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CoO/reduced graphene oxide nanocomposites as effective phosphotriesterase mimetics for degradation and detection of paraoxon Ting Wang, Jiangning Wang, Ye Yang, Ping Su, and Yi Yang Ind. Eng. Chem. Res., Just Accepted Manuscript • DOI: 10.1021/acs.iecr.7b02223 • Publication Date (Web): 07 Aug 2017 Downloaded from http://pubs.acs.org on August 8, 2017

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Industrial & Engineering Chemistry Research

Co3O4/reduced graphene oxide nanocomposites as effective phosphotriesterase mimetics for degradation and detection of paraoxon

Ting Wang, Jiangning Wang, Ye Yang, Ping Su*, and Yi Yang*

Beijing Key Laboratory of Environmentally Harmful Chemical Analysis, College of Science, Beijing University of Chemical Technology, Beijing 100029, China

*E-mail: [email protected]；[email protected]

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ABSTRACT: In this work, Co3O4 nanoparticles supported on reduced graphene oxide (Co3O4/rGO) were prepared through a simple hydrothermal method. The phosphotriesterase (PTE) mimetic activity of the Co3O4/rGO nanocomposites was investigated for the first time, and the catalytic performance was evaluated by hydrolyzing paraoxon. The synergic effects of Co3O4 and rGO greatly improved the binding capacity of the nanocomposites with paraoxon and water molecules, and facilitated electron transfer in the hydrolysis process, resulting in an approximately five-fold enhancement in the hydrolysis efficiency of Co3O4/rGO nanocomposites compared with that of rGO. Co3O4/rGO nanocomposites also exhibited excellent reusability and stability than other PTE mimetics. Based on the PTE activity of Co3O4/rGO nanocomposites, a simple and sensitive colorimetric sensor of paraoxon was developed and successfully used to determine the paraoxon in cabbage and river water, demonstrating its potential applicability for food and environmental analyses.

1. INTRODUCTION Organophosphate triesters (e.g., paraoxon, parathion, and malathion) have been widely used as plasticizers, flame retardants, pesticides, and chemical warfare agents. 1-3 However, these compounds are extremely toxic to mammals because of their irreversible inhibition of acetylcholine esterase, which contributes to the accumulation of the neurotransmitter, ultimately resulting in nerve failure, paralysis, and eventually death.4-6 Moreover, organophosphate triester residues in the environment also cause significant damage to the ecosystem as well as prodigious agricultural and environmental pollution.7 There is thus an urgent need for the development of highly sensitive detection techniques as well as efficient detoxification methods for these compounds. 2


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Phosphotriesterase (PTE, EC 3.1.8.1), a binuclear zinc enzyme, which can hydrolyze the organophosphate triesters into less toxic diesters, has been extensively studied for biosensing and biodegradation of organophosphate triesters.8-10 Although native PTE exhibits excellent performance in most cases, the activities of PTE can be attenuated by denaturation and autodigestion during their application and storage, leading to poor stability as well as low recovery and reusability.11,12 Alternatively, artificial mimetic enzymes with PTE activity have grasped great attention because of their high stability and excellent catalytic activity. Recently, a large number of inorganic and metal materials, such as metal complexes, heterogeneous catalysts, metal organic frameworks, and some nanomaterials have been shown to be capable of hydrolyzing organophosphorus triesters.13-29 However, in most of these cases, the catalytic activity and efficiency of these materials are hindered by the reaction products, which may block part of the active sites, thereby limiting the catalytic activity or leading to irreversible hydrolysis.28 Therefore, development of a novel PTE mimetic enzyme for efficient degradation and detection of organophosphorus triesters is of significant importance. Graphene-supported cobalt/cobalt oxide nanohybrids have exhibited remarkable catalytic activities in the oxidation of CO, hydrogenation of CO2, degradation of phenol, hydrolysis of ammonia borane, oxygen reduction reaction, and oxygen evolution reaction, etc.30-40 This advantageously catalytic performance is ascribed to the synergic effects of graphene and cobalt/cobalt oxides. Although these nanohybrids provide satisfactory catalytic activities in most cases, to our best knowledge, the PTE mimetic catalytic activity of graphene-supported cobalt/cobalt oxides has never been reported. In this study, we synthesized Co3O4/rGO nanocomposites via a facile hydrothermal 3



