Unique dual responsive activity of a platinum nanozyme stabilized by a green solvent: Deep eutectic solvents

Article abstract of DOI:10.1039/c8nj01902a

Cubical Pt nanozymes with a size distribution of 6-8 nm synthesized in and stabilized by eco-friendly choline chloride/glycerol deep eutectic solvent displayed outstanding catalytic performance in the oxidation of uric acid with an activation energy of 37.473 kJ mol^-1 and decomposition of en route generated H₂O₂. Significantly, the exploited one pot nanozyme cascade reaction has proven the novel uricase and peroxidase dual responsive enzyme mimetic activity of the ensuing Pt nanocubes unravelling future prime applications.

Full text of DOI:10.1039/c8nj01902a

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Unique dual responsive activity of a platinum
nanozyme stabilized by a green solvent: deep
Cite this:DOI: 10.1039/c8nj01902a
eutectic solvents†
Henam Sylvia Devia and Henam Premananda Singh
Received 20th April 2018,
Accepted 22nd June 2018
b
*
DOI: 10.1039/c8nj01902a
rsc.li/njc
Cubical Pt nanozymes with a size distribution of 6–8 nm synthe- In fact, a survey of the literature manifests the availability of very
sized in and stabilized by eco-friendly choline chloride/glycerol limited resources on the mimicking behaviour of uricase or urate
deep eutectic solvent displayed outstanding catalytic performance oxidase enzyme that plays a pivotal role in the chain of enzymes
in the oxidation of uric acid with an activation energy of responsible for the metabolism of purines.11,12 Nonetheless, it
37.473 kJ molÀ1 and decomposition of en route generated H2O2. remains a great challenge to achieve highly active nanomaterials
Significantly, the exploited one pot nanozyme cascade reaction has based enzyme mimetic activity. Often, it is emphasized that the
proven the novel uricase and peroxidase dual responsive enzyme synthesis methodology adopted remains a salient element for
mimetic activity of the ensuing Pt nanocubes unravelling future the successful application of nanoscale particles. Furthermore,
prime applications.
worldwide environmental cognizance has ceaselessly propelled
scientists in search of ‘‘greener solvents’’ for use in chemistry
The discovery of ribozymes, in the early 1980s, marked a con- and chemical technology. Veritably, deep eutectic solvents (DESs)
ceptual turning point in enzyme studies and immense interest are one of such alternative solvents and are a briskly emerging
was sparked in artificial enzyme mimics.1 The last decade new class of green solvent liquids with fascinating properties of
has witnessed unprecedented growth in the realm of artificial minimum volatility, non-toxicity, biodegradability, suitability and
enzymes based on nanomaterials known as nanozymes. Nano- low cost.13,14 They are an extended class of ionic liquids which are
zymes offer improved properties relative to natural enzymes in simply prepared by mixing Lewis or Bronsted acids and bases
several aspects such as robustness to a harsh environment, low containing a variety of anionic and/or cationic species.15,16
price, easy preparation, mass production, simpler purification Interaction of a hydrogen bond donor with the quaternary salt
and storage, better stability as well as tunable catalytic in DES subsequently abated the electrostatic forces between the
potential.2,3 Such nanozymes which are expected to link nano- anion and the cation leading to the formation of a eutectic
materials with biological systems and functions opened a mixture with considerably lower melting point and intrinsic
promising window of biosensing, imaging, anti-biofilm, cancer viscosity than those of the initial components.17 Furthermore,
diagnostics, neuroprotection, stem cell growth, pollutant removal strong associative properties of DES through hydrogen bonds
and catalysis applications.4–6 Hitherto, varieties of metal, metal among the components make it a potent stabilizing agent.
