Ratiometric Electrochemical Sensors Associated with Self-Cleaning

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Functional Nanostructured Materials (including low-D carbon)

Ratiometric electrochemical sensor associated with self-cleaning electrode for simultaneous detection of adrenaline, serotonin and tryptopha Junjie Zhang, Dongyang Wang, and Yingchun Li ACS Appl. Mater. Interfaces, Just Accepted Manuscript • DOI: 10.1021/acsami.8b22572 • Publication Date (Web): 15 Mar 2019 Downloaded from http://pubs.acs.org on March 15, 2019

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Ratiometric electrochemical sensor associated with self-cleaning electrode for simultaneous detection of adrenaline, serotonin and tryptophan Junjie Zhanga, 1, Dongyang Wangc, 1, Yingchun Lia,b, * a

Key Laboratory of Xinjiang Phytomedicine Resources for Ministry of Education, School of

Pharmacy, Shihezi University, Shihezi, 832000, China b

College of Science, Harbin Institute of Technology, Shenzhen, 518055, China

c

Key Laboratory for Green Processing of Chemical Engineering of Xinjiang Bingtuan, School of

Chemistry and Chemical Engineering, Shihezi University, Shihezi 832003, China Keywords: zeolite imidazole skeleton; polydimethylsiloxane; methylene blue; electrocatalytic oxidation; rat analysis; superhydrophobic

Abstract: Electrochemical sensors have long suffered from issues such as non-specific adsorption, poor anti-interference ability, as well as internal and external disturbances. To address these challenges, we developed a facile electrochemical method, which integrated ratiometric strategy with self-cleaning electrode. In the novel sensing system, the self-cleaning electrode was realized via forming a hydrophobic layer on carbonized ZIF-67@ZIF-8 (cZIF) by PDMS precursor vaporization. As for ratiometry, it is worth to mention that the measurements were conducted by adding interior reference (methylene blue, MB) directly into electrolyte solution, which is more facile and flexible to


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operate compared with conventional ones. Sensing performance of the self-cleaning electrode as well as the newly established ratiometric strategy was explored fully and it turned out that PDMS@cZIF nanocomposites provided decent electrocatalytic ability, superhydrophobic property and stability. Furthermore, ratiometric strategy significantly elevated the robustness and reproducibility of electrochemical sensing. Simultaneous detection of Adr, 5-HT and Trp was performed under the optimum experimental conditions with wide linear ranges and low detection limits. Finally, the original R-ECS was successfully applied for monitoring the three target molecules in biological samples.

1. Introduction Electrochemical method is very convenient and applicable in the currently developed methods thanks to the advantages of fast detection, easy handling, low cost and miniaturization1-2. During the past a few decades, the introduction of nanomaterials and the specific interactions on the electrode surface led to a significant promotion in the sensitivity and selectivity to analytes.

Unfortunately,

due to the poor anti-interference ability of electrochemical sensor, intrinsic or extrinsic factors such as sensor concentration, instrumental efficiency and environmental condition, and electrode pollution and passivation by the substances in testing samples, accuracy and stability are often seriously threatened. Therefore, measurement in complex biological samples based on electrochemical sensor is still challenging. Since Barthlott and Neihuist’s group3 were inspired by the self-cleaning lotus leaves, the hydrophobic materials have been extensively studied and reported. Recently, Li’s group4 applied hydrophobic organic polymer to fabricate electrochemical sensors and the research shows that the developed sensors can effectively avoid molecular adsorption and electrode passivation.



