An Unusual Two-Step Hydrolysis of Nerve Agents by a Nanozyme

Article abstract of DOI:10.1002/cctc.201801220

Organophosphate-based nerve agents irreversibly inhibit acetylcholinesterase enzyme, leading to respiratory failure, paralysis and death. Several organophosphorus hydrolases are capable of degrading nerve agents including pesticides and insecticides. Development of stable artificial enzymes capable of hydrolysing nerve agents is important for the degradation of environmentally toxic organophosphates. Herein, we describe a Zr-incorporated CeO₂ nanocatalyst that can be used for an efficient capture and hydrolysis of nerve agents such as methyl paraoxon to less toxic monoesters. This unusual sequential degradation pathway involves a covalently linked nanocatalyst-phosphodiester intermediate.

Full text of DOI:10.1002/cctc.201801220

Communications
DOI: 10.1002/cctc.201801220
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An Unusual Two-Step Hydrolysis of Nerve Agents by a
Nanozyme
Kritika Khulbe,[a] Punarbasu Roy,[a] Anusree Radhakrishnan,[a] and Govindasamy Mugesh*[a]
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Organophosphate-based nerve agents irreversibly inhibit acetyl-
cholinesterase enzyme, leading to respiratory failure, paralysis
and death. Several organophosphorus hydrolases are capable of
degrading nerve agents including pesticides and insecticides.
Development of stable artificial enzymes capable of hydrolysing
nerve agents is important for the degradation of environ-
mentally toxic organophosphates. Herein, we describe a Zr-
incorporated CeO2 nanocatalyst that can be used for an efficient
capture and hydrolysis of nerve agents such as methyl paraoxon
to less toxic monoesters. This unusual sequential degradation
pathway involves a covalently linked nanocatalyst-phospho-
diester intermediate.
24 Organophosphate-based nerve agents such as tabun (1), sarin
25 (2), VX (3) (Figure 1a) were used as chemical warfare agents
26 since World War II.[1] These compounds and other highly toxic
27 organophosphate-based pesticides/insecticides such as methyl
28 paraoxon (4) and its analogues (5–7), dichlorvos (8), methyl
29 chlorpyrifos (9) affect the nervous system, leading to paralysis,
30 convulsions, and death by asphyxiation.[1–3] This is due to the
31 irreversible inhibition of the enzyme acetylcholinesterase
32 (AChE), which is responsible for the breakdown of the neuro-
33 transmitter acetylcholine (Figure S1).[1c] It is known that soil
34 bacteria survive in extremely toxic conditions because they
35 produce several enzymes that can detoxify nerve agents by
36 hydrolysis.[4] The importance of nerve agent detoxification
37 related to environment and human health led to the develop-
38 ment of catalysts that can efficiently hydrolyse the phospho-
39 triester bonds, which include metal complexes, organic poly-
40 mers, metal-organic frameworks (MOFs) and nanomaterials.
41 Initial studies on the hydrolysis of organophosphates were
42 mainly focused on metal complexes having mono and
43 binuclear Zn(II) and Cu(II) centres.[5] Farha, Hupp, and co-
44 workers as well as Barea, Navarro, and co-workers reported the
45 design and synthesis of a series of MOFs capable of hydrolysing
46 nerve agents such as methyl paraoxon, soman and VX, under
47 catalytic conditions.[6,7] Also, clay-based catalytic systems have
48 emerged as promising candidates for the oxidative abatement
49 of nerve agent simulants.[8] Recently, nanozyme-mediated
50 catalysis attracted significant research interest.[9] We reported
51
Figure 1. a) Chemical structures of some common nerve agents. b, c) TEM
image and SAED pattern (inset) of VEC, 1ZC nanoparticles respectively. d)
HRTEM image of 1ZC showing (111) planes. e) FT-Raman spectra of 1ZC
showing characteristic peak for CeÀO and oxygen vacancies (inset). f) XRD
pattern of VEC and 1ZC. *Represents the characteristic shift in 2q (from 59.88
to 58.78) of (222) plane upon Zr-doping. g) Quantification of vacancies on
the surface of different nanoceria by XPS. h) Representation of surface
vacancy (purple) on Zr-incorporated nanoceria.