oxide, polymer and composite nanodimensional particles have Owing to their promising properties, numerous research
been explored to resemble catalase, superoxide dismutase, studies were carried out using DES as the solvent but so far
peroxidase and other oxidases.3,7–9 Qingqing Wang et al.10 few studies have been reported on the synthesis of NPs utilizing
developed nickel–palladium hollow nanoparticles (NPs) exhi- DES. Monodispersed star-like Au NPs bounded with low index
biting triple enzyme mimetic activity for wide linear range (111) facets have been reported by Hong G. Liao et al.18 using
detection of glucose with a low detection limit but was limited DES in the presence of L-ascorbic acid at room temperature.
to familiar glucose oxidase, peroxidase and catalase like activity. Au NPs of various shapes and surface structures including
snowflake-like NPs and nanothorns were also obtained by
adjusting the content of water in the DES. Recently, solvothermal
synthesis of profoundly active 1-D ceria nanostructures in the
DES of choline chloride (ChCl)/urea for catalytic CO oxidation
was described by Oliver S. Hammond et al.19 Additionally,
Karimi et al.17 elucidated the superior fabrication of calcium
a Functional Materials and Device Lab, Department of Electrical Engineering,
IIT Delhi, Delhi-110016, India
b Department of Chemistry, National Institute of Technology, Manipur-795001,
India. E-mail: henam_boynao@yahoo.co.in; Tel: +91 8974303534
† Electronic supplementary information (ESI) available. See DOI: 10.1039/c8nj01902a
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phosphate NPs in deep-eutectic ChCl/urea medium compared to
water concerning their crystalline behavior and control of
particle size.
Inspired by these studies by eminent scientists, we have
explored a novel route to tailoring ultra small cubical Pt NPs in
the DES of ChCl/glycerol utilizing L-ascorbic acid as a green
reducing agent gratifying the basic principles of green chemistry.
It is noteworthy that in spite of the challenge of manipulating
the chemical behavior of NPs at an atomic scale, the synthesized
ultra small Pt NPs unambiguously possess excellent uricase
and peroxidase enzyme mimetic activities. To the best of our
knowledge, this artificial dual responsive enzyme mimicking
behavior of Pt nanocubes is reported for the first time paving a
promising path that can be capitalized on successfully in indus-
trial catalysis and biomedical fields. Indeed, this report further
expands the repertoire of such solvents as sustainable media for
tailoring appealing nanoscale particles with participation in the
emerging field of artificial enzymes.
In this work, a bluish black solution of Pt particles stabilized
by a DES of ChCl:glycerol exhibiting a typical plasmonic excita-
tion band at 257 nm has been successfully tailored as shown in
Fig. S1 (ESI†). Nanoscale Pt particles are known to display
absorption spectra in the entire range of the ultraviolet-visible
(UV-vis) spectrum and the characteristic band in this region
was in good agreement with early investigations.20,21 The high
viscosity compared to other common solvents such as water
and the latent supramolecular catalyst role of the DES were
expected to play a crucial fundamental role during the for-
mation of NPs.19,22 The formation of particles was succinctly
confirmed using the recorded scanning electron microscopy
(SEM) image displayed in Fig. 1(A).
The particles primarily appeared to exhibit rough surfaces,
edges and kinks to some extent. Nevertheless, no discernable
morphology can be observed from the obtained image. In fact,
the size and morphology of the ensuing Pt particles were
analyzed by transmission electron microscopy (TEM), and the
result is depicted in Fig. 1(B). A close inspection reveals that the
shape of all Pt particles is nearly cubical with a narrow size
distribution of 6–8 nm. The nanodimensional cubical particles
appear to form a halo over their clusters stabilizing them due to
the DES under these synthesis conditions, see Fig. S2 (ESI†).
Powder X-ray diffraction (XRD) analysis illustrated the perfectly
crystalline nature of the particles; no phase interference or any
other impurities were detected, as shown in Fig. 1(C). The
prepared Pt nanocubes belonged to the fcc crystal lattice with
the (111), (200), (220), (311) and (222) facets corresponding to
2y diffraction values of 39.9251, 46.41, 67.6051, 81.3791 and
85.8791 respectively. The broad and sharp peaks in the pattern
further substantiated the nanocrystalline nature of the synthe-
sized particles. In addition, the interesting information gained
from their XRD data was that the peak intensity ratios of the
(200), (220) and (311) crystal facets with respect to the most
intense (111) peak were 0.40, 0.28 and 0.26 respectively.