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Polydimethylsiloxane (PDMS) as a widely used hydrophobic organic polymer has a good application potential in the sphere of electrochemical sensing owing to its favorable cost-effectiveness, biocompatibility and stability. In addition, various modifiers have been adopted for electrode decoration to enhance sensing performance, among which MOF derived nanocomposites have received great attention5. Zeolite imidazole skeleton (ZIF), a MOF subfamily based on transition metal and imidazole linkers, features well-defined morphology, excellent chemical stability and permeability to guest molecules6-7. ZIF-derived materials have raised rapidly growing interest for catalysis, energy storage and sensing8-9, with ZIF-67 and ZIF-8 act as superior precursors which own large surface area, readily available and low cost10-13. What’s more, compared with the reported self-cleaning electrode, the constructed self-cleaning electrode by combining a hydrophobic material PDMS with carbonized ZIF-67@ZIF-8 (cZIF) shows a superior performance (Table S1). Therefore, PDMS@cZIF can be used as an ideal electrode modification material. Reproducibility of traditional electrochemical analysis strategies is often limited as they are liable to

suffer

from

interference

of

substrates

and

systems.

These

disturbances

include

micro-environmental contamination, complex operations and slow electron transfer kinetics, which cause the differences between lab and lab, person and person, electrode and electrodes, etc. In order to promote sensing reproducibility, ratiometric electrochemical method has achieved remarkable development in recent years, which surmounts the inherent systematic errors by obtaining a proportional response between the analytes and the internal reference, thereby significantly elevating accuracy and reproducibility of electrochemical sensing14. Most, ratiometric electrochemical sensors (R-ECSs) are prepared by immobilizing the internal reference molecules on electrode and they are often chemically combined with the target molecule recognizer to provide a built-in correction15-16.


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Although R-ECSs allow for high elevation of sensing performance including repeatability, robustness, reproducibility, etc., the drawbacks they have such as long detection time and arduous electrode modification, hinder their practical application. Wang’s group constructed a facile ratiometric method by adding reference molecules (methylene blue, MB) directly into electrolyte solution, which avoids the cumbersome steps of electrode modification and simplifies the experimental operation17-18. Inspired by this work, and aiming to develop an electrochemical sensor with satisfying repeatibility and stability, we first integrated super-hydrophobic polymer with ZIF derived material as sensing platform, and then introduced the facile ratiometric tactic into determination. The developed sensor shows a promising application prospect, and this is the very first report on combining self-cleaning electrode with R-ECS for electrochemical sensor application. We built a superhydrophobic working electrode which eliminates the non-specific adsorption of molecules, and further developed it into R-ECS, thereby acquiring satisfactory accuracy and reproducibility. Specifically, a common used glassy carbon electrode (GCE) was modified by using a hydrophobic material PDMS combined with cZIF to obtained self-cleaning electrode. MB serving as an internal reference was added directly in the testing solution to correct detection errors for Adr, 5-HT and Trp. The proposed sensor exhibited accurate and sensitive determination with broad-range linear calibration and low limit of detection (LOD). What’s more, it was favorably used for quantification of the three compounds in brain tissue and blood specimens.

2. Experimental Section 2.1 Reagents and apparatus Adrenaline (Adr), tryptophan (Trp), methylene blue (MB) and 2-methyl imidazole were bought from Shanghai Adamas Reagent Co. Ltd. (Shanghai, China). Serotonin (5-HT) was purchased from