that a vacancy-engineered nanoceria (VEC) functionally mimics
the phosphotriesterase (PTE) activity and can convert paraoxon
to diethylphosphate.[9g] While a rapid hydrolysis of phospho-
triesters to diesters is important, it is equally important to
convert the diesters further to monoester and phosphate as
phosphodiesters are very persistent in the environment, and
they retain significant cholinesterase inhibitory effect.[10] How-
ever, the diesters produced by a single-step hydrolysis of the
triesters often have unactivated ester bonds that resist further
hydrolysis. Furthermore, the diesters generally act as catalyst
poisons, leading to an incomplete decontamination of the
nerve agents.[5c,6a] In this paper, we show that a simple
modification in the vacancy-engineered nanoceria (VEC) by Zr
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[a] K. Khulbe, P. Roy, A. Radhakrishnan, Prof. Dr. G. Mugesh
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Department of Inorganic and Physical Chemistry
Indian Institute of Science
Bangalore 560012 (India)
E-mail: mugesh@iisc.ac.in
Supporting information for this article is available on the WWW under
https://doi.org/10.1002/cctc.201801220
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incorporation not only alters the substrate preference, but also
facilitates the hydrolysis of diesters to monoesters through an
unusual two-step process. This study represents the first
example of a nanozyme-mediated two-step hydrolysis of a
nerve agent, leading to the formation of a monoester.
In the CeO2-mediated decontamination of nerve agents, it is
important to generate Ce3+ on the surface in addition to the
existing Ce4+ ions.[9g] This can be achieved by creating oxygen
vacancies (OVs) as surface defects in the material. It has been
10 shown theoretically that the introduction of Zr4+ ions to CeO2
11 not only can increase the oxygen release, but also can enhance
12 the thermal stability of its surface vacancies.[11] Furthermore, the
13 OVs created near the Zr4+ sites are expected to be less mobile
14 than that of the surface having only cerium ions.[11c] The Zr-
15 incorporated CeO2 nanomaterials (ZCs) required for this study
16 were prepared by co-precipitation method by changing the Zr/
17 Ce molar ratio at 1%, 5% and 10% to give 1ZC, 5ZC and 10ZC,
18 respectively. The X-ray diffraction (XRD) pattern (Figure 1f and
19 Figure S2a) showed all three Zr-doped CeO2 nanoparticles have
20 a cubic fluorite structure similar to the previously reported
21 CeO2 (JCPDS no. 01-0800) and Zr-doped CeO2 (JCPDS no. 02-
22 1311). Transmission electron microscopy (TEM) images show
23 spherical morphologies of all the three ZCs with particle
24 diameter in the range of 4.0 to 5.0 nm, which is in good
25 agreement with the crystallite size obtained from XRD (Figur-
26 es 1b, c, S3–S4 and S2b–d). High-resolution TEM (HRTEM)
27 images indicate that there is no major morphological variation
28 upon Zr-doping (Figure 1d and S5). The presence and distribu-
29 tion of Ce, Zr and O were confirmed by energy dispersive
30 spectroscopy (EDS) and X-ray mapping respectively (Figures S6
31 and S7). In FT-Raman spectra, the characteristic peak for CeÀO
32 vibration appears at 460 cmÀ1 in ZCs and bulk CeO2 (Figure 1e
33 and S8).[12] With an increase in the Zr content, the peak position
34 is shifted slightly to higher wavenumber with respect to VEC,
35 indicating that the introduction of Zr4+ strengthens the CeÀO
36 bond and thereby enhances the thermal stability (Table S2). In
Figure 2. a) Reaction scheme of the hydrolysis of 4 by nanocatalyst. b)
Comparison of the conversion of 4 by 5 mol% of 1ZC, 5ZC and 10ZC and c)
Conversion of 6 by different mol % of 1ZC determined using UV-Vis
spectroscopy. d) 31P NMR spectra of products formed after 24 h of hydrolysis
of 4: (I) without catalyst, 0.035 mol% of nanocatalysts (II) VEC, (III) 0.5ZC, (IV)
