Journal of Hazardous Materials 384 (2020) 121505

Contents lists available at ScienceDirect

Journal of Hazardous Materials

journal homepage: www.elsevier.com/locate/jhazmat

Enhanced passivation of lead with immobilized phosphate solubilizing T

bacteria beads loaded with biochar/ nanoscale zero valent iron composite
Zedong Tenga,b, Wen Shaoa,c, Keyao Zhanga,b, Fulu Yua,d, Yaoqiang Huoa,e, Min Lia,b,*
a
College of Environmental Science and Engineering, Beijing Forestry University, Beijing 100083, China
b
Beijing Key Lab for Source Control Technology of Water Pollution. Beijing Forestry University, Beijing 100083, China
c
Department of Geoscience, Eberhard Karls Universität Tübingen, Tübingen 72074, Germany
d
School of Chemical Science and Engineering, Tongji University, Shanghai 200940, China
e
Department of Environmental Science and Engineering, Fudan University, Shanghai 200438, China

GRAPHICAL ABSTRACT

ARTICLE INFO ABSTRACT

Editor: Daniel CW Tsang Phosphate solubilizing bacteria (PSBs) can effectively enhance the stability of lead via the formation of insoluble
Keywords: Pb-phosphate compounds. This research presents a bio-beads, which was implemented with the help of a self-
Phosphate solubilizing bacteria designed porous spheres carrier, by immobilized PSBs strains Leclercia adecarboxylata (hereafter referred as L1-
Passivation 5). In addition, the passivation efficiency of lead via bio-beads under different conditions and its mechanism
Lead were also investigated in this study. The results indicated that phosphate solubilized by bio-beads could reach
Biochar 30 mg/L in Ca3(PO4)2 medium containing 1 mM Pb2+, and the highest removal rate of Pb2+ in beef peptone
Nanoscale zero valent iron liquid medium could reach 93%, which is better than that of free bacteria. Furthermore, it was also concluded
that the lead could be transformed into stable crystal texture, such as Pb5(PO4)3Cl and Pb5(PO4)3OH. Both
hydrophobic and hydrophilic groups in the bio-beads could capture Pb2+, which indicated that electrostatic
attraction and ion-exchange were also the mechanism of Pb2+ adsorption. All the experimental findings de-
monstrated that this bio-bead could be consider as an efficient way for the lead immobilization in contaminated
soil in the future.

⁎
Corresponding author at: College of Environmental Science and Engineering, Beijing Forestry University, 35# Qinghua East Rd., Haidian District, Beijing 100083,
China.
E-mail address: minli@bjfu.edu.cn (M. Li).