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method and investigated their PTE activities for the first time. The catalyst performance of the nanocomposites was evaluated by the hydrolysis of paraoxon, and the results indicate that the Co3O4/rGO nanocomposites possessed excellent hydrolysis efficiency even in the presence of a high concentration of diethyl phosphate, which is one of the main hydrolysis products. In addition, the Co3O4/rGO nanocomposites exhibited excellent reusability and stability in the hydrolysis of paraoxon, and the Co3O4/rGO nanocomposites showed an approximately five-fold enhancement in the catalytic efficiency compared with that of rGO, which was attributed to the synergic effects of Co3O4 and rGO. The effect of the rGO content in the Co3O4/rGO nanocomposites on the catalytic activity was also examined, and the catalytic mechanism was explored in detail. Based on the PTE mimetic activity of the Co3O4/rGO nanocomposites, we developed a simple and sensitive colorimetric sensor of paraoxon and provided a potential degradation pathway.

2. EXPERIMENTAL SECTION 2.1 Materials. All of the chemicals were of analytical grade and used without further purification. Graphite and Co(CH3COO)2·4H2O were purchased from Alfa Aesar Co. Ltd. (Massachusetts, USA). Triethylamine, NH3·H2O (28 wt%), KMnO4, NaNO3, NaOH, Na2CO3, NaHCO3, H2SO4 (98%), HCl (37.5%), and H2O2 (30%) were supplied by Beijing Chemical Works (Beijing, China). N-methylmorpholine (NMM) and diethyl p-nitrophenyl phosphate (paraoxon) were purchased from Sigma–Aldrich (Shanghai, China). The paraoxon was stored in the dark to avoid photolysis and was diluted with ethanol before use. For safety, direct contact and inhalation of paraoxon should be avoided by taking appropriate precautions and operating in a fume hood. Deionized water was used in all of the experiments. 4


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2.2 Synthesis of graphene oxide. Graphene oxide (GO) was synthesized from natural graphite powder using a modified Hummers method.41 Briefly, concentrated H2SO4 (92 mL) was added to a mixture of graphite powder (4 g) and NaNO3 (2 g). With vigorous stirring, KMnO4 (12 g) was slowly added to the mixture to keep the reaction temperature below 20 °C. The mixture was stirred vigorously for 1 h and then warmed to 35 °C for 1.5 h. After that, deionized water (100 mL) was slowly added, producing a large exotherm to 98 °C. External heating was introduced to maintain the reaction temperature at 98 °C for 30 min, and then additional deionized water (200 mL) was added. The mixture was subsequently washed with H2O2, HCl (5%), and deionized water to remove remaining metal ions and the acid. Following drying under vacuum for several hours, the resulting products were ground into powders. 2.3 Synthesis of Co3O4/rGO nanocomposites. Co3O4/rGO nanocomposites were synthesized using a general two-step method.31 In the first step of the Co3O4/GO hybrid synthesis, Co3O4 nanoparticles (NPs) were mildly grown on the GO sheets via the hydrolysis and oxidation of Co(OAc)2. In detail, 59.5 mg of GO was dispersed in 72 mL of anhydrous ethanol using ultrasonication. Co(OAc)2 aqueous solution (3.6 mL, 0.2 M), NH3·H2O (1.5 mL), and H2O (2.1 mL) were sequentially added to the above GO/ethanol dispersion with intense stirring at room temperature. Then, the mixture temperature was maintained at 80 °C with vigorous stirring for 10 h. In the second step, the reaction mixture was transferred to an autoclave for hydrothermal reaction at 150 °C for 3 h, leading to the crystallization of Co3O4 and reduction of GO. After cooling to room temperature, the synthetic product was collected by centrifugation, thoroughly washed several times with ethanol and deionized water, and then dried under vacuum at 60 °C for 4 h. The rGO and Co3O4 NPs were synthesized using the same procedure without using Co salt or GO, respectively. The synthesis conditions of Co3O4/rGO nanocomposites with different reactant ratios of Co3O4 and rGO are provided in the Supporting Information. 5



