Intentional hydrolysis to overcome the hydrolysis problem: Detection of Ce(iv) by producing oxidase-like nanozymes with F-

Article abstract of DOI:10.1039/c9cc06167c

Polyvalent metal ions are susceptible to hydrolysis, making their reproducible detection by sensors and biosensors difficult. We herein turned this disadvantage into an advantage to develop a high selectivity colorimetric method for detecting Ce(iv) by intentionally hydrolyzing it through heating, where subsequent addition of F^- recovered the activity, allowing a detection limit of 3.8 μM Ce(iv).

Full text of DOI:10.1039/c9cc06167c

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Intentional hydrolysis to overcome the hydrolysis
problem: detection of Ce(IV) by producing
oxidase-like nanozymes with Fꢀ†
Cite this: Chem. Commun., 2019,
55, 13434
Received 8th August 2019,
Accepted 14th October 2019
ab
b
Yawen Wang,
Jianzhong Yang,a Yilin Zhaob and Juewen Liu
*
DOI: 10.1039/c9cc06167c
rsc.li/chemcomm
Polyvalent metal ions are susceptible to hydrolysis, making their and unstable aqueous chemistry even in strong acids.21,22 Such
reproducible detection by sensors and biosensors difficult. We herein hydrolysis makes reliable and reproducible measurements
turned this disadvantage into an advantage to develop a high selectivity difficult. We recently reported that Fꢀ can drastically improve
colorimetric method for detecting Ce(IV) by intentionally hydrolyzing the oxidase-like activity of CeO2 nanoparticles.23 This accelera-
it through heating, where subsequent addition of Fꢀ recovered the tion is unique to CeO2 among all the tested metal oxides.24
activity, allowing a detection limit of 3.8 lM Ce(IV).
Since the hydrolysis of Ce(IV) may lead to products similar to
CeO2, it might be possible to use this reaction to detect Ce(IV).
Analysis of polyvalent metal ions is often complicated by metal We herein intended to turn the disadvantage of Ce(IV) hydrolysis
hydrolysis, leading to time-dependent results and poor repro- into an advantage in efforts to distinguish it from other metal ions.
ducibility. A typical method to deal with this problem is using
We first reacted various metal ions (50 mM each) with a
strong acids to fully dissolve the hydrolyzed products. While chromogenic substrate 2,20-azino-bis(3-ethylbenzothiazoline-6-
this method is ideal for certain analytical instruments such sulphonic acid) (ABTS, 0.2 mM). The freshly prepared Ce(IV)
as inductively coupled plasma (ICP) spectroscopy, strong acids produced a green color, indicating oxidation of ABTS,25 while
are less desirable for sensors and biosensors. We reason that the Fe(III) sample also appeared slightly green (inset of Fig. 1A).
another possibility is to achieve full hydrolysis, which may also However, the other metals, including Ce(III) and Cu(II), did not
help avoid time-dependent changes in sample properties.
show much color change. This is reasonable since Ce(IV) is
Cerium is an important rare earth metal with rich redox and a strong oxidant, and Fe(III) has a lower oxidation potential.
catalytic activities.1–3 Cerium dioxide (CeO2) is a very useful We then performed the same reactions with 400 mM Fꢀ added.
material for catalysis4–7 and sensing applications.8–10 It has two The color of the Ce(IV) sample became even more intense, while
stable oxidation states: Ce(III) being similar to other trivalent the Fe(III) sample appeared less green. To quantitatively measure
lanthanides as a hard Lewis acid, whereas Ce(IV) is a strong them, we used UV-vis spectrometry and plotted the absorption
oxidant that is easily hydrolyzed in water. A lot of efforts have peak at 415 nm (Fig. 1). Indeed, Fꢀ promoted the activity of
been made for detecting cerium.11 For example, determination Ce(IV) but inhibited Fe(III), while none of the other metals had
of Ce(IV) was achieved through oxidation of organic compounds, any activity, regardless of Fꢀ. We then tested more metal ions,
such as arsenazo III,12 H2-receptor antagonists,13 tribromo- including Sc(III), Y(III), La(III), Pr(III), Nd(III), Sm(III), Eu(III), Gd(III),
arsenazo,14 sulphanilic acid,15 phenothiazine derivative,16 and Tb(III), Dy(III), Ho(III), Er(III), Tm(III), Yb(III), Al(III), Ca(II) and Mg(II).