https://doi.org/10.1016/j.jhazmat.2019.121505
Received 24 August 2019; Received in revised form 3 October 2019; Accepted 18 October 2019
Available online 24 October 2019
0304-3894/ © 2019 Elsevier B.V. All rights reserved.
Z. Teng, et al.
1. Introduction
Lead accumulated in soil was originated from solid waste disposal,
wastewater irrigation, pesticide application and atmosphere deposition,
threatening the health of animals and humans through food chains (Niu
et al., 2013). Lead ion binds strongly to albumin protein in body
(Ayranci and Duman, 2004) and changes the functions of protein. It is
well known that high levels of exposure can cause problems that jeo-
pardize the kidneys, gastrointestinal tract, joints and reproductive
system, and cause acute or chronic damage to the nervous system,
especially for children (Park et al., 2011; Chen et al., 2016a). Therefore,
the effects of lead pollution on the environment are significant, and
remediation of lead-contaminated soils is significant to improve the soil
environment. Up to now, various methods have been applied to at-
tenuate the lead contamination, such as chemical precipitation, ad-
sorption and bio-treatment (Yuan et al., 2017a; Wang et al., 2018,
2019a). Bioremediation technology was widely concerned by many
researches due to its economically feasibility, easier application in
contaminated sites and less secondary pollution (Oh et al., 2016).
Microbial remediation strategy was suggested as a promising tech-
nology for heavy metal contamination (Park and Bolan, 2013; Manasi
et al., 2014). Previous researches have reported that various micro-
organisms have high Pb tolerance and biosorption capacity (Edris et al.,
2013). Phosphate solubilizing bacteria, as functional bacteria, can re-
lease P from insoluble P rock slowly via the production of organic acids
and phosphatases. Those P released by PSB could reduce Pb mobility
and bioavailability via forming of stable Pb compounds such as pyr-
omorphite (Pb5(PO4)3Cl), hydroxypyromorphite (Pb10(PO4)6(OH)2)
and fluoropyromorphite (Pb10(PO4)6F2) (Park et al., 2011; Zhu et al.,
2018). Lead immobilized by P compounds has been studied in waste-
water and soil (Chen et al., 2007; Li et al., 2017a; Shen et al., 2018),
and the effects of microorganisms on these processes, such as pre-
cipitation and adsorption, were also concerned in recent years (Zeng
et al., 2017).
Although phosphate solubilizing bacteria (PSB) was used to re-
mediate lead contaminated soil, high concentration of lead could in-
hibit the activities of PSB, especially in free cells (Yuan et al., 2017b). In
order to enhance the tolerance of microbial to high concentration of
heavy metals, immobilized microorganism was explored for many re-
searchers. Comparing with free cells, microbial immobilization on
carrier materials has unique superiority, which can protect micro-
organisms from harmful substance and maintain high microbial activity
and stability (Kim et al., 2016; Zhang et al., 2016; Bai et al., 2018).
Furthermore, PSB was immobilized on composite carrier materials
which support the growth of the bacteria communities and provide
them an ideal habitat, in close their nature environment may be helpful
for enhancing the efficiency of phosphorus dissolution and Pb im-
mobilization (Zhang et al., 2017a; Li et al., 2018a). Bacillus sp. with the
capacity of biomineralization can convert the organic phosphorus into
phosphate in the process of metabolism, which can remove Pb2+ from
the solution in the form of coprecipitation Pb5(PO4)3OH (Zhang et al.,
2019). When the added content of PSB-immobilized biochar (PIB) was
equal to or greater than 800 mg/kg, the PSB could proliferate effec-
tively and the NH4NO3-extractable Pb concentration was decreased to
below 1 mg/kg (Zhang et al., 2017a). Biochar immobilized PSB were
prepared in previous researches: Biochar and liquid growth medium
were mixed and inoculated with PSB, and then grown to log phase and
the inoculated biochar was harvested (Zhang et al., 2017a; Li et al.,
2018a). Biochar can provide a protective niche for PSB and hence re-
duce competition from indigenous microorganisms. Therefore, the
creation of an ideal microbial environment inside an advanced porous
carrier may efficiently enhance the contaminants adsorption and bio-
mineralization.
Furthermore, recent researches showed that engineered structured
nanomaterials are promising in removing contaminants, such as heavy
metals and organic contaminants (Xu et al., 2016a; Zhan et al., 2016;
2
Journal of Hazardous Materials 384 (2020) 121505
Zou et al., 2016). Nanomaterial treatment technology can immobilize
heavy metals (Xu et al., 2016b) and prevent them from entering the
deeper soil layers and groundwater (Mukherjee et al., 2015). Among
these smart nanomaterials, nanoscale zero valent iron (nZVI) and their
composites have attracted global attention due to their fascinating
features, such as high surface area (Hu et al., 2017) and adsorption
capabilities (Gil-Diaz et al., 2017; Xue et al., 2018; Sun et al., 2017;
Yang et al., 2018; Sun et al., 2019). In addition, it can quickly reduce
the toxic effect of lead on PSB and reduce the redox potential of soil,
which can enhance the release of phosphate. However, in order to
prevent aggregations occurring, the nZVIs must be hybridized with a
micro-characteristics carrier. Biochar (BC) has the advantages of large
specific surface area, high porosity, porous structure and strong ad-
sorption capacity (Kookana, 2010; Mohan et al., 2014; Shen et al.,
2017; Lefevre et al., 2018; Li et al., 2018b). Therefore, it is usually
chosen as the ideal carrier. Over the past few years, biochar-supported
nZVI (BC/nZVI) was used to remove heavy metals due to its better
stability and effective immobilized ability (Dong et al., 2017; Zhu et al.,
2017). Highly efficient performance for Pb2+ removal can be realized
by using corn stalk-derived hydrophilic biochar supported nZVI (e.g.,
291.3 mg·g−1 for Pb2+ following pseudo-second order kinetics) (Yang
et al., 2018) and sewage sludge-derived biochar immobilized nZVI (e.g.,
nearly 82% Pb2+ removal within 30 min) (Diao et al., 2018). It can be
proved that multiple interaction mechanisms including adsorption,
precipitation, reduction and complexation are responsible for lead re-
moval by BC/nZVI composites. In addition, several recent studies have
shown that BC/nZVI composites have the potential to control phos-
phate and lead pollution. However, these studies did not examine the
potential of release P by bacteria as a lead passivator in soil, which
mainly depends on the synergistic effect of these materials and phos-
phate solubilizing bacteria.
In order to form a porous spheres carrier and combine BC/nZVI with
PSB together, polymeric matrixes such as polyvinyl alcohol (PVA) and
sodium alginate (SA) was added as embedding materials of immobilized
bacteria due to its cheaper, less poisonous and better biocompatible
(Huang et al., 2016), which can efficiently mitigate Fe0 oxidation and
enhanced the stability of bio-beads (Lv et al., 2013; Yu et al., 2018; Sun
et al., 2018). These attractive features make it useful as a microbial
immobilization carrier to enhance biomineralization efficiency. It is
believed that an intergrated immobilization of PSB with PVA-SA con-
taining BC/nZVI composite is an innovative and promising idea. The
feasibility and efficiency of bio-beads for immobilizing of lead was
explored in this research. This article was mainly focused on: (1) pre-
paration of bio-beads (2) lead removal analysis under different treat-
ment (3) underlying mechanisms. This research is respect to provide a
new way for dealing with soil contaminated by lead.
2. Materials and methods
2.1. Cultivation of phosphate solubilizing bacteria
The strain of PSB, Leclercia adecarboxylata, used in this experiment
was isolated in our laboratory. The species preservation number is
CGMCC No.14561. PSB was cultivated in a 150 mL conical flask at 28 °C
on beef extract peptone medium in a shake incubator for 12 h. The
bacteria concentrations were controlled OD600 = 0.6-0.8 for further
study.
2.2. Synthesis of biochar/nZVI and immobilization of bacterial