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2.4 Characterization. The particle sizes and morphologies were investigated using scanning electron microscopy (SEM; Zeiss Supera55, Oberkochen, Germany) and transmission electron microscopy (TEM; Philip Tecnai 20, Amsterdam, Netherlands). The crystal structures were investigated using X-ray diffraction (XRD; Rigaku D/MAX-2500, Tokyo, Japan) with Cu Kα radiation (α = 1.54178 Å). Fourier-transform infrared (FT-IR) spectra were collected on a Nicolet spectrometer (Thermo Fisher Scientific, Waltham, USA) using the KBr disk method. The elemental compositions were determined using an elemental analyzer (EA; Vario EL Cube, Elementar, Hanau, Germany) and X-ray photoelectron spectroscopy (XPS; Thermo VG ESCALAB-250, Massachusetts, USA) with an Al Kα radiation source. Thermogravimetric analysis (TGA) was conducted using a Mettler Toledo thermal analyzer (1100SF, Columbus, USA) that was adapted to a nitrogen purge gas at a heating rate of 10 °C min-1. Dynamic light scattering (DLS) measurements were performed in ethanol at 25 °C using a Zetasizer (Malvern Zetasizer Nano-ZS, Malvern, UK). Absorption spectra were recorded on a TU-1950 spectrophotometer (Beijing Purkinje General Instrument Co., Ltd., Beijing, China). 2.5 Catalyst performance. To evaluate the catalytic activity for the hydrolysis of paraoxon, batch experiments were performed as follows. Co3O4/rGO nanocomposites (6 mg) were added to a Na2CO3–NaHCO3 buffer (1 mL, pH = 10.5, 0.1 M) in a glass vial and then stirred for 5 min to achieve a homogeneous dispersion. Next, paraoxon (1.27 mg) was added to the mixture solution and swirled at 60 °C. Periodic monitoring was performed by removing a 10 µL aliquot from the reaction mixture and diluting it with the buffer (0.99 mL) before ultraviolet–visible (UV–vis) spectroscopy measurements (the formation of p-nitrophenolate was studied at 401 nm) and 31P NMR analysis (the hydrolysis of paraoxon was monitored at –7.8 ppm, and the formation of diethyl phosphate occurred at –0.3 ppm). Steady-state kinetics experiments were performed, and the effect of the buffer, reaction temperature, and time were 6


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evaluated using a similar procedure. The experimental procedure used to investigate the effect of the presence of hydrolysis product (diethyl phosphate) on the catalytic activity is provided in Supporting Information. 2.6 Detection of paraoxon and assessment of practicability. Paraoxon was detected using colorimetric analysis as follows: (a) Co3O4/rGO nanocomposites (6 mg) were added to a Na2CO3–NaHCO3 buffer (1 mL, pH = 10.5, 0.1 M) in a glass vial and then stirred for 5 min to achieve a homogeneous dispersion. (b) Paraoxon samples (100 µL) were added to the mixture solution and swirled for 20 min at 60 °C. (c) 10 µL of the supernatant liquid was removed and diluted with the buffer (0.99 mL) for UV–vis spectroscopy measurements at 401 nm.

3. RESULTS AND DISCUSSION 3.1 Characterization of catalysts. The surface morphology and size characteristics of the as-prepared Co3O4/rGO nanocomposites were investigated using SEM, TEM, and energy-dispersive X-ray spectroscopy (EDS). The SEM image (Figure 1) shows that the rGO sheets resembled silk veil wave and exhibited the typical wrinkle morphology. The TEM images confirm the presence of large rGO sheets homogeneously decorated with well-dispersed Co3O4 nanocrystals (3–5 nm in size). A lattice structure is observed in the high-resolution TEM image, and the lattice space of Co3O4/rGO nanocomposites are measured with a spacing of 0.24 nm, which is consistent with the theoretical spacing of the (3 1 1) planes of Co3O4 nanocrystals.

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Figure 1 (a) Low-magnification and (b) high-magnification TEM images of Co3O4/rGO nanocomposites. (c) SEM image of Co3O4/rGO nanocomposites. (d) EDS elemental mapping of C, O, and Co of Co3O4/rGO nanocomposites.