ascorbate.17 However, these reactions are unlikely to be specific They also failed to oxidize ABTS even with Fꢀ (Fig. S1, ESI†). In this
and other oxidation reagents might also achieve similar results. regard, Ce(IV) is unique for its high activity for oxidation of ABTS,
Recently, Ce(III) was detected by promoting the catalytic activity and its selectivity was further increased by Fꢀ.
of gold nanoparticles.18 Biological probes for cerium, such as
DNAzymes, have also been reported.19,20
Ce(IV) has a strong tendency to hydrolyze, making the
reproducible measurement difficult. For example, after storing
freshly prepared Ce(IV) in water for 2 weeks, its activity towards
2+
Ce(IV) in water exists mainly as Ce(OH)3+ and Ce(OH)2
along with a dimer (Ce–O–Ce)6+, leading to time-dependent oxidation of ABTS was significantly decreased (comparing the
first and second sets of tubes in Fig. 1B). Interestingly, Fꢀ still
a College of Chemistry and Chemical Engineering, Taiyuan University of Technology,
boosted the oxidation activity of this aged Ce(IV) to a similar
level to the fresh Ce(IV). We further challenged the system by
Taiyuan 030024, China
b Department of Chemistry, Waterloo Institute for Nanotechnology, Water Institute,
heating freshly prepared Ce(IV) in water at 90 1C for 1 h and 6 h to
University of Waterloo, Ontario, N2L 3G1, Canada. E-mail: liujw@uwaterloo.ca
† Electronic supplementary information (ESI) available. See DOI: 10.1039/c9cc06167c further promote hydrolysis (the last two sets of tubes in Fig. 1B).
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Fig. 2 (A) Effect of pH on ABTS (0.2 mM) oxidation monitored at 415 nm
in the presence of 50 mM Ce901h without and with 400 mM Fꢀ. pH 1.29 to
3 was adjusted by HNO3, pH 4 and 5 were with acetate buffer (20 mM), and
pH 6 and 7 were with MES buffer (20 mM). (B) Effect of Fꢀ concentration
on ABTS (0.2 mM) oxidation in the presence of Ce901h (50 mM) in pH 4
acetate buffer. (C) Effect of various anions (400 mM each) on the oxidation
of ABTS by Ce(IV). The anions were incubated with Ce901h for 15 min
before measurement.
Fig. 1 (A) Oxidation of 0.2 mM ABTS (monitored at 415 nm) by various
metal ions (50 mM each) without and with Fꢀ (400 mM). Inset: The
corresponding photographs. (B) Photographs of reacting Ce(IV) (50 mM)
after various thermal treatments with 0.2 mM ABTS without and with Fꢀ
(400 mM). Effect of 400 mM Fꢀ on the kinetics of 0.2 mM ABTS oxidation
(monitored at 415 nm) in the presence of 50 mM (C) freshly prepared Ce(IV),
and (D) Ce(IV) heated for 1 h at 90 1C. The buffer was 20 mM acetate, pH 4.
After heating, the activity of ABTS oxidation was also reduced At pH 4, the effect of Fꢀ was the greatest. When the pH was
significantly. Yet, Fꢀ still brought the activity back. The color of higher than 6, the reaction was inefficient even in the presence
the products was quantitatively measured (Fig. S2, ESI†). In all of Fꢀ, since ABTS oxidation generally requires an acidic pH.
the cases, the samples with Fꢀ showed a similar level of activity. Thus pH 4 was used for most reactions.25 Therefore, the best
Therefore, Fꢀ can normalize the activity of hydrolyzed Ce(IV) with condition to show the effect of Fꢀ was pH 4.
a different extent of hydrolysis.
We then studied the effect of Fꢀ concentration. For 50 mM
We further measured the reaction kinetics. For the freshly Ce901h, we added up to 3 mM Fꢀ (Fig. 2B). Initially the activity
prepared Ce(IV), the reaction was very fast and most of the increased linearly with the logarithm of Fꢀ concentration and
reaction took place immediately after mixing (Fig. 1C, red saturated activity was observed at around 20 mM Fꢀ. After that,
trace). After that, although the reaction continued, the rate even 3 mM Fꢀ did not further increase or inhibit activity.