1ZC, (V) 5ZC and (VI) 10ZC. Conditions: nanocatalyst, organophosphate,
0.9 M NMM in water, 458C.
hydrolysis was performed by using 5 mol% of 1ZC in the
presence of 0.9 M N-methyl morpholine (NMM) as a general
base in water at 458C. When the progress of the hydrolysis of 4
was followed by UV-Vis spectroscopy, a peak at 405 nm
indicated the formation of p-nitrophenol (Figure S11). It was
observed that the amount of Zr present in the nanozymes has
a significant effect on the activity, which follows the order:
1ZC>5ZC>10ZC (Figure 2b and c). Interestingly, 1ZC catalyses
an unusual two-step hydrolysis of 4 and 6 at faster rate (t1/2
value of 1.2 min and 3.5 min for methyl paraoxon and methyl
parathion hydrolysis, respectively) as compared to the one-step
hydrolysis by other materials reported earlier (Table 1). When
the hydrolysis of 4 was followed by 31P NMR, a rapid decrease
in the peak corresponding to 4 at À4.3 ppm was observed with
two new peaks appearing at 2.5 ppm and 4.5 ppm correspond-
ing to the diester 10 (DMP) and monoester 11 (MMP),
respectively (Figure 2d). While no hydrolysis was observed in
the presence of bulk CeO2 or ZrO2 nanomaterial even after 24 h
(Figure S12), the formation of monoester 11 produced in the
presence of 5ZC and 10ZC was relatively lower than that of
1ZC.
37 contrast to bulk CeO2,
38 characteristic of OVs in the CeO2 lattice, was observed for all
39 nanoceria.
a
broad peak around 600 cmÀ1,
40
The presence of OVs on nanoceria surface was also
41 confirmed by X-ray photoelectron spectroscopy (XPS) and the
42 increase in the OVs upon Zr doping was quantified by
43 measuring the ratio of Ce3+/Ce4+ from the deconvolution of Ce
44 3d spectra (Figure 1g, S9, Table S3 and S4).[12] The removal of
45 oxygen from CeO2 lattice during the formation of vacancy leads
46 to an electronically dense environment by the reduction of
47 Ce4+ to Ce3+ as well as a partial reduction of Zr4+ to Zr(4Àx)+ [12b]
.
48 This suggests that Zr ions are localized in the vicinity of the
49 vacancies on the surface of nanoceria as proven by XPS analysis
50 and shown in Figure 1h. The Zr/Ce ratio on the surface of
51 different ZCs as calculated from the XPS was found to correlate
52 well with the amount determined by inductively-coupled
53 plasma mass-spectrometry (ICP-MS) (Table S5).
While higher amount (%) of unreacted methyl paraoxon
was detected in the presence of 5ZC and 10ZC, no change in
the concentration of intermediate 10 was observed (Figure 3a).
The rate of the formation of MMP (11) was also found to be
lower for 5ZC and 10ZC as compared to that of 1ZC (Fig-
ure S15c). This is probably due to the substitution of Ce4+ by
Zr4+ which can decrease the number of catalytically active Ce4+
on the surface of nanoceria. It should be noted that optimum
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The nerve agent degradation by structurally stable ZCs was
55 studied using the common pesticides/insecticides, dimethyl p-
56 nitrophenyl phosphate (DMNP) also known as methyl paraoxon
57 (4) and its thio-analogue, methyl parathion (6) (Figure 2). The
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0.5% in the nanoceria (Figure S10), a complete hydrolysis of 4
was observed. However, in this case, there was a significant
decrease in the amount of monoester 11 produced in the
reaction. Interestingly, when VEC (0% Zr) was used as catalyst,
the amount of monoester 11 produced in the reaction was
found to decrease further such that it was almost identical to
that of the diester 10 even after 24 h (Figures 3a, S15b). These
observations suggest that an optimum Zr doping is important
for the complete two-step degradation of methyl paraoxon 4
to monoester 11. Thus, among different ZCs, 1ZC was found to
be the most effective catalyst for the two-step hydrolysis of
nerve agent stimulants. The steady state kinetics studies at a
fixed concentration of nanoparticles (0.035 mol%) indicated
that the hydrolysis of 4 by the nanocatalysts leading to the
formation of 11 follows a typical Michaelis-Menten kinetics
(Figure S18 and Table S6).