Rice hull is among the most abundant types of biomass in China,
where tons of rice hulls remain as a by-product in the rice production
industry. These results are most likely helpful for understanding the
potential contributions of biochar amendments to lead passivation. In
this research, the rice hull biochar was purchased from Nanjing
Forestry University (China). To prepare the BC/nZVI, 3.6 g BC (passed
Z. Teng, et al.
through a standard 70-mesh stainless steel sieve) was firstly dissolved in
a 100 mL 0.08 g/L FeSO4·7H2O solution (The mass ratio of biochar to
iron is 3:1). Then, the solution was transferred to a three-necked flask,
and N2 was purged into the solution for 1 h to remove dissolved O2 with
vigorous stirring. 100 mL 0.5 mol/L NaHB4 solution was added into the
slurry using peristaltic pump (approximately 1 drop/s). After fully re-
acting for 40 min, the composited particles were separated from the
liquid phase. Then, the particles were washed for several times with
distilled water and ethanol, respectively, and finally vacuum-dried
(Dong et al., 2017; Hussain et al., 2017; Wei et al., 2018).
PVA 1788 and SA were purchased from Beijing Chemical works,
China. PVA (5 g, 4.5 g and 4 g) was added to 50 mL distilled water and
heated at 95 °C for 30 min until the PVA was completely dissolved. And
then, SA (1 g, 0.5 g, 0.45 g and 0.2 g) was added to the PVA solution
above, and the solution was cooled to about 35 °C. The mixture was
sterilized before adding bacteria suspension. BC/nZVI (0.75 g, 0.6 g,
0.5 g, 0.4 g and 0.25 g, ultraviolet sterilization) was mixed with 25 mL
PSB culture and added to the cooled PVA-SA solution and mixed
thoroughly. All percentages are calculated with respect to distilled
water. Drops of the resulting mixture were extruded into saturated
boric acid solution containing 3% calcium chloride using peristaltic
pump, and cross-linking reaction for 24 h to form bio-beads. Bio-beads
were next rinsed several times and soaked thoroughly in sterile saline to
remove boric acid and calcium chloride. Ultimately, the bio-beads were
lyophilized and stored at sample bottle for the further experiment (Sun
et al., 2018; Li et al., 2017b). The results only showed the experimental
group which could form bio-beads.
2.3. Experimental design in liquid medium
To explore the phosphate solubilizing ability of bio-beads, the
comparative experiment of five treatments was carried out (1) in-
cubation with biochar (0.4 g/L); (2) incubation with BC/nZVI (0.4 g/L);
(3) incubation with free bacteria (20 mL/L); (4) incubation with bio-
beads (5 g/L) and (5) medium without added anything as a control
(CK). In order to control the mass of BC/nZVI and bacteria was similar
to the content of bio-beads group, 0.4 g/L BC/nZVI and 20 mL/L bac-
teria culture was added. Three replicates were used to avoid experi-
mental error. Bio-beads and free bacteria were added into 150 mL tri-
angle bottle containing 50 mL Ca3(PO4)2 liquid medium and then
incubated for 24 h at 30 °C. The P content was determined by spectro-
photometry at 700 nm on a U-1000 spectrophotometer (T6, China)
(Teng et al., 2018). In order to explore the effect of lead stress on the
ability of bio-beads to solubilize phosphate, phosphate solubilized ex-
perimental was carried out by adding 0.5%, 1%, 2.5%, 5% bio-beads,
respectively. The bio-beads were cultured in 50 mL Ca3(PO4)2 liquid
medium containing 1 mM Pb2+, no lead added experiment was treated
as a control.
In addition, the other experiment was designed to examine the
ability of the bio-beads to remove Pb2+. Three treatments were carried
out (1) incubation with free bacteria cells; (2) incubation with bio-
beads and (3) a control. In order to avoid the effect of Ca3(PO4)2 on
mineralization of lead, replacing the Ca3(PO4)2 medium with beef ex-
tract peptone medium (3 g/L Beef extract, 10 g/L peptone, 5 g/L NaCl)
containing 1 mM Pb2+. Residual Pb2+ concentration was determined
by atomic absorption spectrophotometer (AAS). The influence of dif-
ferent dosages (0, 0.5%, 1%, 2.5%, 5%), different initial Pb2+ con-
centrations (0 mM, 0.5 mM, 1 mM, 3 mM, 5 mM), pH (5, 6, 7, 8, 9) and
the temperature (4 °C, 15 °C, 30 °C) on removal rate of Pb2+ were also
explored. Each batch experiment was carried out under the same con-
ditions, and only a single influencing factor was changed. These results
are discussed in detail in this paper.
2.4. Related characterization of bio-beads
Scanning electron microscope (SEM) was used to characterize the
3
Journal of Hazardous Materials 384 (2020) 121505
porous structure and surface morphology of the bio-beads, and Energy-
dispersive spectroscopy (EDS) was used to determine the surface ele-
mental composition of the bio-beads. In addition, functional group of
lyophilized bio-beads was analysis using a Nicolet 6700 Fourier
Transform infrared spectrometer (FT-IR, Bruker, vertex-70 with an ATR
accessory). The crystalline structure was obtained using a power X-ray
diffractometer (XRD, Shimadzu, XRD-7000) at a scan rate of 2°·min−1.
3. Results and discussion
3.1. Characteristics of biochar, biochar/nZVI and bio-beads
3.1.1. Properties of biochar/nZVI
Nano zero-valent iron was coated on the biochar carrier to play a
dual role i.e. adsorption- passivation. Specific surface area of BC and
BC/nZVI were analyzed using Brunauer-Emmett-Teller (BET), which
can be investigated the suitability of BC/nZVI for adsorption and bac-
teria adhesion. The mean surface area of the BC/nZVI was
116.83 m2·g−1, compared to 33.40 m2·g−1 of BC. The increased surface
area of the composite is likely due to non-aggregation of the nZVI
particles (Kim et al., 2013). The BC/nZVI was composed of well-ar-
ranged network of BC with an average pore size of 5.24 nm, which was
larger than BC in Table 1, enhancing the carrier adsorption properties.
It probably because the structure of pore may be changed by the che-
mical bond between nZVI and biochar.
SEM photographs showed that the biochar looked like the sieve
plates with a polyporous structure (Fig. 1a). Some particulate matter
was loaded on the surface and pores of biochar after load iron (Fig. 1b).
In addition, EDS was carried out to explore the element distribution
characteristics of the reaction products (Fig. 1c). The element mapping
showed that BC was eventually covered by O and Fe elements, which as
a background material, indicating Fe could settle on the surface and
pores of biochar.
The effect of synthesis conditions on the biochar and iron forms was
analysis by XRD and FT-IR. Fig. 2A showed the functional groups of the
BC and BC/nZVI. The FT-IR spectrum showed the peak shifted from
1583.40 cm−1 to 1552.54 cm-1 marking amide II bond NeH bending
and C–N stretching from peptides (Bai et al., 2014), which involve in
the iron binding. The peaks shifted from 1436.82 to 1488.90 cm-1, and
1353.89 to 1321.11 cm-1, indicated that carboxyl groups and C]O
group of the amino acids and amine were involved in the binding of
iron. A significant change in the peak value from 1078.20 cm-1 to
1047.24 cm-1 may be related to the C–O of carboxyl groups, which in-
volved in the iron binding. The key peak at 673.09 cm-1, ascribed to Fe-
O stretching, is observed in the BC/nZVI spectrum (He et al., 2017),
indicating the presence of a chemical bonds between the nZVI and
biochar (Rajapaksha et al., 2016; Yan et al., 2015; Su et al., 2016). This
observation also reflects the present of iron oxide in the composite.
Previous studies have shown that the diffraction peaks at 15-30° and
43-45°were attributed to the present of amorphous organic components
and α-Fe0, respectively (Qian et al., 2017; Gong et al., 2018). In this
research, the diffraction peak (2θ) at 22° exists in both BC and BC/nZVI.
By observing and analyzing the characteristic peak existing at ap-
proximately 44.9°for BC/nZVI in Fig. 3B, it was corresponded to the
formation of ZVI.
Table 1
Brunauer-Emmett-Teller (BET) specific surface area of biochar.
Carrier SBET(m2· g−1) Vtotal(cm3· g−1) Average pore size (nm)
BC 33.40 0.032 1.92
BC/nZVI 116.83 0.306 5.24
SBET, indicates BET specific surface area, Vtotal, indicates total pore volume.
Z. Teng, et al. Journal of Hazardous Materials 384 (2020) 121505