decreased significantly. With Fꢀ, the reaction finished in B1 min, Although the concentration of Ce(IV) was only 50 mM, after
where no color change was observed after that. Meanwhile, for the hydrolysis, the surface exposed cerium might be significantly less.
heat treated samples, the initial oxidation became much lower Therefore, to activate hydrolyzed Ce(IV), a sub-stoichiometric
(Fig. 1D). After adding Fꢀ, the extent of oxidation became similar amount of Fꢀ was sufficient.
to that of the freshly prepared Ce(IV), although the initial kinetics
To test if this reaction is specific for Fꢀ, we also added
were slightly slower. With Fꢀ, it took B2 min for the heated 9 types of other sodium salts to Ce901h (Fig. 2C). Only Fꢀ
sample to fully oxidize ABTS. The kinetics were overall quite robust promoted the activity. Na2S2O3 is a strong reducing agent and
with different heating temperatures (40 1C to 90 1C) and times could react with Ce(IV), resulting in a suppressed color change.
(0.1 to 6 h) (Fig. S3, ESI†). Although we cannot ensure that all Ce(IV) Phosphate also showed a significant inhibition effect, likely
in solution were hydrolyzed by the heat treatment, the insensitivity due to its strong affinity with Ce(IV), forming cerium phosphate
to the heating time and temperature is important for reproducible to inhibit the oxidation activity. EDTA also showed a weak
analytical assays.
inhibition effect. Therefore, this system has excellent selectivity
Since both hydrolysis of Ce(IV) and oxidation of ABTS are for Fꢀ. We previously showed that we can detect Fꢀ with CeO2.24
strongly pH-dependent,26–28 we then studied the effect of pH. The mechanism of action could be similar here, especially for the
We heated fresh Ce(IV) at 90 1C for 1 h and denoted this sample heat treated samples. Since directly using CeO2 might be more
Ce901h. Ce901h (50 mM) was then dispersed in buffers of convenient than using heat treated Ce(IV) for detecting Fꢀ, the
different pH. At very low pH (e.g. below 3), the activity was high analytical work here focused on the detection of Ce(IV).
regardless of Fꢀ (Fig. 2A), suggesting that the acid dissolved the
The change in absorbance after a 15 min reaction was
hydrolyzed products and recovered the activity.2 Thus, the plotted as a function of the concentration of Ce901h (Fig. 3A).
effect of Fꢀ was not obvious under very low pH conditions. With 200 mM ABTS, a linear relationship was obtained in the
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Fig. 3 (A) Effect of Ce901h (prepared by heating Ce(IV) at 90 1C for 1 h)
concentration on ABTS (0.2 mM) oxidation monitored at 415 nm in the
presence of 400 mM Fꢀ. (B) Selectivity test for the detection of Ce(IV). All the
metal ions were heated for 1 h. Concentration of ABTS: 0.2 mM, Fꢀ:
400 mM and the ABTS oxidation was performed for 15 min in 20 mM
acetate buffer, pH 4.
0–200 mM Ce(IV) range. The regression equation is DAbs =
0.00339 + 0.00823 [Ce(IV)] (mM), r2 = 0.996. The limit of detection
(LOD) was calculated to be 3.8 mM according to 3s/slope (s =
standard deviation of the blank signal). The relative standard
deviation (RSD) was 1.75% for the detection of 50 mM Ce(IV) (n = 9).
For the selectivity test, we measured 16 heated metal ions
(Fig. 3B). Only a high concentration of Fe3+ (500 mM) gave an
obvious signal, but the signal was still weaker than that of
50 mM Ce(IV), indicating high specificity. We also compared
50 mM Ce(IV) and its mixtures with various metal ions (Fig. S4,
ESI†). The coexistence of other metal ions (50 mM) had no
obvious influence on the activity of heated Ce(IV), although
500 mM Ca(II) showed some inhibition. Ce(III) showed no activity
and thus this method is specific for Ce(IV). One may still use it
for Ce(III) by first oxidizing it to Ce(IV) followed by hydrolysis.
To test its application in real samples, a water sample collected
from a contaminated site in Quebec was spiked with Ce(IV) (Fig. S5,
ESI†), and a detection limit of 2.1 mM Ce(IV) was obtained.
