Synthesis of a superparamagnetic ultrathin FeCO3 nanorods-enzyme bionanohybrid as a novel heterogeneous catalyst

Article abstract of DOI:10.1039/c8cc02851f

Herein we report a straightforward synthesis of an ultrathin protein-iron(ii) carbonate nanorods (FeCO₃-NRs) heterogeneous bionanohybrid at room temperature and in aqueous media. The enzyme induced the in situ formation of well-dispersed FeCO₃ NRs on a protein network. The addition of NaBH₄ as a reducing agent allowed us to obtain nanorods (5 × 40 nm) with superparamagnetic properties. This bionanohybrid showed excellent catalytic results in reduction, oxidation and C-C bond reactions.

Full text of DOI:10.1039/c8cc02851f

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Synthesis of a superparamagnetic ultrathin FeCO3 nanorods-
enzyme bionanohybrid as novel heterogeneous catalyst†
Rocio Benavente, a David Lopez-Tejedor a and Jose M. Palomo* a
Received 00th January 20xx,
Accepted 00th January 20xx
DOI: 10.1039/x0xx00000x
www.rsc.org/
Herein we report a straightforward synthesis of ultrathin protein- which are attracting much attention due to their enhanced
iron (II) carbonate nanorods (FeCO3-NRs) heterogeneous optical, magnetic, catalytic and mechanical properties over
bionanohybrid at room temperature and aqueous media. The spherically shaped nanoparticles.16-22 However, only a few
enzyme induced the in situ formation of well-dispersed FeCO3 NRs examples of these kind of nanostructures of iron (mainly iron
on a protein network. The addition of NaBH4 as reducing agent oxides) has been reported.23-30 In addition, there are rare cases
allowed getting nanowires (5x40 nm) with superparamagnetic of the application of iron nanorods or nanowires in catalytic
properties. This bionanohybrid showed excellent catalytic results process that have been described. 27-29
in reduction, oxidation and C-C bond reactions.
We have developed for the first time the successful design and
green synthesis of ultrathin iron (II) carbonate nanorods based
on an enzyme (Candida antarctica B lipase) and iron salts (Fig.
1). Well-dispersed nanowires on the biological matrix -avoiding
any aggregation problem- were obtained using this
methodology at room temperature in aqueous media. The
structure of the new heterogeneous nanocatalyst was
confirmed by transmission electron microscopy (TEM)
combined with other characterization techniques.
Iron catalysis has gained an extraordinary attention in the last
years in organic chemistry 1-4 and the design and development
of new types of iron catalysts is highly desirable.
In particular, iron nanostructures with controlled size and
composition have been recently developed with different
interesting properties.5-10 They are of great interest for
heterogeneous catalysis because of their unique
nanostructure-dependent properties which drastically
differentiate their catalytic performance from that of bulk
metals. 11
Different synthetic methods have been described for iron
nanoparticles, where controlling the morphology and size of
the nanoparticles exerts tremendous impact on their catalytic
properties. 12-15 Many of these methods involve the application
of hard conditions (e.g. high temperatures or the presence of
organic solvents) and the necessity of highly controllable
conditions or the utilisation of special equipment. 14-15
Therefore, the application of new technologies where the iron
nanostructures can be synthesized in mild conditions and
simple processes where no additional equipment is required
for the synthesis of the nanoparticles would represent a
tremendous impact. Furthermore, nanorods (NRs) and
nanowires (NWs) represent a new class of 1D-nanomaterials
Fig. 1. Graphic illustration of the synthesis of CAL-B-FeCO3
nanorods bionanohybrid.
Iron (II) carbonate nanorods (FeCO3NRs) with a diameter of 5-7
nm and length from 40-93 nm were observed by TEM. The iron
nanobiohybrid was reduced for 15 min and generated a highly
stable and superparamagnetic catalyst, with excellent catalytic
efficiency in hydrogenation process in aqueous media, the
degradation of aromatic compounds and C-C bond formation.
A similar synthetic approach has been recently applied for
designing bionanoconstructs from gold nanoclusters and
proteins.31-32 However, this represents the first time the
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synthesis of magnetic nanorods in a biological environment The catalytic activity of this bionanohybrid in the reduction of
and with catalytic properties. p-nitrophenol was tested and no conversion was observed.