The structure of the Co3O4/rGO nanocomposites was examined using XRD, Raman spectroscopy, and FT-IR spectroscopy. The XRD patterns of the Co3O4/rGO nanocomposites (Figure 2a) contain (111), (220), (311), (400), (511), and (440) diffraction peaks, which can be indexed as cubic-phase Co3O4 (JCPDS #43-1003). The results therefore confirm the crystalline nature and phase purity of the Co3O4 NPs in the Co3O4/rGO nanocomposites. The Raman spectra of the rGO and Co3O4/rGO nanocomposites (Figure 2b) show two prominent peaks at 1354 and 1594 cm−1, which are attributed to the typical D and G bands of graphene, respectively. The other Raman peaks at 464 and 513 cm−1 are assigned to the F2g modes of Co3O4, and the peak at 693 cm−1 is assigned to A1g mode. These results clearly confirm the

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existence of both graphene and Co3O4 in the as-prepared composites. FT-IR spectra of the rGO, Co3O4 NPs, and Co3O4/rGO nanocomposites (Figure 2c) were obtained to further investigate the structure of the nanocomposites. The broad absorption peak at 3420 cm–1 corresponds to the –OH group, and the peak at 1620 cm–1 is attributed to O–H bending vibration of absorbed water molecules and contributions from the vibration of aromatic C=C. The peaks at 660 and 570 cm–1 are assigned to Co–O vibrations. The FT-IR analysis further confirms that Co3O4 NPs were successfully deposited on the surfaces of rGO.

Figure 2 (a) XRD patterns, (b) Raman spectra, (c) FT-IR spectra, and (d) TGA profile of Co3O4/rGO nanocomposites.

The composition and surface electronic states of the Co3O4/rGO nanocomposites were examined using TGA and XPS. The mass ratio of graphene in the Co3O4/rGO nanocomposites was determined using TGA (Figure 2d). The initial decline of the TG curve below 100 °C 9



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corresponds to the weight loss of water (2.34 wt%). In the following step, a sharp drop stage (39.02 wt%) is observed, corresponding to the removal of graphene from the composites. Therefore, the results suggest that the composites contained 58.64 wt% Co3O4 loading. The XPS survey spectra (Figure S1a) reveals the presence of Co, O, and C in the composites with no other impurities. The regional Co 2p spectrum in Figure S1b shows two major peaks with binding energies at 797.5 eV (Co 2p1/2) and 781.5 eV (Co 2p3/2), which are consistent with those reported for Co3O4.42 The presence of two shakeup satellite peaks at 787.6 and 804.1 eV further confirms the formation of the Co3O4 crystal phase. The high-resolution XPS spectra of Co 2p3/2 reveal the presence of Co2+ (782 eV) and Co3+ (779 eV) in the composites. The carbon bonds can be deconvoluted as follows: C–C (284.7 eV), C–O (286.1 eV), and C=O (288.1 eV) (Figure S1c), and the oxygen bonds can be deconvoluted as follows: Co–O (530.2 eV), C=O (531.8 eV), and C–OH (533.8 eV) (Figure S1d).

Figure 3 Hydrolysis process of paraoxon by Co3O4/rGO nanocomposites.

3.2 Catalytic Evaluation. Co3O4/rGO nanocomposites have been shown to act as peroxidase and catalase mimetic enzymes.33,38 However, the use of Co3O4/rGO nanocomposites to mimic hydrolytic enzymes has not been investigated in detail. In this work, the PTE-like activity of Co3O4/rGO nanocomposites was studied and measured by the formation of p-nitrophenol using UV-vis spectroscopy (Figure 3). As shown in Figure 4a, the 10


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maximum absorption peak was observed at 401 nm when the paraoxon was treated with Co3O4/rGO nanocomposites, indicating that the nanocomposites exhibit good catalytic activity. The hydrolysis of paraoxon was further studied using

31

P NMR spectroscopy in a

time-dependent manner. As shown in Figure S2, the signal at –7.8 ppm for paraoxon decreased with time, and a new peak at –0.3 ppm appeared because of the formation of diethyl phosphate. In this process, no further hydrolysis reaction was observed, indicating that the Co3O4/rGO nanocomposites exhibited high efficiency for PTE model systems. In addition, the initial rate of the reaction did not obviously change when a high concentration of diethyl phosphate was added to the reaction system (Figure 4b), indicating that the Co3O4/rGO nanocomposites could efficiently hydrolyze paraoxon and that the reaction rate was unaffected by the product, which may also be responsible for its stability against further hydrolysis.