Fig. 4 Kinetics of 0.2 mM ABTS oxidation (monitored at 415 nm) of 50 mM
CeH1 incubated in 20 mM pH 4 acetate buffer (A) in the absence of Fꢀ for a
different time, then adding 400 mM Fꢀ and ABTS to initiate the reaction,
and (B) in the presence of 400 mM Fꢀ for a different time and then adding
ABTS to initiate the reaction. (C) Reaction of freshly prepared 50 mM Ce(IV)
and fresh Ce(IV) incubated with 400 mM Fꢀ for 2 and 12 h before reacting
with ABTS. (D) Effect of Fꢀ incubation time with 200 mg mLꢀ1 CeO2 for
oxidation of ABTS (0.5 mM). (E) A scheme showing the principle of heat
promoted hydrolysis and boosted activity by Fꢀ, while Fꢀ inhibited the
activity of free Ce(IV).
After showing the concept of intentional hydrolysis using some unstable partially hydrolyzed species. At last, we can
Ce(IV) as an example, we then wanted to understand the role of conclude that forming CeF4 was not the reason for the promoted
Fꢀ. Two possibilities exist: (1) breaking the hydrolyzed products activity of hydrolyzed Ce(IV). We then studied the extent of hydrolysis
to produce Ce(IV) ions and (2) acting on the hydrolyzed products by centrifugation and measuring the UV-vis spectrum of the super-
just like how Fꢀ acts on CeO2.29 To understand the mechanism, natant (Fig. S7, ESI†). It appeared that most of the Ce(IV) remained
we designed the following experiments. First, Ce901h was in the supernatant and thus the hydrolyzed products were not large
respectively incubated in pH 4 acetate buffer for 0, 1.5, 3 and particles.
12 h before Fꢀ and ABTS were added (Fig. 4A). The kinetics of
Finally, we did the same experiment with CeO2 nanoparticles.
the reactions were all very fast and showed no obvious difference Fꢀ promoted the activity of CeO2 to a similar extent regardless of
between these samples. Without Fꢀ, however, even storing Ce901h the pre-incubation time with Fꢀ (Fig. 4D). The kinetic trend of the
at pH 4 for 12 h still resulted in low activity (Fig. S6, ESI†). We then heated Ce901h sample (Fig. 4B) behaved between those of Ce(IV)
incubated Ce901h at pH 4 with Fꢀ for various periods of time before and CeO2. Therefore, we reason that the hydrolyzed Ce(IV) did not
adding ABTS (Fig. 4B). Interestingly, the reaction rate decreased with form rigid CeO2, and thus can still be slowly disrupted by Fꢀ. The
longer pre-incubation of Fꢀ.
mechanism of promotion should be similar for the hydrolyzed
After adding Fꢀ to freshly prepared Ce(IV) and immediately Ce(IV) and CeO2. The hydrolyzed Ce(IV) had a more stable response
testing it, its activity slightly increased (Fig. 1C). However, when than freshly prepared Ce(IV) upon incubation with Fꢀ (comparing
fresh Ce(IV) and Fꢀ were incubated for 2 h, its activity was the 12 h incubation data in Fig. 4B and C). Therefore, intentional
significantly decreased, and was nearly fully inhibited after hydrolysis can produce more robust sensors. The effect of hydro-
overnight incubation with Fꢀ (Fig. 4C). Fꢀ can form stable lysis and Fꢀ is summarized in Fig. 4E.
complexes with Ce(IV),30–32 and such complexes are inactive.
In conclusion, we proposed the concept of intentional
The kinetics of the complex formation appeared to be slow, yet hydrolysis of polyvalent metal ions to achieve a more stable sensor
adding Fꢀ had a transient promoting effect, likely by breaking response. Using this concept, we have developed a highly selective
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colorimetric method to detect Ce(IV) based on the effect of Fꢀ to
boost the oxidation ability of hydrolyzed Ce(IV). Compared to other
sample treatment approaches using strong acids, this detection
system can avoid strong acidic conditions. The products of hydro-
lysis for other metal ions might also have their own unique
properties to be explored for their detection. In particular, the
concept of nanozymes can be explored for these metal containing
species.27,28,33–37
We thank GeoSyntec Consultants (Guelph, Canada) for
supplying the environmental water sample and Miss Kellie
Chong for proofreading this paper. Funding for this work was
from the Natural Sciences and Engineering Research Council of
Canada (NSERC) and the Canada First Research Excellence
Fund (CFREF) for the Global Water Future project. Y. W. was
supported by the Shanxi Scholarship Council in China to visit
the University of Waterloo.
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