The synthesis of this bionanohybrid was performed in aqueous Therefore, a treatment with sodium borohydride was applied
media by adding lipase from Candida antartica B (CAL-B, 33 after the previous incubation of the Fe2+ salt with CAL-B in
kDa, monomeric enzyme) to an aqueous solution of fully water bicarbonate buffer. The reduction step was evaluated at
soluble iron salts at room temperature and under gentle different times after adding an aqueous solution of 1.2 M of
stirring. Initially (NH4)2Fe (SO4)2 (Mohr´s salt) was used at sodium borohydride (to achieve a final concentration of 0.12
different concentrations. 3.6 mL of CAL-B solution (approx. 18 M). After that, the solid was washed three times with water to
mg protein calculated by Bradford assay33) was added to 60 mL remove the remaining borohydride, frozen and lyophilized for
of iron salt solution at room temperature. In order to control 18
h to obtain the so called reduced CAL-B-FeCO3NRs
the pH of the final solution -which must be higher than the bionanohybrids. When the reduction was performed at 15
isoelectric point of the lipase pI=6 to obtain a negatively min, small nanorods (approx. 5 nm diameter x 40 nm length) -
charged protein and ≤10 to avoid iron oxide nanoparticle as far as we know the smallest reported iron nanorods- were
formation in solution-, different buffers and different determined by TEM (Figure 3). XRD and XPS demonstrated
concentrations were tested, with 100 mM of sodium that FeCO3 was the unique iron species (Figure 3, Figure S2).
bicarbonate (pH 10) the best option. At concentration of 10 This reduced bionanohybrid (CAL-B-FeCO3NRs-2) presented
mg/mL of iron salts, the solution started to turn cloudy (first excellent magnetism in the presence of a magnetic field
step in the bionanohybrid formation) after 30 min with a permitting a fast recovery. The analysis of magnetic properties
decrease in the pH of the solution to around 8, which was revealed a superparamagnetic material with a saturation
conserved unaltered during all the incubation time. After 16- magnetization (Ms) value of 125 emu/g considering the total
hours of incubation, a solid was obtained, washed several mass of the sample or 267 emu/g considering the iron content
times with distilled water, centrifuged and lyophilized of the sample (Figure S3), extremely higher compared with
overnight. ICP-OES analysis revealed that this new bulk siderite (Ms <0.5 emu/g).
bionanohybrid contained 47% (wt.) of iron. The solid was not
formed in any case for the enzyme without iron or for the iron
salt without protein. XRD and XPS analysis of the solid
demonstrated the presence of iron (II) carbonate (siderite,
FeCO3)34 as the main iron species, although some minor
contamination of iron oxide (magnetite or maghemite) was
found (Figure 2A-B, Figure S1). The agitation speed of the
solution in the preparation was an important parameter to
avoid the formation of oxidative species (e.g. magnetite). TEM
analysis confirmed the formation of nanorods (FeCO3NRs) with
a size of approx. 7 nm in diameter by 59 nm in length induced
by the protein matrix, obtaining the so-called CAL-B-FeCO3NRs-
1 bionanohybrid (Figure 2C-D). This hybrid did not possess any
evidence of magnetism in the presence of a magnetic field.
Fig. 3. Characterization of CAL-B-FeCO3NRs-2 bionanohybrid.
A) XRD. B) Full XPS spectrum. C) XPS spectrum of Fe2p. D)
TEM/HRTEM.
The reduction was also performed for 30, 45, 60 and 360
minutes recovering the iron bionanohybrids following the
same protocol as previously described. The different
bionanohybrids were called CAL-B-FeCO3NRs-3, CAL-B-
Fig.
2
.
FeCO3NRs-4 CAL-B-FeCO3NRs-5 and CAL-B-FeCO3NRs-6,
Characterization of CAL-B-FeCO3NRs-1 bionanohybrid.A) XRD. respectively. XRD analysis of the different bionanohybrids
● FeCO3, iron oxide impurity. B) XPS. C) TEM. D) HRTEM.
*
showed the presence of the same contaminants of Fe(III)
observed in CAL-B-FeCO3NRs-1 which had increasing residual
2 | J. Name., 2012, 00, 1-3
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the presence of the iron oxides contaminant was observed. Around
33% slower rate was obtained for the bionanohybrids reduced for
30 or 45 min in comparison with the results of CAL-B-FeCO3NRs-2.
The CAL-B-FeCO3NRs-5 bionanohybrid (1h reducing incubation)
achieved full reduction after 90 seconds, whereas 95% conversion
after 240 seconds was obtained using the CAL-B-FeCO3NRs-6 (6h
reducing incubation).
amounts in CAL-B-FeCO3NRs-4 to CAL-B-FeCO3NRs-6 (Fig S4-
S7). TEM analysis revealed the generation of longer-size
nanorods, from around 40 to 93 nm, apparently increasing in
length according to the reducing time but keeping the same
diameter (Table S1, Figures S4-S7).
The activity in the reduction of p-NP by the different reduced
CAL-B-FeCO3NRs bionanohybrids were also tested (Table 1).