Figure 4 (a) Absorption curves of different amounts of Co3O4/rGO nanocomposites in NaHCO3–Na2CO3 buffer containing 1 mg of paraoxon after reaction for 20 min. (b) Effect of externally added diethyl 11



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phosphate on the initial rate. (c) Effect of different nanomaterials as catalysts on the conversion of paraoxon (the amounts of catalysts are all 6 mg). (d) Conversion of paraoxon by Co3O4/rGO nanocomposites with different reactant ratios of GO:Co3O4.

To explore the role of each component in the hydrolysis of paraoxon, the catalytic activities of the pure individual components (Co3O4 NPs and rGO) were evaluated at 60 °C and compared with those of the Co3O4/rGO nanocomposites. As demonstrated in Figure 4c, the pure Co3O4 NPs were inefficient in catalyzing the hydrolysis of paraoxon. And the hydrolysis ratio of paraoxon by pure rGO was 24% within 20 min. However, the paraoxon was hydrolysed completely by Co3O4/rGO nanocomposites under the same conditions, indicating an approximately five-fold enhancement in catalytic activity compared with that of rGO. These results demonstrated that the Co3O4/rGO nanocomposite possessed excellent PTE-like catalytic activity ， which was derived from rGO and the Co3O4 played an indispensable role in improving the catalytic activity. The excellent adsorption capacity of rGO prefers to binding of paraoxon and water molecules, allowing an efficient nucleophilic attack of water (or hydroxide) at the phosphorus center (Figure S3),43,44 and the Co3O4 on the rGO facilitates the electron transfer during the catalytic reaction. The synergistic of Co3O4 and rGO is conducive to accelerating the electron transfer and promoting the catalytic reaction rate, which is consistent with previous reports.31-33 The relationship between the catalytic activity and the ratio of Co3O4 and rGO in the nanocomposites was further investigated, as shown in Figure 4d. The results demonstrated that the catalytic activity was significantly affected by the reactant molar ratio of Co3O4 and rGO, and a ratio of 1:2.5 has a higher PTE-like activity under the same conditions. It has been reported that the hydrolysis of paraoxon was carried out in the presence of a general base,28,45 and histidine has been used as a general base to enhance the rate of reaction.46,47 Nevertheless, the poor solubility of histidine would decrease the catalytic 12


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activity in aqueous solution. Therefore, N-methylmorpholine (NMM), ammonia, and Na2CO3–NaHCO3 buffer were studied as reaction media in the hydrolysis reaction. As observed in Figure 5a, the hydrolytic activity was significantly enhanced when the Na2CO3–NaHCO3 buffer was employed as a general base. In addition, the pH value of the buffer affected the hydrolysis activity. A relatively higher pH promoted the hydrolysis activity, and the buffer with pH 10.5 was selected as the reaction medium for the following experiments (Figure 5b).

Figure 5 PTE-like activity of Co3O4/rGO nanocomposites for various: (a) reaction media, (b) pH values, (c) temperatures, and (d) amounts of Co3O4/rGO nanocomposites.

To further study the catalytic performance of the Co3O4/rGO nanocomposites, the reaction was tested at different temperatures, and the results are presented in Figure 5c. The reaction in the absence of Co3O4/rGO nanocomposites was extremely slow at 20 °C, and the catalytic performance remained slow even at higher temperature (60 °C) (Figure S4). 13