Table 1. Comparison of t1/2 and product formed after hydrolysis of nerve
agents by Zr-doped nanoceria with previous studies.
Catalyst
Cat.
[mol%]
Substrate Hydrolysis of Tries- Formation
ter[d]
of mono-
ester
Product[c] t1/2
[min]
UiO-66
6.0
6.0
6.0
–
4
4
4
5
DMP
DMP
DMP
DEP
45[6a]
15[6b]
1[6f]
not ob-
served
not ob-
served
not ob-
served
not ob-
served
NU-1000
UiO-66-NH2
10
11
12
13
14
15
16
17
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19
20
21
22
23
24
25
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MIL-101(Cr)-
DAAP[b]
VEC
1ZC
1ZC
300[13]
–
5
4
6
4
4
DEP
10[9g]
1.2
3.5
10.2
14.5
observed[e]
observed[e]
observed[e]
observed[e]
observed[e]
5.0[a]
5.0[a]
5.0[a]
5.0[a]
MMP
MMTP
MMP
MMP
5ZC
10ZC
To further understand the role of base in the two-step
hydrolysis of 4, experiments were performed with an increasing
concentration of NMM. It was observed that the two-step
hydrolysis is more favoured at higher concentrations of NMM
(Figure 3b, S14 and S15a). A control reaction with NMM alone
indicated that the hydrolysis of 4 in the absence of nanocatalyst
is extremely slow. Similarly, no hydrolysis was observed in the
presence of nanocatalyst when NMM was not used. These
observations suggest that NMM plays a key role in generating a
nucleophilic OHÀ species on the catalyst surface, which can
attack at the electrophilic P(V) centre of the nerve agent.
[a] Number of moles of catalyst in 1 mL=(number of catalyst particles /
NA), NA=Avogadro’s number. [b] DAAP: dialkylaminopyridine. [c] DMP:
dimethyl phosphate, MMP: monomethyl phosphate, MMTP: monomethyl
thiophosphate, DEP: diethyl phosphate. [d] Hydrolysis monitored through
p-nitrophenol formation, lmax =405 nm, 458C, 0.9 M NMM, pH 10 by UV-Vis
spectroscopy. [e] Products confirmed by 31P NMR spectroscopy.
A time-dependent investigation of the hydrolysis of 4 by
1ZC and other nanocatalysts showed some interesting features.
A careful analysis of the hydrolysis mediated by all ZCs by 31P
NMR indicates that small amount of 10 produced during the
initial period of the reaction remains constant even after 24 h,
indicating that the hydrolysis of 4 to 11 occurs in a single cycle
without removal of 10 from the active site pocket of the
nanozyme (Figures 3c–e, S17 and S16b–c). In other words, DMP
generated from methyl paraoxon remains attached to the
reaction sites throughout the catalytic cycle. These observations
suggest the possibility of a covalent nanozyme catalysis, in
which a covalent bond formation between the substrate and
nanomaterial may favour the further hydrolysis of the diester to
the corresponding monoester. In contrast, a significant increase
in the amount of 10 was observed for VEC, suggesting that a
covalent attachment of 10 at the active site pocket of VEC is
not a favoured process (Figure 3d and S16a). The increase in
the formation of monoester in the presence of ZCs can be
ascribed to the Zr-doping in CeO2 lattice, which can stabilize
the covalent nanozyme-DMP complex. This is in agreement
with the earlier report that Zr present on the catalyst surface
acts as anchoring site for organophosphates.[7b] If the DMP gets
released from the active site after the first step hydrolysis, the
subsequent hydrolysis is not a favoured process as observed for
VEC. This is in agreement with our earlier observation that
greater amount of 10 is produced in the VEC catalysed
hydrolysis of 4 (Figure 3a).