Meanwhile, the respective ratios for the bulk standard values
are 0.30, 0.16 and 0.16 which preliminary established better
exposure of the active (200), (220) and (311) surfaces relative to
Fig. 1 (A) SEM, (B) TEM image, and (C) XRD pattern of the as-synthesized
Pt particles. (D) Size histogram of the particles. [Synthesis experiment:
A clear, homogenous colorless solution of ChCl: glycerol DES was prepared
by heating 12 g of ChCl in 12.5 mL of glycerol at 80 1C with constant stirring.
0.05 g of PtCl2 + 1 mL of DES + 1 mL of 20% L-ascorbic acid solution
prepared in DES was stirred for 2 h at 60 1C for formation of NPs].
the (111) surface in the as-synthesized Pt nanocubes compared
to bulk Pt particles.23 Besides, Fig. 1(D) highlights the size
distribution histogram of the aforementioned Pt nanocubes
with an average particle size of 6 nm.
The FT-IR spectrum of the DES liquid sample is shown in
Fig. 2 giving the following information: the band in the 3000–
3500 cmÀ1 region was attributed to OH stretching vibrational
frequencies and the presence of a H-bond, the band in the
1020–1200 cmÀ1 region was attributed to C–OH stretching, the
band in the 1300–100 cmÀ1 region was attributed to C–-N
stretching, and also the band located at around 1440–1465 cmÀ1
was attributed to CH3 bending. Additionally, the bands around
500–1305 cmÀ1 were also reported due to the formation of
H-bonds.24 Most of the characteristic bands of the DES were
noticed in the spectrum of the prepared Pt nanocubes, as shown
in Fig. 2(B). Interestingly, in the case of the aerially dried particles
Fig. 2 Staking FT-IR plots of (A) DES and (B) aqueous dispersed Pt
nanocube samples.
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the signature band of OH appeared to be shifted slightly toward a signifies the plausible applications of the tailored Pt nanozyme
longer wavelength compared to the DES suggesting the interaction under higher temperature conditions in contrast to natural
of the DES with the particle’s surface through OH groups. The uricase enzyme, which exhibits an optimal temperature of
overall observations clearly evidenced the presence of the DES 40 1C.25
on the surface of the nanocubes stabilizing them from further
changes. Moreover, we observed that our colloidal nanocubes the corresponding Pt nanocube catalyzed oxidation reaction of
dispersed in DDW remained unsettled for more than a week.
UA from the linear plot of Àln kapp vs. 1/T based on the
An activation energy of 37.473 kJ molÀ1 was estimated for
To investigate the dual responsive enzyme-like activity of Arrhenius equation, as displayed in Fig. 3(D). Yongqiang Dong
the Pt nanocubes, a single pot cascade type reaction was carried et al.11 reported an activation energy of 61.73 kJ molÀ1 and a
out systematically. Of note, initially under mild experimental rate constant which was almost 29 000 times higher in the
conditions, the Pt nanocubes could quickly catalyze the oxida- absence of 4 to 5 nm Pt particles for a similar UA oxidative
tion of uric acid (UA) as illustrated in Fig. 3(A) indicating degradation reaction. Thus, our Pt nanocubes could dramati-
intrinsic uricase-like activity. In this real time analysis, notably cally accelerate the reaction rate by facilitating a lower energy
the two characteristic absorption bands of UA at 231 nm pathway. This property of small size Pt nanocubes may serve as
and 285 nm alleviate subsequently with time and finally a rostrum for the treatment of hyperuricemia cases that can
disappeared within 3 h without concomitant appearance of any lead to gout, obesity, diabetes, high cholesterol, high blood
absorption bands. In our case, UA almost remained unaltered in pressure, kidney disease, hypertension, and cardiovascular
the absence of nanocubes during the incubation time constraint disease.26 Meanwhile, in order to corroborate the peroxidase