Fig. 1. The scanning electron microscope (SEM) images of: (a) biochar, (b) BC/nZVI and (c) EDS images with element mapping of BC/nZVI; The energy dispersive
spectrometer (EDS) images with element mapping of the products after load iron.

3.1.2. Preparation and optimization conditions of bio-beads

PVA and SA has been selected for synthesizing the outer-layer
porous carrier composite. At the same time, biochar was used as the
carrier material of zero-valent iron, providing a good inner barrier for
L1-5 biomineralized lead. The optimized embedding condition was
morphology, such as particles, shape, mass and so on. The key three
parameters were concentrations of PVA, SA and BC/nZVI. As showed in
Table 2, the bio-beads gradually became larger with the increasing of
PVA content. However, the excessive content of PVA caused the me-
chanical strength of the bio-beads to become lower, while the viscosity
of the mixed gel increased, making the preparation of bio-beads more
difficult. The mechanical strength of the bio-beads was getting better
Fig. 3. The appearance and morphology of immobilized bio-beads.
and better with the increasing of SA content, but the excessive SA
content would make the tailing of the bio-beads more serious. In ad-
dition, the solution would become sticky and not easy to drip out, and optimal composition of bio-beads was the mass fraction of 9% PVA, 1%
the shape of the bio-beads was irregular when the BC/nZVI content was SA and 0.8% BC/nZVI, respectively. The immobilized beads prepared
too high. In summary, only the concentration of the three factors (PVA, according to the optimum amount of the material was shown in Fig. 3,
SA, BC/nZVI) that affected the structural properties of the bio-beads they were almost spherical with a diameter approximately 6 mm.
were within the appropriate range, a perfect bio-beads can be formed.
After comprehensive comparative analysis, the results showed that the 3.1.3. Characteristics of bio-beads
FT-IR spectrums of the bio-beads was showed in Fig. 4A, rich

Fig. 2. Characterization of the synthesized materials: (A) Fourier transform infrared spectroscopy of (a) BC and (b) BC/nZVI; (B) X-ray diffractometer patterns of (a)
BC and (b) BC/nZVI.