Figure 3. a) Amount (%) of DMP (10) and MMP (11) formed after hydrolysis
of methyl paraoxon (4) by different nanocatalysts. b) Effect of [NMM] on the
relative ratio of [MMP] to [DMP] in the reaction mixture formed after
hydrolysis of methyl paraoxon by 1ZC. c) Time-dependent 31P NMR
spectroscopy analysis to monitor hydrolysis of 4 by 1ZC up to 24 h. d–e)
Speciation of reaction mixture during hydrolysis of 4 by VEC and 1ZC
respectively. Conditions: Nanocatalyst (0.035 mol%), methyl paraoxon (4)
(27.0 mM), 0.9 M NMM in water, 458C, total reaction time=24 h.
55 amount of Ce4+ ions is important for the hydrolysis of organo-
56 phosphate triester to diester, i.e., the first step of the
57 catalysis.[9g] In contrast, when the Zr content was decreased to
To study the versatility of 1ZC for the hydrolysis of nerve
agents, we carried out the hydrolysis of extremely toxic
pesticides and insecticides, methyl parathion (6), dichlorvos (8),
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and methyl chlorpyrifos (9).[14] As observed for methyl paraoxon,
its thio analogue, methyl parathion underwent a two-step
hydrolysis to produce the monoester, O-methyl thiophosphate
(17) (Figure S19–S20). Similarly, 1ZC was also able to hydrolyse
other related phosphotriesters, dichlorvos (8) and methyl
chlorpyrifos (9) to the corresponding monoester derivatives
(Figure S20–21). While all the triesters underwent the two-step
hydrolysis by 1ZC to form monoesters, the rate of conversion
was found to be dependent on the nature of leaving group
an ion-pair between 1ZC and 10 is responsible for the
hydrolysis, we used 1,3-dimethylimidazolium salt of DMP as
substrate and found that the formation of an ion-pair does not
mediate the reaction (Figure 4d and S24c). Interestingly,
externally added DMP did not inhibit the hydrolysis of 4,
indicating that DMP may not interact with the nanoceria
catalyst surface, which is in contrast to a few other catalysts
that are poisoned by DMP (Figure S25).[5c,6a] These observations
indicate that an activated phosphoester bond is essential for
the first step to form a covalent intermediate. Once such an
intermediate is formed, even an unactivated diester such as
DMP can undergo hydrolysis to produce the corresponding
monoester.
10 attached to the phosphorus centre as well as the P=X (X=O or
11 S) moiety in the molecule (Figure S23).
12
To understand the unusual two-step hydrolysis of nerve
13 agents by 1ZC in detail, we synthesized the mixed triester,
14 ethylmethyl p-nitrophenyl phosphate (12) and the activated
15 diester, methyl p-nitrophenyl phosphate (14) (Figure 4a). When
To understand the mode of binding between the nanozyme
and in-situ generated DMP (10), FT-Raman spectroscopic
analysis[15] was performed using the nanomaterial isolated from
the reaction mixture at different time points. The spectra
indicated the presence of two types of PÀO bonds on CeÀO
surface: PÀOÀC at 865 cmÀ1 and PÀO*ÀCe at 1190 cmÀ1 for the
phosphodiester bound to 1ZC during the hydrolysis of 4 and 6
(Figures 5a-b).[16] It was also observed that the intensity of the
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Figure 4. a) Chemical structures of substrates used in the reaction and their
corresponding products (12-15). b) Hydrolysis of 12 (20.0 mM) by 1ZC as a
function of time showing selective formation of 13. c) Hydrolysis of 14
(20.0 mM) by 1ZC, showing an increase in the formation of 15 as a function
of time. The reactions were monitored by 31P NMR spectroscopy. d)
Attempted hydrolysis of the unactivated ester DMP (10) (25.0 mM) by: (from
top to bottom) VEC, 1ZC, 1ZC with 2.5 mM p-nitrophenol (2.5 mM DMP used
in this case), 1ZC with ionic salt of DMP as substrate. Conditions:
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Nanocatalyst (2.5 mgmLÀ1), substrate, 0.9 M NMM in water, 458C, 24 h.