(data not reported). The experiment was repeated thrice for mimetic activity of the ensuing cubical nanodimensional Pt
reproducibility. Next, a set of steady state kinetics experiments particles further, a typical cascade reaction of the above UA
was carried out to study the dose dependence of nanocube oxidation reaction was adopted. The reaction was examined
loading and temperature influenced by the oxidation of UA by utilizing 200 mL of horse radish peroxidase enzyme substrate
noting the fall in absorbance at 285 nm, a prominent charac- o-phenylenediamine (OPD, 1 Â 10À2 M) in the presence of
teristic band of UA. Under identical reaction conditions, it was 1500 mL UA (1 Â 10À4 M) and 500 mL Pt nanocubes
found that the first order initial reaction rate escalated with (0.3 mg mLÀ1). The total volume of the reaction mixture was
an increase in Pt nanocube dose, as shown in Fig. 3(B). maintained at 3 mL using DDW. The progress of the reaction
Compellingly, the experimental results also manifested that was also monitored in real time using UV-vis spectroscopy, as
the apparent rate constants (kapp) obtained from the plot of shown in the time evolution curves (Fig. 4) at room temperature
lnA285 vs. time increased with temperature in the range from by following the enhancement of the band at 425 nm, which
20 1C to 60 1C, as presented in Fig. 3(C). Verily, this result originated from the OPD oxidized product 2,3-diaminophenazine
(DAP).27 En route liberated hydrogen peroxide (H2O2) during the
oxidation of UA to 5-hydroxyisourate (5HIU) subsequently inter-
acted with OPD in the presence of Pt nanocubes to form its
yellowish oxidized product. For further verification, a controlled
experiment was performed utilizing H2O2 and OPD directly in the
absence and presence of the as-synthesized Pt nanocubes. Under
the standard conditions employed, the matched characteristic
peak of DAP at 425 nm for the Pt nanocube catalyzed reaction
elevated rapidly compared to the uncatalyzed reaction that was
trivial, as presented in Fig. S3 (ESI†).
Fig. 3 (A) Time dependent UV-vis spectra for oxidative degradation of UA
at 35 1C. The reaction mixture consists of 1.5 mL UA (1 Â 10À4 M), and
500 mL Pt nanocubes (0.3 mg mLÀ1). The total volume of the reaction
system was maintained at 3 mL with DDW. (B) Dose response plot of
reaction rate with amount of NPs introduced in the reaction system at
35 1C. (C) Plot of ln A285nm vs. time at different temperatures ranging from
20 1C to 60 1C. Concentration of UA: 1 Â 10À4 M (1.5 mL); concentration of
Pt nanozyme: 0.3 mg mLÀ1 (500 mL); and DDW (1 mL). (D) Relationship
between the temperature of the reaction system and the apparent rate
constant for the UA oxidative degradation reaction.
Fig. 4 UV-vis absorption time course curves for the peroxidase mimicking
activity of Pt nanocubes; inset shows the reaction scheme and digital image of
the reaction product.
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multiple washing and centrifugation processes involved during
the test. Vividly, our experimental results for the uricase and
peroxidase dual responsive enzyme mimetic activity of Pt
nanocubes evinced its uniqueness and was beneficial considering
parallel reports on the basis of excellent performance, recyclability,
robustness to the environment, cost effectiveness and the lack
of auxiliaries since such a reported archetypal one pot cascade
reaction for other nanoscale materials occurred generally in
combination with the natural uricase enzyme.32,33
In summary, we have used the notable ecologically and
economically profitable DES of ChCl/glycerol for tailoring exciting
functional nanomaterials. The generated cubical Pt NPs belong to
the fcc crystal system with a narrow size distribution and an
average size of 6 nm. In particular, there is no doubt that the DES
of ChCl/glycerol, besides being a legitimate greener solvent, is also
Fig. 5 Recyclability test for Pt nanozyme in the oxidation of UA.