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However, the catalytic activity in the presence of Co3O4/rGO nanocomposites was remarkably enhanced and temperature dependent. The conversion of paraoxon by Co3O4/rGO reached 73% in 10 min at 60 °C, which was more than 70-fold higher than that for the reaction without Co3O4/rGO. In the presence of the Co3O4/rGO nanocomposites, the t1/2 values (time required to hydrolyze 50% of the paraoxon) for the reaction were 25, 12, and 5 min at 40 °C, 50 °C, and 60 °C, respectively. Therefore, 60 °C was considered the optimum temperature for the degradation reaction. The hydrolysis of paraoxon was further studied for different amounts of the Co3O4/rGO nanocomposites (Figure 5d). The hydrolysis rate of paraoxon gradually increased with increasing catalyst amount (2, 4, 6, 8 mg). It has been reported that the interaction between the catalysts may play a key role in enhancing the reaction rate.48 The correlation curve between the catalyst amount and initial rate is presented in Figure S5, and the catalyst-dependent sigmoidal behavior is attributed to the formation of an enzyme dimer or oligomers.28 Steady-state kinetics assays were performed by changing the paraoxon concentration in the catalytic system and keeping the other reaction conditions constant (Na2CO3–NaHCO3 buffer, pH 10.5, 60 °C, 20 min, and 6 mg of Co3O4/rGO nanocomposites). A typical Michaelis–Menten plot was obtained when the concentration of the paraoxon solution was increased from 17 to 280 µM (Figure S6). Furthermore, the straight line in the Lineweaver–Burk plot suggests that the Co3O4/rGO nanocomposites follow enzyme-like kinetics. The Michaelis–Menten constant (Km = 83.2 µM), maximum velocity (Vmax = 0.02 mM min-1), and turnover number (kcat = 1.54×10-2 s-1) were calculated from these plots and the concentration of catalysts. These values are comparable to those of some PTE mimics (Table S2),18,25 showing a good catalytic activity of the Co3O4/rGO nanocomposites. The reusability and robustness of a mimetic enzyme are particularly important for 14


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practical application. The batch hydrolysis of paraoxon was used as a model reaction to evaluate the reusability of the Co3O4/rGO nanocomposites. The Co3O4/rGO nanocomposites were thoroughly washed with deionized water and ethanol after each reaction, and then, their catalytic efficiency was measured. As observed in Figure 6, the Co3O4/rGO nanocomposites retained more than 75% of their original enzymatic activities after 10 cycles, demonstrating the excellent reusability of Co3O4/rGO nanocomposites compared with other PTE mimetic enzymes. This phenomenon may be attributed to the inability of diethyl phosphate to interact with the active sites; therefore, the catalytic sites were not blocked by the reaction product, leading to the reversible activity. In addition, the SEM, TEM, XRD, and XPS analyses of Co3O4/rGO nanocomposites after reused were performed for characterizing their morphology and structure, and the results demonstrated that the Co3O4/rGO nanocomposites were almost unchanged after the reaction (Figure S7–S9).

Figure 6 Reusability of Co3O4/rGO nanocomposites.

To assess the robustness of the Co3O4/rGO nanocomposites, they were first incubated at a range of temperatures (20–100 °C) and pH values (2.0–10.0) for 2 h, and then, their PTE-like activities were measured. The results showed that the activities of the Co3O4/rGO 15



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nanocomposites remained stable over a wide range of temperatures and pH values (from 20 °C to 100 °C and from 2.0 to 10.0, respectively), demonstrating that the Co3O4/rGO nanocomposites preserved excellent temperature and pH resistance (Figure S10 and S11). Furthermore, the Co3O4/rGO nanocomposites retained more than 90% of their catalytic activities after a month of storage in air, water, and ethanol (Figure S12), showing their high stability during long-term storage. The robustness of the Co3O4/rGO nanocomposites makes them suitable for a broad range of applications for environmental and biological analyses. In addition, the relative standard deviation (RSD) of the relative catalytic activity was below 4% for five batches of the Co3O4/rGO nanocomposites (Figure S13), demonstrating their excellent reproducibility. 3.3 Detection of paraoxon and assessment of practicability. Based on the PTE-like activity of the Co3O4/rGO nanocomposites, the materials can be potentially applied in the degradation and detection of paraoxon. A simple colorimetric sensor for paraoxon detection was developed in the current study by measuring the generated p-nitrophenol, which was the hydrolysis product of paraoxon and determined at 401 nm. The feasibility of this strategy for paraoxon detection was investigated under optimized conditions. The regression equation used in this case was ∆A = 0.0093 C(µM) + 0.038. A good linear relationship (R2 = 0.9996) and low detection limit (0.8 µM, signal/noise = 3) were obtained for paraoxon (Figure 7). The linear range was 8–140 µM. The proposed sensor was used to determine the paraoxon levels in cabbage, river water, and tap water under the optimal experimental conditions, and the results are listed in Table 1. The RSD values were within 3%, demonstrating the high precision and good reproducibility of the proposed method for practical detection. The sample recoveries ranged from 98.2% to 103.8%, indicating that the proposed colorimetric sensor for paraoxon quantification is reliable in practical application.