The reduced bionanohybrid CAL-B-FeCO3NRs-2 showed high
catalytic capacity, being able to complete the total
transformation of p-NP to p-AP within 30 seconds (Figure S8).
Thus, these results demonstrated that a short-time reducing
step strongly improved the catalytic efficiency of these CAL-B-
FeCO3NRs bionanohybrids for the p-NP degradation where the
non-reduced bionanohybrid was not efficient.
Table 1. Synthesis of anilines by reduction of nitroarenes using
FeCO3NRs biohybrids as catalysts.a
The heterogeneous CAL-B-FeCO3NRs-2 catalyst showed
excellent recyclability, conserving more than 95% of its
catalytic efficiency in the p-NP reduction after six reaction
cycles.
NO2
NH2
FeCO3NRs Biohybrid
NaBH4
R
Distilled water
r.t
R
The catalytic activity of this nanobiohybrid was tested with
other nitroarenes with high yields in most of cases (Table 1).
One important point for a possible industrial application of this
bionanohybrid is the stability against oxidation. The CAL-B-
FeCO3NRs-2 bionanohybrid was stored in aerobic conditions
for 30 days. After that time, XRD analysis showed a very small
residual impurity of iron oxide in the FeCO3 spectra (Fig. S9).
The catalytic efficiency of the CAL-B-FeCO3NRs-2 in the
reduction of p-NP was still excellent, with a full conversion of
the substrate in 45 seconds. TEM analysis revealed the slight
increase of the length in the iron NRs which could explain the
slight decrease in its efficiency (Fig. S10).
Catalyst
CAL-B-
Substrate
T
[sec]
Product
Yield
[%]
0
NH2
NO2
360
FeCO3NRs-1
OH
1a
1a
OH
2a
CAL-B-
FeCO3NRs-2
CAL-B-
FeCO3NRs-3
CAL-B-
FeCO3NRs-4
CAL-B-
FeCO3NRs-5
CAL-B-
FeCO3NRs-6
CAL-B-
30
2a
2a
2a
2a
>99
>99
>99
>99
92
1a
1a
1a
45
45
In order to broaden the scope of application of these new iron
nanocatalysts, in particular importance the environmental
remediation processes, the catalytic capacity of this iron
bionanohybrid in the degradation of p-aminophenol (pAP)
(hydrolytic product of acetaminophen (paracetamol)) was
evaluated. The CAL-B-FeCO3NRs-2 biohybrid was used to
catalyse the reaction in the presence of different hydrogen
peroxide concentrations (Figure S11) in aqueous solution at
different pHs. The best results were found using 0.5 mM of
hydrogen peroxide as an oxidant at room temperature in
sodium acetate buffer pH 4 (Figure S11). This nanocatalyst
exhibited very good performance under these conditions and
more than 98% degradation of pAP was achieved in 2 min
(Figure S11). No traces of any compounds were detected by
HPLC after 50 min.
90
1a
240
120
2a
NO2
NH2
63
FeCO3NRs-2
HO
HO
1b
2b
NH2
NO2
CAL-B-
FeCO3NRs-2
60
72
HO
HO
1c
2c
NH2
>99[b]
NO2
CAL-B-
FeCO3NRs-2
900
OH
1d
OH
2d
NH2
83[b]
NO2
CAL-B-
FeCO3NRs-2
300
300
The elimination of Bisphenol-A (BPA), an important monomer
in the manufacture of plastics, food cans, and other daily-used
chemicals, is complex but crucial due to its high toxicity.
Therefore, its degradation to other non or less-toxic products
is necessary. However, only a few catalytic processes have
been developed. In this way, we tested the application of our
nanocatalyst in this process. The CAL-B-FeCO3NRs-2 biohybrid
obtained almost complete degradation of BPA after 3 days
using 150 mM of hydrogen peroxide (Figure S12).
CHO
CHO
2e
1e
NO2
NH2
CAL-B-
FeCO3NRs-2
74
O
O
O
O
1f
2f
aConditions: 1.0 mM (3 mg) nitroarene, 40 mM (3 mg) NaBH4, 2 mL of
distilled water, 3 mg of bionanohybrid, air and room temperature. b 80 mM
NaBH4
Finally,
heterogeneous nanocatalyst in C-H functionalization was
performed. The Heck reaction between ethylacrylate ( ) and
iodobenzene ( ) was performed using CAL-B-FeCO3NRs-2 as a
a very interesting application as a magnetic
3
4
A clear correlation between the decrease in the activity of the iron
nanocatalyst and the increase in the length of the nanorods with
catalyst (entry 3, Table 2). Conversion of 56% producing
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