41 compound 12 was treated with 1ZC, an almost quantitative
42 conversion to monoethyl phosphate (13, MEP) was observed
43 (Figure 4b). The selective formation of 13 from 12 can be
44 ascribed to the better leaving group ability of methoxy group
45 as compared to that of an ethoxy substituent. If the reaction
46 proceeds through the formation of a covalent bond between
47 the product of the first step, i.e. a diester and the nanozyme, a
48 direct hydrolysis of diester to phosphate must occur when an
49 activated diester is used for the hydrolysis. As expected, methyl
50 p-nitrophenyl phosphate (14) readily underwent hydrolysis by
51 1ZC to produce 15 (Figure 4c). In contrast, 10 did not undergo
52 any hydrolysis by VEC, 1ZC (Figure 4d), 5ZC or 10ZC even after
53 48 h in the presence of higher concentrations of NMM (Fig-
54 ure S24a–b and d–e). The role of p-nitrophenol (eliminated in
55 the first step) in the hydrolysis of 10 was ruled out as 10 did
56 not undergo hydrolysis in the presence of externally added p-
57 nitrophenol. Furthermore, to verify the possibility of whether
Figure 5. a–d) FT-Raman spectra of catalyst-bound phosphodiester inter-
mediates using nanocatalysts recovered at different time points from the
reaction mixture containing nanocatalyst (2.0 mgmLÀ1), substrate (25.0 mM),
0.9 M NMM in water, 458C. a) Hydrolysis of 4 by 1ZC. b) Hydrolysis of 6 by
1ZC. c) Hydrolysis of 4 by 10ZC. d) Hydrolysis of 4 by VEC. e) FT-IR analysis of
the reaction mixture during hydrolysis of 4 (25 mM) by 1ZC (2.0 mgmLÀ1),
0.9 M NMM in water, 458C. f) Representation of the phosphodiester
intermediate bound to nanocatalyst. g) After the hydrolysis of 4 by VEC (top)
and 1ZC (bottom), isolated nanocatalysts were treated with 10 M NaOH and
analysed by 31P NMR spectroscopy after a complete dissolution.
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PÀO*ÀCe peak at 1190 cmÀ1 decreases during the course of the
reaction, which is due to the dissociation of the diester from
the catalyst surface, leading to the formation of monoester. The
minor difference in the peak position for the phosphate-bound
catalyst (PÀO*ÀCe) upon changing the substrate from 4 to 6
suggests that both the substrates form similar covalent
intermediates with the catalyst surface after the first step of
hydrolysis (Figures 5a–b and S26a–b). Similar changes were
observed for the intermediates isolated for 4 in the presence of
mechanism (shown for methyl paraoxon (4) and methyl para-
thion (6), a Ce3+-bound oxygen nucleophile, generated by a
proton abstraction by NMM, attacks at the phosphorus centre
of a Ce4+-bound phosphotriester to form a covalent bond, with
the elimination of p-nitrophenol. The coordination of P=X (X=O
or S) to Ce4+ ions polarises the P=X bond, making the
phosphorus centre electron deficient. As the phosphodiester
bound to the nanozyme is insoluble, we were unable to detect
this intermediate by 31P NMR spectroscopy under given reaction
conditions. However, we were able to confirm the formation of
a covalent intermediate by FT-Raman and FT-IR spectroscopy. In
the second step, the nucleophilic attack of another hydroxyl
group at the metal-bound diester leads to the cleavage of PÀO
bond to produce the corresponding monoester and methanol.
Once the monoester is formed, it is cleaved from the active site
by a base-mediated hydrolysis, leading to the regeneration of
reaction sites. As shown in Figure S30, no changes in the crystal
structure, arrangement of planes or morphology were ob-
served, indicating that the catalytically active surface of the
nanozyme is very stable under the reaction conditions. The ICP-
MS analysis also confirmed that there was no leaching of Ce
and Zr ions during the hydrolysis (Table S7).