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Table 1. Detection of paraoxon in real samples Real Samples

Detected

Added

Found

RSD

Recovery

(µM)

(µM)

(µM)

(%, n = 3)

(%)

50.0

49.1

2.8

98.2

100.0

103.1

2.9

103.1

50.0

51.9

2.6

103.8

100.0

101.7

2.5

101.7

50.0

49.4

2.5

98.8

100.0

100.1

2.6

100.1

Cabbages

–a

River

–a

Tap water

–a

a

Not detectable

Figure 7 Dose–response curve for the detection of paraoxon by Co3O4/rGO nanocomposites under the optimum conditions. Inset: linear calibration plot for the detection of paraoxon. The error bars represent the standard deviation values of three measurements.

4. CONCLUSION In summary, Co3O4/rGO nanocomposites were prepared using a facile hydrothermal synthesis method, and their PTE-like activity was investigated for the first time. The synergic combination of Co3O4 and rGO greatly improved the binding capacity with paraoxon and water molecules, and facilitated the electron transfer in the hydrolysis process. The 17



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Co3O4/rGO nanocomposites exhibited not only excellent activity for the hydrolysis of paraoxon but also good reusability, which was attributed to the inability of diethyl phosphate to interact with the active sites. In addition, the Co3O4/rGO nanocomposites present several advantages, such as low cost, easy preparation, high temperature resistance, remarkable long-term stability and reproducibility. These advantages of Co3O4/rGO nanocomposites make them highly suitable for catalytic applications, and they are thus expected to open new opportunities for the efficient degradation and sensitive detection of organophosphorus compounds in real samples.

ASSOCIATED CONTENT Supporting Information The synthetic method of Co3O4/rGO nanocomposites with different reactant ratios of Co3O4 and rGO; The investigation of the phosphotriesterase activity in the presence of diethyl phosphate and the reusability of Co3O4/rGO nanocomposites; Sample preparation for the detection of paraoxon; The XPS spectra of Co3O4/rGO nanocomposites; The 31P NMR spectra of the time-dependent hydrolysis of paraoxon by Co3O4/rGO nanocomposites; Possible mechanism of Co3O4/rGO nanocomposites on the hydrolysis of paraoxon; The phosphotriesterase activity in the absence and presence of Co3O4/rGO nanocomposites at different temperature; The effect of the amount of Co3O4/rGO nanocomposites on the initial rate; Steady-state kinetic assay and the double reciprocal plots of Co3O4/rGO nanocomposites; SEM, TEM, XRD, and XPS characterizations of Co3O4/rGO nanocomposites after reused; The relative activity of Co3O4/rGO nanocomposites treated with different temperatures and different pH values; The relative activity of Co3O4/rGO nanocomposites after long-term storage in air, water and ethanol; The repeatability of Co3O4/rGO nanocomposites in different 18


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batches; The elemental analysis of Co3O4/rGO nanocomposites with different reactant ratios of Co3O4 and rGO; Comparison of the Michaelis–Menten constant (Km) and maximum reaction rate (Vmax) with other catalysts.

AUTHOR INFORMATION Corresponding Author * E-mail: [email protected]; [email protected]. Tel.: +86 10 64441521. Notes The authors declare no competing financial interest.

ACKNOWLEDGMENT This work was supported by the National Nature Science Foundation of China (Grant No. 21075008) and the Beijing Natural Science Foundation (Grant No. 2132048).

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Reduced Graphene Oxide ... - ACS Publications

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