Experimental conditions: 1.5 mL of UA (1 Â 10À4 M), 300 mL of nanozyme
(0.3 mg mLÀ1) and 1.2 mL of DDW at 35 1C.
This ultimately well concluded significant intrinsic peroxidase a perspective stabilizing agent for nanoscale particles. We have for
behaviour of the synthesized ultra small cubical Pt nanozyme and the first time demonstrated the efficient uricase and peroxidase
additionally suggested remarkable competence for the effective dual enzyme mimetic activity of ultra small Pt nanocubes
decomposition of H2O2, a reactive oxygen species (ROS). exercising a one-pot nanozyme cascade reaction. The corres-
We speculated that the relatively large surface area-to-volume ponding activation energy for Pt nanocube catalyzed oxidative
ratio of the ultra small size Pt nanocubes and their strong Lewis degradation of UA was found to be 37.473 kJ molÀ1. The key to
acid properties initiated the oxidation reactions making Pt nanozyme activity was reckoned to be its small size with
them catalytically more efficient.28,29 Plus, facet-dependent fascinating morphology, strong Lewis acid behaviour and exposed
adsorption, activation, and rearrangement processes between facets. Indeed, this report not only partook in the next wave of
the reacting species and the exposed crystal facets of the artificial enzyme research but also offered significant insights for
cubical NPs might have played a crucial integrated role in their future applications in environmental chemistry, biotechnology,
enzyme mimicking activities as noticed by Yushi Yang et al.7 in industrial catalysis, sensing, chemopreventive agents and multi-
the case of exposed facets of CeO2 nanostructures mimicking drug therapy for biomedical purposes.
the redox enzyme. In our case, 5HIU and allantoin were
contemplated not to be the end product of the oxidation
reaction as none of their characteristic absorption band(s) at Conflicts of interest
around 292 nm and 216 nm, respectively, were observed.26,30
The exact oxidative degradation product of UA by the Pt
There are no conflicts to declare.
nanozyme was not known but it was anticipated to form
2-amino-2-(carbamoylamino)-2-hydroxyacetic acid via the for-
mation of 5HIU and H2O2 since the observed absorption
Acknowledgements
spectrum of our oxidative degradation product was in good
The authors are grateful to the Director, the National Institute
of Technology, Manipur, India, for his encouragement and
support in the form of financial assistance. We are also thankful
concordance with the available literature.11 Fig. S4 (ESI†)
demonstrates a plausible mechanism for the uricase mimicking
behaviour of the Pt nanozyme. Apparently, further extensive
to Dr Th. David Singh, the National Institute of Technology,
investigations through various techniques are needed to under-
Manipur, India, for facilitating the UV-vis spectrophotometer
stand and enunciate this notion. Nonetheless, the natural
experiments. Furthermore, we highly appreciate AIIMS, New Delhi,
peroxidase-like activity of the prepared Pt nanocubes for the
India, for recording our TEM images.
oxidation of OPD in the presence of the liberated H2O2 can be
attributed to the generation of an OHꢀ radical from the decom-
position of H2O2, as displayed in Fig. S5 (ESI†). Adsorption of
H2O2 molecules on the surface of nanoscale particles occurred
Notes and references
and the O–O bond of H2O2 might be broken up into a double OHꢀ
radical. Partial electron exchange interaction stabilized the
generated OHꢀ radical by Pt nanocubes which supplemented
their catalytic ability.28,31 Fig. 5 shows the recyclability test of
the Pt nanozyme for the oxidative degradation of UA. It was found
that 88.7% of catalytic activity was profoundly retained even after
the fifth cycle evincing the excellent catalytic performance of the
tailored Pt nanocubes. Such mitigation in catalytic behaviour after
repeated cycles might be due to the loss of particles during
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