4
Z. Teng, et al. Journal of Hazardous Materials 384 (2020) 121505

Table 2
Comparison of the characteristics of bio-beads.
Experiment PVA SA BC/nZVI Bio-bead morphology
(%) (%) (%)

1 10 1 0.5 Larger particles, not round, more broken

2 10 1 1 Smaller, brittle, and 1/3 broken
3 10 2 1 Not broken, but tailing
4 9 1 1 Forming, good flexibility, mostly floating
5 9 1 0.8 Forming, good flexibility, mostly sinking
6 8 1 1.5 Tailing severely, irregular shape, not
sinking
7 8 1 1.2 Tailing severely, irregular shape
8 8 1 0.8 Tailing severely, irregular shape
9 8 0.9 0.8 Tailing weakened, shape improved.
10 8 0.4 0.8 Soft and tailing severely

function groups (such as −OH, −CH2, C]O, C–N, PO2− and C–O-C)
were found in surface of bio-beads. The XRD pattern of bio-beads as
presented in Fig. 4B. The diffraction peaks at 44.67 and 64.68° (2θ)
were corresponded to the formation of iron in its zero-valent form. The
characteristic peaks of 2θ at 31.74°, 56.44° and 66.17°, corresponded to
the formation of Fe(OH)2. As shown by the SEM images in Fig. 4C and
Fig. 4D, the bio-beads had a rough and relative dense surface (Fig. 4C),
improving its stability and capability for avoiding the violation from
heavy metals. In addition, the BC/nZVI particles were embedded in the
surface of the bio-beads. The inside of the bio-beads formed a mesh-like
porous with large and small pores (Fig. 4D), providing a convenient
channel for matter transmission.
Fig. 5. Phosphorus solubilized by free bacteria, BC, BC/nZVI and Bio-beads in
Ca3(PO4)2 liquid medium.
3.2. Phosphate solubilizing ability and lead passivating capacity of bio-
beads
3.2.1. Phosphate solubilizing ability of bio-beads
To evaluate the performance of the bio-beads, the phosphate solu-
bilizing ability of free bacteria, BC, BC/nZVI and bio-beads were ob-
tained. As showed in Fig. 5, the concentration of soluble phosphate in
the free bacteria and the bio-beads group was significantly higher than
that in the CK, the BC and the BC/nZVI group, indicating that phos-
phate solubilizing bacterial still had the phosphate solubilizing capacity
after immobilization, and the bio-beads had significantly enhanced the

Fig. 4. Characterization of the bio-beads: FT-IR spectrum (A) and XRD pattern (B) of bio-bead; SEM images of surface (C) and inner structure (D) of bio-bead.

5
Z. Teng, et al.
Fig. 6. Effect of different dosage of bio-beads on P-solubilized and pH in
Ca3(PO4)2 liquid medium in absence and presence of 1 mM Pb(NO3)2.
phosphate-solubilizing capacity, reaching more than 200 mg/L. It might
be because there were numerous bacteria in the bio-beads, which can
increase the stability and activity of cells and enhance biological
functions of bacteria, and produce more organic acids and phospha-
tases, compared with free bacteria group (Zhang et al., 2017b; Chen
et al., 2019). At the same time, the porous structure of bio-beads pro-
vided a provides a pathway for the migration of PSB and nutrients.
The bio-beads dosage plays an important role in the process of
phosphorus solubilizing. The concentration of soluble phosphate in the
lead-free medium decreased gradually with the bio-beads dosage in-
creased, while it was increased gradually in the medium containing
lead (Fig. 6). The reason may be that most of the soluble phosphate was
absorbed by the bio-beads in the culture medium without lead. For
instance, the PSB can consume the mobile P in medium, which results
in a lower P content after increase bio-beads addition (Chen et al.,
2019). In addition, previous studies have shown that biochar has a
strong adsorption capacity for phosphate (Rajapaksha et al., 2016;
Ngatia et al., 2017). The amount of biochar increased with the in-
creasing of bio-beads dosage, which can enhance the adsorption of
soluble phosphate, leading to the low content of soluble phosphate.
However, most of the soluble phosphate in the culture medium con-
taining lead formed precipitation with lead (Chen et al., 2019), and the
precipitation was largely absorbed by the bio-beads, thus preventing
the bio-beads from adsorbing the remaining small amount of soluble
phosphate in the solution. Therefore, the soluble phosphate content in
the solution showed an increasing trend with the increase of bio-beads
dosage.
In this research, pH played a key role in the process of phosphorus
solubilizing. Previous studies have shown that a significant negative
correlation was observed between pH and phosphate solubilized con-
centrations (Teng et al., 2019a). The pH value of samples in absence
lead medium was 7.23 before sterilization, after addition lead, the pH
value was slightly changed to 6.83 (not showed in this research). After
24 h incubation, all treatments pH values with bio-beads addition were
evaluated (Fig. 6). The pH was maintained at about 4 in the lead-free
samples, while it was tended to decrease with bio-beads dosage in-
creasing in lead present sample. It may be that the high bio-beads do-
sage can immobilize amount of Pb2+ in solution, which reduces the
toxic effects on PSB and enhances their metabolic capacity. But anyway,
pH values were decreased compared to the initial value in all samples,
which can be attributed to the ability of microorganisms to produce
organic acids when decomposing organic matter (Li et al., 2016a, b).
3.2.2. Lead passivation capacity of bio-beads
To investigate the functions of lead immobilization and the activity
6
Journal of Hazardous Materials 384 (2020) 121505
Fig. 7. Effects of bio-beads on lead immobilization in beef extract peptone
medium containing 1 mM Pb2+.
of the bio-beads, a batch experiment was performed in a flask for 24 h.
The results presented here, clearly indicating strong immobilization
ability of bio-beads due to removal 70% of Pb2+. Very surprisingly,
straightforward comparison the immobilization of lead between free
bacteria and bio-beads demonstrated that the bio-beads could sig-
nificantly improve the ability of PSB to remove lead ions. In preliminary
experiment, BC/nZVI showed a higher Pb2+ removal percentage than
BC, i.e., 84.54% vs. 1.33% (data not shown). BC/nZVI has higher
capability of sorption, together with its more abundant P content (Chen
et al., 2019). PVA and SA could prevent lead ions from contacting PSB
directly and reduce the toxic effect of Pb2+ on PSB (Ali et al., 2015;
Karim et al., 2019), which provided a more appropriate way to attract
PSB to immobilized lead (Fig. 7).
3.2.3. Effect of different factors on lead passivation
In order to further improve the removal capacity of bio-beads to
lead ions, the reaction conditions were optimized. As expected, the
Pb2+ removal rate was increased with the dosage increasing (Fig. 8A).
When the dosage was 5% (2.5 g bio-beads in 50 mL medium), Pb2+
removal rate can reach above 80%. In order to consider economic costs,
the dosage of bio-beads in subsequent experiments was controlled at
5%. The effect of initial Pb2+ concentration on the adsorption capa-
cities of Pb(II) are illustrated in Fig. 8B. The removal efficiency in-
creases with the initial concentration increasing from 0.5 mM to 5 mM
with the maximum removal efficiency of 93%, and then it decreases as
the initial concentration increases further. This might be attributed to a
larger driving force that promotes mass transfer between solid and
aqueous phase at higher Pb(II) concentration (Chen et al., 2016b). But
excessive high concentration of lead inhibits the activity of phosphate
solubilizing bacteria (Li et al., 2016a). The initial pH significantly af-
fected the speciation of lead and surface active sites of the bio-beads.
When the initial pH ranged from 5 to 9, the removal rate of lead ion was
first decreased and then increased, and the maximum sorption capacity
is at pH 5. This phenomenon could be explained that Pb2+ was exists in
the form of PbOH+ when pH was higher than 5, and this form was not
easier transformed by bio-beads comparing to Pb2+ (Wang et al.,
2019b). However, too low pH could inhibit the microbial activity
causes the adsorption process to reverse. Therefore, the optimal pH
value for the passivation of lead by bio-beads was selected as pH 5 in
this research. Fig. 8C showed that the temperature effect on the re-
moval of lead was not significant, but also with the rise of temperature
increasing trend. This phenomenon can be explained as follows: the
growth and metabolism ability of PSB is enhanced with the increase of
temperature, which can solubilize more phosphate, thereby enhancing
the passivation of Pb2+. Based on comprehensive analysis, when the
dosage is 5%, the initial lead concentration is 1 mM, the pH is 5, and the
temperature is 30 °C, the Pb2+ removal rate can reach 93%. As shown
Z. Teng, et al. Journal of Hazardous Materials 384 (2020) 121505

Fig. 8. Effect of different (A) bio-beads dosages; (B) initial lead concentration; (C) initial pH; (D) temperature on lead immobilization in beef extract peptone
medium.

