nitrogen removal methods and the biological or
bio-chemical coupled nitrogen removal processes have been
also been summed up.
(3)From the perspective of mechanism, pertinent possible
chemical reaction equations referring to multifarious chemical
and biochemical coupled pathways or processes have been
summed. Four main mechanisms were analyzed and elucidated
(depicted in Fig. 4).
5.2. Challenges and future prospects
There is no doubt that iron plays a crucial role in nitrogen
removal. At the same time, challenges or drawbacks are inevitable
during the nitrogen removal process. Conditions that iron works in
denitrification systems coupling with iron, such as pH and co-
existence substrate, make a great difference. Surface passivation
is a key limiting factor for nFe0 involved technology of for water
remediation. Although some common oxidants, surface proton-
ation and ligands have been investigated to address the problem, it
was also still in the stage of research and experiment. Furthermore,
the appearance of some undesired products hinders the exchange
of substances and electron transportation. For example, the ferric
 H. Liu et al. / Chemosphere 204 (2018) 51e62
oxide formed on the cell strongly affects the exchange between
inner and outer cell of bacteria. Thus, the selection of appropriate
iron-based material in denitrification process for different type of
wastewater is required to find the countermeasures, as well as the
researches looking into the underlying biochemical mechanisms,
i.e. the effect of iron on the activities of various enzymes, electron
transport and microbial community.
Acknowledgements
This work was co-supported by the Bill and Melinda Gates
Foundation RTTC-China 2014 project and Projects of International
Cooperation Shanghai (STCSM, 18230712300). The authors are very
grateful to Dr. Stephan Kueppers from Juelich Research Center,
Germany and Prof. Cong Li from Zhejiang University, China for their
help in refining language of the manuscript.
Appendix A. Supplementary data
Supplementary data related to this article can be found at
https://doi.org/10.1016/j.chemosphere.2018.04.019.
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