10 VEC and 10ZC (Figures 5c–d and S26c–d). Furthermore, the FT-
11 IR analysis of the reaction mixture at different time points
12 showed a decrease in the intensity of the peak for the
13 nanozyme-bound phosphodiester intermediate (PÀO*ÀCe) at
14 1110 cmÀ1 as the reaction proceeded towards completion due
15 to dissociation of the bound intermediate, which is in agree-
16 ment with the FT-Raman spectra discussed earlier (Figure 5e,
17 S27–S28).[16] Interestingly, while the treatment of VEC-based
18 intermediate with 10 M NaOH released phosphodiester also
19 from the nanozyme, treatment of 1ZC-based intermediate
20 released exclusively the monoester (Figure 5g and S29), indicat-
21 ing that the covalent bond between the diester and 1ZC is
22 much stronger than that of VEC. This is consistent with the
23 earlier observation that no major change in the percentage
24 composition of products 10 and 11 was observed between 12
25 and 24 h during the hydrolysis of 4 by VEC. This observation
26 indicates that the diester 10 produced in the reaction is
27 resistant to further hydrolysis by VEC (Figure 3d and S16a). As
28 1ZC is expected to be structurally more stable than VEC, a
29 ZrÀOÀCe mixed surface may facilitate the formation of a
30 covalent intermediate with the diester during the catalysis
31 (Figure 5f). Together, the above observations suggest that just
32 1% Zr-doping is sufficient not only to alter the rate of the
33 reaction, but also to remarkably change the mechanistic
34 pathway.
To understand the ability of the ZC to act as a catalyst for
the purification of contaminated water, a small column was
designed using 1ZC nanomaterial. The treatment of 1% methyl
paraoxon-containing water (56.0 mM, i.e.105 times greater
concentration than usually found in waste water) was carried
out. When methyl paraoxon solution was passed through the
column packed with 1ZC, the 31P NMR spectroscopy analysis of
the eluted pesticide showed that more than 90% methyl
paraoxon has been converted to monomethyl phosphate. Thus,
such covalent catalysis mediated hydrolysis by Zr-doped nano-
ceria (1ZC) has the potential for the removal of pesticide/
insecticide residues from drinking water. (Figure S31).
35
On the basis of above observations, a catalytic cycle
In conclusion, we showed for the first time that a simple Zr-
incorporation in CeO2-based nanozymes can facilitate the two-
step hydrolysis of nerve agents, leading to the formation of less
toxic monoesters. The mechanism of the degradation involves
an unusual formation of a covalent intermediate with the
nanomaterial, which makes it possible for the cleavage of an
unactivated diester bond. The efficiency of two-step covalent
catalysis depends not only on the nature of leaving group, but
also on the groups attached to the phosphorus centre as well
as on the nature of the P=X (X=O or S) moiety in the molecule.
While the vacancies in CeO2 can act as reactive sites for nerve
agent degradation, Zr doping can fine-tune the activity as well
as the selectivity of the nanozymes.
36 involving a covalent intermediate is proposed for the hydrolysis
37 of organophosphate nerve agents (Figure 6). According to this
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Caution: The organophosphates described in this paper are
highly hazardous in nature and necessary precautions should
be taken at all stages when working with these compounds.
Acknowledgements
This study was supported by the Science and Engineering
Research Board (SERB) and DST Nano Mission (SR/NM/NS-1380/
2014). G.M. acknowledges the SERB/DST for the award of J. C.
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Figure 6. Mechanism of organophosphate ester hydrolysis by 1ZC.
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Keywords: catalysis · enzyme models · nanozymes · nerve
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Manuscript received: July 26, 2018
Accepted Article published: August 29, 2018
Version of record online: August 29, 2018
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COMMUNICATIONS
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The first two steps! Zr-incorporation
in nanoceria facilitates the two-step
hydrolysis of nerve agents, by which
unactivated diesters can be hydro-
lysed to monoesters. This study rep-
resents the first example of a
K. Khulbe, P. Roy, A. Radhakrishnan,
Prof. Dr. G. Mugesh*
1 – 7
An Unusual Two-Step Hydrolysis of
Nerve Agents by a Nanozyme
nanozyme-mediated two-step hy-
drolysis of a nerve agent.
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