Table 3
Comparison of Pb(II) removal capacities with other materials.
Materials pH Time (h) Initial concentration of Pb2+ (mg/L) Removal rate of Pb2+ (%) Adsorption capacity (mg/g) Ref.

a
RB +PSB 8.71 72 1000 24.11 4.84 (Chen et al., 2019)
SBb+PSB 8.64 72 1000 60.85 12.17 (Chen et al., 2019)
Spongy microbe-based biosorbents 5.00 1 100 61.54 354.02 (Wang et al., 2019b)
Z-NZVIc 6.00 10 100 58.32 85.37 (Li et al., 2018c)
Phosphate-embedded beads 4.00 24 200 94.2 236.16 (Wang et al., 2016)
PSB beads 5.00 24 200 93 27.50 This study

a
An alkaline biochar produced from rice husk.
b
Slightly acidic biochar produced from sludge.
c
Zaeolite-supported nanoscale zero-valent iron.

in Table 3, phosphate solubilizing bacteria beads loaded with biochar/
nanoscale zero valent iron composite has high lead ion removal capa-
city comparing with other biological and chemical adsorbents, in-
dicating that this bio-bead could be consider as an efficient material for
the lead passivation.
3.3. Mechanism of lead passivation by bio-beads
3.3.1. SEM and ESD analysis of bio-beads before and after removal Pb2+
SEM-EDS spectra of bio-beads surface and inner before and after
reaction with Pb2+ were studied to investigate the removal mechanism
(Fig. 9(A–D)).
The beads had a lots of pleats on the surface (Fig. 9A), which en-
larges the surface area. It is important that the bio-beads was embedded
with BC/nZVI particles. And a network with large and small pores was
formed in the bio-beads (Fig. 9B), providing a convenient channel for
matter transmission. Furthermore, the reticular structure had a huge
specific surface area, which provides more reaction site to promoted
high immobilization efficiency between lead and bio-beads. SEM
images of the surface and inner structure of the bio-beads after removal
of Pb2+ were shown in Fig. 9C and D, respectively. A large number of
pleats were also found on the surface of the bio-beads after equili-
brating with culture medium containing Pb2+, and there were some
impurities on the surface, possibly due to the adsorption of some sub-
stances such as bacterial metabolites and lead-containing compounds in
the removal process. It was also found that a lot of granular materials in
the interior of the bio-beads, especially in the near of BC/nZVI, after
Pb2+ removal. In order to further analyze those substances existed on
the surface (Fig. 9E) and inside (Fig. 9F) of the bio-beads, energy
spectrum analysis was carried out. It is confirmed that lead, phos-
phorous and chlorine were present on the surface and inner of the bio-
bead, and suggest the formation of phosphate precipitation.

7
Z. Teng, et al. Journal of Hazardous Materials 384 (2020) 121505

Fig. 9. SEM images of bio-beads: (A) surface and (B) inner structure of bio-bead before using; (C) surface and (D) inner structure of bio-bead after using for Pb2+
removal; EDS spectra of the bio-bead after using for Pb2+ removal: (E) surface of bio-bead and (F) inner of bio-bead.

3.3.2. FT-IR and XRD analysis of lead bounded to bio-beads
FT-IR spectrum of bio-beads was recorded to investigate possible
interactions between Pb2+ and surface functional groups (Fig. 10A).
The complex changed of multiple functional groups were shown in
Table 3. The band at 3355.18 cm−1 in bio-beads and the composite
resulted from OeH stretching may be attributed to H2O and MeOH (M
short for Metal), while the band at 1272.89 cm−1 can be attributed to
C–OH bending (Adams et al., 2017). The strong peak at 1635.47 cm−1,
ascribed to C]O stretching vibration of the protein or amine (He et al.,
2017). The C–N bending vibration at 1336.53 cm−1 was disappear,
which is attributed to the effective interaction of lead with the aromatic
amine. Previous study has shown that aromatic amine is a cationic
surfactant with a hydrophobic and a hydrophilic group (Teng et al.,
2019b). Therefore, it is suggested that the bio-beads is an amphiphilic
material, which could also capture Pb2+ via electrostatic attraction and
ion-exchange. The peak at 1112.81 cm−1 disappeared after treatment,
indicated that the C–O of carboxyl group involved in Pb2+ binding.
Therefore, it can be concluded that the carboxyl, hydroxyl, and acyl
amino groups may be the main function groups for Pb2+ binding.
To identify Pb2+ compounds formed during the biosorption process,
the bio-beads were scanned by XRD (Fig. 10B). By observing and ana-
lyzing the characteristic peak of 2θ = 30.115°, 30.178°, corresponding
to Pb5(PO4)3OH and Pb5(PO4)3Cl, it was demonstrated that lead was
mineralized into a stable phosphorus compound. The concludes were

8
Z. Teng, et al.
Fig. 10. The Fourier transform infrared spectroscopy (A) and X-ray diffractometer patterns (B) of bio-beads before (CK) and after (1 mM Pb2+) reaction with Pb2+.
consistent with the results of EDS, which further demonstrated that
Pb2+ was precipitated by phosphate.
Fig. 11 is presented the whole experiment procedure and the me-
chanism of passivation of lead(II). Specifically, lead could be trans-
formed into stable crystal texture, such as Pb5(PO4)3Cl and
Pb5(PO4)3OH, which was the main mechanism of Pb2+ passivation. In
addition, both hydrophobic and hydrophilic groups in the bio-beads
was observed, which indicated that electrostatic attraction and ion-
exchange were also the mechanism of Pb2+ adsorption.
4. Conclusion
In this research, a unique approach was proposed for highly efficient
treatment of lead contamination by novel carrier with high surface area
and porous structure derived from the BC/nZVI embedded in the PVA-
SA. The novel carrier successfully protected the PSB from the lead
toxicity and provided a highly favorable environmental to enhance the
simultaneous adsorption-passivation. The optimal conditions of pre-
pared bio-beads were 9% PVA,1% SA, 0.8% BC/nZVI and 50% cell
suspension. Obviously, the immobilized PSB beads could achieve
Fig. 11. Schematics of the preparation of bio-beads and their application for immobilization of Pb.
9
Journal of Hazardous Materials 384 (2020) 121505
excellent remediation efficiency, and the highest removal rate of Pb2+
in beef peptone liquid medium could reach 93%, which is better than
that of free bacteria. Lead ions were converted to lead hydroxyapatite
and pyromorphite by adsorption and precipitation of bio-beads. It is
strongly believed that this work will provide a new horizon to the
search of highly active, biocompatible carriers for passivation of lead in
soil by enhancing synergic adsorption and precipitation. Further study
should be focus on the metal species competition in multi-metal solu-
tion and the performance in remediation of the multi-metal con-
taminated soil.
Acknowledgements
This work was supported by the National Natural Science
Foundation of China (No. 51279004) and the Fundamental Research
Funds for the Central Universities (No. 2015ZCQ-HJ-01). We wish to
thank Kedong Gong & Yushan Wang for their assistance in laboratory
work.
Z. Teng, et al.
Appendix A. Supplementary data
Supplementary material related to this article can be found, in the
online version, at doi:https://doi.org/10.1016/j.jhazmat.2019.121505.
References
Adams, E.M., Wellen, B.A., Thiraux, R., Reddy, S.K., Vidalis, A.S., Paesani, F., Allen, H.C.,
2017. Sodium–carboxylate contact ion pair formation induces stabilization of pal-
mitic acid monolayers at high pH. Phys. Chem. Chem. Phys. 19, 10481–10490.
Ali, M., Oshiki, M., Rathnayake, L., Ishii, S., Satoh, H., Okabe, S., 2015. Rapid and suc-
cessful start-up of anammox process by immobilizing the minimal quantity of bio-
mass in PVA-SA gel beads. Water Res. 79, 147–157.
Ayranci, E., Duman, O., 2004. Binding of lead ion to bovine serum albumin studied by ion
selective electrode. Protein Peptide Lett. 11, 331–337.
Bai, J., Yang, X.H., Du, R.Y., Chen, Y.M., Wang, S.Z., Qiu, R.L., 2014. Biosorption me-
chanisms involved in immobilization of soil Pb by Bacillus subtilis DBM in a multi-
metal-contaminated soil. J. Environ. Sci.-China 26, 2056–2064.
Bai, N., Wang, S., Sun, P., Abuduaini, R., Zhu, X., Zhao, Y., 2018. Degradation of non-
ylphenol polyethoxylates by functionalized Fe3O4 nanoparticle-immobilized
Sphingomonas sp. Y2. Sci. Total Environ. 615, 462–468.
Chen, S., Xu, M., Ma, Y., Yang, J., 2007. Evaluation of different phosphate amendments
on availability of metals in contaminated soil. Ecotoxicol Environ. Saf. 67, 278–285.
Chen, Z., Pan, X., Chen, H., Guan, X., Lin, Z., 2016a. Biomineralization of Pb(II) into Pb-
hydroxyapatite induced by Bacillus cereus 12-2 isolated from lead-zinc mine tailings.
J. Hazard. Mater. 301, 531–537.
Chen, Z., Pan, X., Chen, H., Guan, X., Lin, Z., 2016b. Biomineralization of Pb(II) into Pb-
hydroxyapatite induced by Bacillus cereus 12-2 isolated from Lead-Zinc mine tailings.
J. Hazard. Mater. 301, 531–537.
Chen, H., Zhang, J., Tang, L., Su, M., Tian, D., Zhang, L., Li, Z., Hu, S., 2019. Enhanced Pb
immobilization via the combination of biochar and phosphate solubilizing bacteria.
Environ. Int. 127, 395–401.
Diao, Z.H., Du, J.J., Jiang, D., Kong, L.J., Huo, W.Y., Liu, C.M., Wu, Q.H., Xu, X.R., 2018.
Insights into the simultaneous removal of Cr6+ and Pb2+by a novel sewage sludge-
derived biochar immobilized nanoscale zero valent iron: Coexistence effect and
mechanism. Sci. Total Environ. 642, 505–515.
Dong, H., Deng, J., Xie, Y., Zhang, C., Jiang, Z., Cheng, Y., Hou, K., Zeng, G., 2017.
Stabilization of nanoscale zero-valent iron (nZVI) with modified biochar for Cr(VI)
removal from aqueous solution. J. Hazard. Mater. 332, 79–86.
Edris, G., Alhamed, Y., Alzahrani, A., 2013. Biosorption of cadmium and lead from
aqueous solutions by chlorella vulgaris biomass: equilibrium and kinetic study. Arab.
J. Sci. Eng. 39, 87–93.
Gil-Diaz, M., Pinilla, P., Alonso, J., Lobo, M.C., 2017. Viability of a nano-remediation
process in single or multi-metal(loid) contaminated soils. J. Hazard. Mater. 321,
812–819.
Gong, K., Hu, Q., Yao, L., Li, M., Sun, D., Shao, Q., Qiu, B., Guo, Z., 2018. Ultrasonic
pretreated sludge derived stable magnetic active carbon for Cr(VI) removal from
wastewater. ACS Sustain. Chem. Eng. 6, 7283–7291.
He, S., Zhong, L., Duan, J., Feng, Y., Yang, B., Yang, L., 2017. Bioremediation of waste-
water by iron oxide-biochar nanocomposites loaded with photosynthetic bacteria.
Front. Microbiol. 8, 823.
Hu, B., Ye, F., Jin, C., Ma, X., Huang, C., Sheng, G., Ma, J., Wang, X., Huang, Y., 2017. The
enhancement roles of layered double hydroxide on the reductive immobilization of
selenate by nanoscale zero valent iron: macroscopic and microscopic approaches.
Chemosphere 184, 408–416.
Huang, D., Xue, W., Zeng, G., Wan, J., Chen, G., Huang, C., Zhang, C., Cheng, M., Xu, P.,
2016. Immobilization of Cd in river sediments by sodium alginate modified nanoscale
zero-valent iron: impact on enzyme activities and microbial community diversity.
Water Res. 106, 15–25.
Hussain, I., Li, M.Y., Zhang, Y.Q., Li, Y.C., Huang, S.B., Du, X.D., Liu, G.Q., Hayat, W.,
Anwar, N., 2017. Insights into the mechanism of persulfate activation with nZVI/BC
nanocomposite for the degradation of nonylphenol. Chem. Eng. J. 311, 163–172.
Karim, M.R., Aijaz, M.O., Alharth, N.H., Alharbi, H.F., Al-Mubaddel, F.S., Awual, M.R.,
2019. Composite nanofibers membranes of poly (vinyl alcohol)/chitosan for selective
lead(II) and cadmium(II) ions removal from wastewater. Ecotoxicol. Environ. Saf.
169, 479–486.
Kim, S.A., Kannan, S.K., Lee, K.J., Park, Y.J., She, P.J., Lee, W.H., Kim, H.M., Oh, B.T.,
2013. Removal of Pb(II) from aqueous solution by a zeolite–nanoscale zero-valent
iron composite. Chem. Eng. J. 217, 54–60.
Kim, I.H., Lee, W.J., Kim, Y.K., Oh, B.K., 2016. Development of recycled aggregate bio-
carrier with sulfate reducing bacteria for the elimination of heavy metals from sea-
water. Biotechnol. Bioprocess Eng. 21, 689–693.
Kookana, R.S., 2010. The role of biochar in modifying the environmental fate, bioavail-
ability, and efficacy of pesticides in soils: a review. Aust. J. Soil Res. 48, 627–637.
Lefevre, E., Bossa, N., Gardner, C.M., Gehrke, G.E., Cooper, E.M., Stapleton, H.M., Hsu-
Kim, H., Gunsch, C.K., 2018. Biochar and activated carbon act as promising
amendments for promoting the microbial debromination of tetrabromobisphenol A.
Water Res. 128, 102–110.
Li, Z., Bai, T., Dai, L., Wang, F., Tao, J., Meng, S., Hu, Y., Wang, S., Hu, S., 2016a. A study
of organic acid production in contrasts between two phosphate solubilizing fungi:
penicillium oxalicum and Aspergillus niger. Sci. Rep. 6, 25313.
Li, Z., Wang, F., Bai, T., Tao, J., Guo, J., Yang, M., Wang, S., Hu, S., 2016b. Lead im-
mobilization by geological fluorapatite and fungus Aspergillus niger. J. Hazard. Mater.
320, 386–392.
10
Journal of Hazardous Materials 384 (2020) 121505
Li, Z., Tang, L., Zheng, Y., Tian, D., Su, M., Zhang, F., Ma, S., Hu, S., 2017a. Characterizing
the mechanisms of lead immobilization via bioapatite and various clay minerals. ACS
Earth Space Chem. 1, 152–157.
Li, X., Dai, L., Zhang, C., Zeng, G., Liu, Y., Zhou, C., Xu, W., Wu, Y., Tang, X., Liu, W., Lan,
S., 2017b. Enhanced biological stabilization of heavy metals in sediment using im-
mobilized sulfate reducing bacteria beads with inner cohesive nutrient. J. Hazard.
Mater. 324, 340–347.
Li, Z., Su, M., Duan, X.F., Tian, D., Yang, M.Y., Guo, J.Y., Wang, S.M., Hu, S.J., 2018a.
Induced biotransformation of lead (II) by Enterobacter sp. in SO4-PO4-Cl solution. J.
Hazard. Mater. 357, 491–497.
Li, G., Khan, S., Ibrahim, M., Sun, T.R., Tang, J.F., Cotner, J.B., Xu, Y.Y., 2018b. Biochars
induced modification of dissolved organic matter (DOM) in soil and its impact on
mobility and bioaccumulation of arsenic and cadmium. J. Hazard. Mater. 348,
100–108.
Li, Z.T., Wang, L., Meng, J., Liu, X.M., Xu, J.M., Wang, F., Brookes, P., 2018c. Zeolite-
supported nanoscale zero-valent iron: new findings on simultaneous adsorption of Cd
(II), Pb(II), and As(III) in aqueous solution and soil. J. Hazard. Mater. 344, 1–11.
Lv, X.S., Jiang, G.M., Xue, X.Q., Wu, D.L., Sheng, T.T., Sun, C., Xu, X.H., 2013. Fe0-Fe3O4
nanocomposites embedded polyvinyl alcohol/sodium alginate beads for chromium
(VI) removal. J. Hazard. Mater. 262, 748–758.
Manasi, V., Rajesh, Rajesh, N., 2014. Adsorption isotherms, kinetics and thermodynamic
studies towards understanding the interaction between a microbe immobilized
polysaccharide matrix and lead. Chem. Eng. J. 248, 342–351.
Mohan, D., Sarswat, A., Ok, Y.S., Pittman Jr, C.U., 2014. Organic and inorganic con-
taminants removal from water with biochar, a renewable, low cost and sustainable
adsorbent—a critical review. Bioresour. Technol. 160, 191–202.
Mukherjee, R., Kumar, R., Sinha, A., Lama, Y., Saha, A.K., 2015. A review on synthesis,
characterization, and applications of nano zero valent iron (nZVI) for environmental
remediation. Crit. Rev. Environ. Sci. Technol. 46, 443–466.
Ngatia, L.W., Hsieh, Y.P., Nemours, D., Fu, R., Taylor, R.W., 2017. Potential phosphorus
eutrophication mitigation strategy: biochar carbon composition, thermal stability and
pH influence phosphorus sorption. Chemosphere 180, 201–211.
Niu, L., Yang, F., Xu, C., Yang, H., Liu, W., 2013. Status of metal accumulation in farm-
land soils across China: from distribution to risk assessment. Environ. Pollut. 176,
55–62.
Oh, S.Y., Seo, Y.D., Kim, B., Kim, I.Y., Cha, D.K., 2016. Microbial reduction of nitrate in
the presence of zero-valent iron and biochar. Bioresour. Technol. 200, 891–896.
Park, J.H., Bolan, N., 2013. Lead immobilization and bioavailability in microbial and root
interface. J. Hazard. Mater. 261, 777–783.
Park, J.H., Bolan, N.S., Chung, J.W., Naidu, R., Megharaj, M., 2011. Environmental
monitoring of the role of phosphate compounds in enhancing immobilization and
reducing bioavailability of lead in contaminated soils. J. Environ. Monit. 13,
2234–2242.
Qian, L., Zhang, W., Yan, J., Han, L., Chen, Y., Ouyang, D., Chen, M., 2017. Nanoscale
zero-valent iron supported by biochars produced at different temperatures: Synthesis
mechanism and effect on Cr(VI) removal. Environ. Pollut. 223, 153–160.
Rajapaksha, A.U., Chen, S.S., Tsang, D.C.W., Zhang, M., Vithanage, M., Mandal, S., Gao,
B., Bolan, N.S., Ok, Y.S., 2016. Engineered/designer biochar for contaminant re-
moval/immobilization from soil and water: potential and implication of biochar
modification. Chemosphere 148, 276–291.
Shen, Z., Zhang, Y., Jin, F., McMillan, O., Al-Tabbaa, A., 2017. Qualitative and quanti-
tative characterisation of adsorption mechanisms of lead on four biochars. Sci. Total
Environ. 609, 1401–1410.
Shen, Z., Tian, D., Zhang, X., Tang, L., Su, M., Zhang, L., Li, Z., Hu, S., Hou, D., 2018.
Mechanisms of biochar assisted immobilization of Pb2+ by bioapatite in aqueous
solution. Chemosphere 190, 260–266.
Su, H., Fang, Z., Tsang, P.E., Zheng, L., Cheng, W., Fang, J., Zhao, D., 2016. Remediation
of hexavalent chromium contaminated soil by biochar-supported zero-valent iron
nanoparticles. J. Hazard. Mater. 318, 533–540.
Sun, Y.Q., Lei, C., Khan, E., Chen, S.S., Tsang, D.C.W., Ok, Y.S., Lin, D.H., Feng, Y.J., Li,
X.D., 2017. Nanoscale zero-valent iron for metal/metalloid removal from model
hydraulic fracturing wastewater. Chemosphere 176, 315–323.
Sun, Y.Q., Lei, C., Khan, E., Chen, S.S., Tsang, D.C.W., Ok, Y.S., Lin, D.H., Feng, Y.J., Li,
X.D., 2018. Aging effects on chemical transformation and metal(loid) removal by
entrapped nanoscale zero-valent iron for hydraulic fracturing wastewater treatment.
Sci. Total Environ. 615, 498–507.
Sun, Y.Q., Yu, I.K.M., Tsang, D.C.W., Cao, X.D., Lin, D.H., Wang, L.L., Graham, N.J.D.,
Alessi, D.S., Komárek, M., Ok, Y.S., Feng, Y.J., Li, X.D., 2019. Multifunctional iron-
biochar composites for the removal of potentially toxic elements, inherent cations,
and hetero-chloride from hydraulic fracturing wastewater. Environ. Int. 124,
521–532.
Teng, Z., Chen, Z., Zhang, Q., Yao, Y., Song, M., Li, M., 2018. Isolation and character-
ization of phosphate solubilizing bacteria from rhizosphere soils of the Yeyahu
Wetland in Beijing, China. Environ. Sci. Pollut. Res. Int. https://doi.org/10.1007/
s11356-018-2955-5.
Teng, Z.D., Shao, W., Zhang, K.Y., Huo, Y.Q., M, Li, 2019a. Characterization of phosphate
solubilizing bacteria isolated from heavy metal contaminated soils and their potential
for lead immobilization. J. Environ. Manage. 231, 189–197.
Teng, Z.D., Shao, W., Zhang, K.Y., Huo, Y.Q., Zhu, J., Li, M., 2019b. Pb biosorption
byLeclercia adecarboxylata: protective and immobilized mechanisms of extracellular
polymeric substances. Chem. Eng. J. 375, 122113.
Wang, Y.Y., Yao, W.B., Wang, Q.W., Yang, Z.H., Liang, L.F., Chai, L.Y., 2016. Synthesis of
phosphate-embedded calcium alginate beads for Pb(II) and Cd(II) sorption and im-
mobilization in aqueous solutions. Trans. Nonferrous Met. Soc. China 26, 2230–2237.
Wang, L., Yu, K.Q., Li, J.S., Tsang, D.C.W., Poon, C.S., Yoo, J.C., 2018. Low-carbon and
low-alkalinity stabilization/solidification of high-Pb contaminated soil. Chem. Eng. J.
Z. Teng, et al.
351, 418–427.
Wang, L., Cho, D.W., Tsang, D.C.W., Cao, X.D., Hou, D.Y., Shen, Z.T., Alessi, D.S., Ok,
Y.S., Poon, C.S., 2019a. Green remediation of As and Pb contaminated soil using
cement-free claybased stabilization/solidification. Environ. Int. 126, 336–345.
Wang, N.N., Qiu, Y.Y., Xiao, T.F., Wang, J.Q., Chen, Y.X., Xu, X.J., Kang, Z.C., Fan, L.L.,
Yu, H.W., 2019b. Comparative studies on Pb(II) biosorption with three spongy mi-
crobe-based biosorbents: high performance, selectivity and application. J. Hazard.
Mater. 373, 39–49.
Wei, A.L., Ma, J., Chen, J.J., Zhang, Y., Song, J.X., Yu, X.Y., 2018. Enhanced nitrate
removal and high selectivity towards dinitrogen for groundwater remediation using
biochar-supported nano zero-valent iron. Chem. Eng. J. 353, 595–605.
Xu, Y., Yan, X., Fan, L., Fang, Z., 2016a. Remediation of Cd(ii)-contaminated soil by three
kinds of ferrous phosphate nanoparticles. RSC Adv. 6, 17390–17395.
Xu, Y., Fang, Z., Tsang, E.P., 2016b. In situ immobilization of cadmium in soil by stabi-
lized biochar-supported iron phosphate nanoparticles. Environ. Sci. Pollut. Res. Int.
23, 19164–19172.
Xue, W., Huang, D., Zeng, G., Wan, J., Zhang, C., Xu, R., Cheng, M., Deng, R., 2018.
Nanoscale zero-valent iron coated with rhamnolipid as an effective stabilizer for
immobilization of Cd and Pb in river sediments. J. Hazard. Mater. 341, 381–389.
Yan, J.C., Han, L., Gao, W.G., Xue, S., Chen, M.F., 2015. Biochar-supported nanoscale
zerovalent compasite used as persulfulfate activator for removing trichloroethylene.
Bioresour. Technol. 175, 269–274.
Yang, F., Zhang, S.S., Sun, Y.Q., Cheng, K., Li, J.S., Tsang, D.C.W., 2018. Fabrication and
characterization of hydrophilic corn stalk biochar supported nanoscale zero-valent
iron composites for efficient metal removal. Bioresour. Technol. 265, 490–497.
Yu, J.P., He, H.J., Yang, W.L., Yang, C.P., Zeng, G.M., Wu, X., 2018. Magnetic bionano-
particles of Penicillium sp. yz11-22N2 doped with Fe3O4 and encapsulated within
PVA-SA gel beads for atrazine removal. Bioresour. Technol. 260, 196–203.
Yuan, J.Z.B., Awad, Y.M., Beckers, F., Tsang, D.C.W., Ok, Y.S., Rinklebe, J., 2017a.
Mobility and phytoavailability of As and Pb in a contaminated soil using pine sawdust
11
Journal of Hazardous Materials 384 (2020) 121505
biochar under systematic change of redox conditions. Chemosphere 178, 110–118.
Yuan, Z., Yi, H., Wang, T., Zhang, Y., Zhu, X., Yao, J., 2017b. Application of phosphate
solubilizing bacteria in immobilization of Pb and Cd in soil. Environ. Sci. Pollut. Res.
Int. 24, 21877–21884.
Zeng, G.M., Wang, J., Huang, D.L., Hu, L., Huang, C., Cheng, M., Xue, W.J., Gong, X.M.,
Wang, R.Z., Jiang, D.N., 2017. Precipitation, adsorption and rhizosphere effect: the
mechanisms for phosphate-induced Pb immobilization in soils—a review. J. Hazard.
Mater. 339, 354–367.
Zhan, L., Li, X., Wang, M., He, Y., Chai, L., Huang, J., Wang, H., Wu, X., Lai, Y., 2016.
Highly flexible and porous nanoparticle-loaded films for dye removal by graphene
oxide-fungus interaction. ACS Appl. Mater. Interfaces 8, 34638–34647.
Zhang, M., Wang, H., Han, X., 2016. Preparation of metal-resistant immobilized sulfate
reducing bacteria beads for acid mine drainage treatment. Chemosphere 154,
215–223.
Zhang, X., Li, Y., Li, H., 2017a. Enhanced bio-immobilization of Pb contaminated soil by
immobilized bacteria with biochar as carrier. Pol. J. Environ. Stud. 26, 413–418.
Zhang, X.Q., Li, Y.S., Li, H., 2017b. Enhanced bio-immobilization of Pb contaminated soil
by immobilized bacteria with biochar as carrier. Pol. J. Environ. Stud. 26, 413–418.
Zhang, K.J., Xue, Y.W., Xu, H.H., Yao, Y.N., 2019. Lead removal by phosphate solubilizing
bacteria isolated from soil through biomineralization. Chemosphere 224, 272–279.
Zhu, S., Ho, S.H., Huang, X., Wang, D., Yang, F., Wang, L., Wang, C., Cao, X., Ma, F., 2017.
Magnetic nanoscale zerovalent iron assisted biochar: interfacial chemical behaviors
and heavy metals remediation performance. ACS Sustain. Chem. Eng. 5, 9673–9682.
Zhu, J., Li, M., Whelan, M., 2018. Phosphorus activators contribute to legacy phosphorus
availability in agricultural soils: a review. Sci. Total Environ. 612, 522–537.
Zou, Y., Wang, X., Khan, A., Wang, P., Liu, Y., Alsaedi, A., Hayat, T., Wang, X., 2016.
Environmental remediation and application of nanoscale zero-valent iron and its
composites for the removal of heavy metal ions: a review. Environ. Sci. Technol. 50,
7290–7304.