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Beijing Innochem Co. Ltd. (Beijing, China). Polydimethylsiloxane (PDMS) was supplied by Shanghai TCI Chemical Industry Co. Ltd. (Shanghai, China). Co(NO3)2·6H2O, Zn(NO3)2·6H2O were provided by Shanghai Shanpu Chemical Co. Ltd. (Shanghai, China). Reagents and materials, such as K3[Fe(CN)6], K4[Fe(CN)6], guanine, adenine, glucose (Glu), NaCl, ascorbic acid (AA), NH4Cl, H2SO4, KNO3, KCl, CaCl2, phosphate buffer (PB, K2HPO4 and KH2PO4) were of the analytical level. All solutions were formulated using double distilled water. Surface morphology of the employed materials was characterized using a Nova Nano FESEM 450 scanning electron microscope (SEM). Energy dispersive spectrometry (EDS) data were recorded with a Nova Nano FESEM 450 at 15 kV equipped with Energy Dispersive X-ray Spectroscopy (Bruker XFlash-SDD-5010, Germany). Phase structures were analyzed by Fourier transform infrared spectroscopy (FTIR; Nicolet iS10, Thermofisher, America). A centrifuge (Anke TGL-16G, Shanghai, China) was used in pretreatment of biological samples. Electrochemical measurements were performed by a computer-controlled Electrochemical Workstation (CHI660, CHI Instruments Co., Shanghai, China). A traditional three-electrode system consist of a bare or modified glassy carbon electrode (GCE, 4 mm in diameter) as working electrode, Pt-wire auxiliary electrode as counter elelctrode and a saturated calomel electrode (SCE) as reference electrode. All potentials given in this paper were referred to SCE19. 2.2 Synthesis of cZIF ZIF-67@ZIF-8 was synthesized in accordance with a previously recorded method20-23. Briefly, Co(NO3)2•6H2O (4.368 g), Zn(NO3)2•6H2O (4.464 g) and 2-methyl imidazole (4.928 g) were dissolved in 60 mL methanol, 60 mL methanol and 120 mL methanol, respectively under ultrasound for 5 min at 25 °C. Afterwards, the Co(NO3)2•6H2O methanol solution was slowly added into ligand


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solution (2-methyl imidazole) within 5 min under ultrasonic and stirring condition. Next, the Zn(NO3)2•6H2O methanol solution was mixed with the solution as mentioned above under the same condition and the final solution was further sonicated for 1 h. The resulting suspension was subjected to centrifugation at 8000 rpm for 10 min, and the sediments were rinsed with methanol for three times followed by drying at 80 °C for 2 h to get ZIF-67@ZIF-8 crystals. cZIF were then obtained after ZIF-67@ZIF-8 crystals were heated for 6 h at 900 °C under nitrogen flow at temperature ranging from 100 to 900 °C for 2.5 h with a heating rate of 6 °C min−1.

2.3 Synthesis of PDMS@cZIF PDMS@cZIF nanoparticles were prepared by following a reported method. Concisely, PDMS precursor was blended with cZIF in a weight ratio of 1:1 and added to a 25 mL reacting kettle. The PDMS layer was formed on cZIF by sealing the kettle and keeping it at 300 ° C for 12 hours. After dropped to 25 °C, the nanoparticles were dispersed in DMF with ultrasonication for 2 h, followed by centrifugation (2000 rpm, 10 min) and filtration. Ultimately, the nanoparticles were ready for use.

2.4 Preparation of self-cleaning PDMS@cZIF/GCE GCE was buffed on a chamois with 0.05 μm Al2O3 powder, and then washed with double-distilled water and dried at room temperature. 1 mg PDMS@cZIF was suspended in 1 mL DMF by ultrasonication for 2 h, and then 10 μL of the suspension was pipetted onto GCE surface and the solvent was evaporated under infrared lamp. A uniform and superhydrophobic surface on GCE was formed by repeating the “drop and dry” steps four times24. The schematic preparation process of PDMS@cZIF/GCE is illustrated in Fig. 1.



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Fig.1 The Schematic diagram for preparing of PDMS@cZIF/GCE.

2.5 Electrochemical measurement Cyclic voltammetry (CV) was used to estimate electrochemical properties of different electrodes in probe solution (containing 5 mM [Fe(CN)6]3−/4− and 0.1M KNO3) from −0.2 V to 0.6 V (vs. SCE) at a scan rate of 50 mV s−1. Electrochemical impedance spectroscopy (EIS) was performed in probe solution with 0.1 M KCl. Electrochemical performance of PDMS@cZIF/GCE towards MB, Adr, 5-HT and Trp was recorded through differential pulse voltammetry (DPV) in PB with the potential scanning range from −0.6 to 0.8 V (vs. SCE). Before each use, working electrodes was processed by CV scanning to return to their initial potential.

2.6 Measurement of Adr, 5-HT and Trp in real samples For purpose of studying the practicability of applying the decorative sensor in complex biological samples, the prepared PDMS@cZIF/GCE was used to detect 5-HT in brain tissue of Sprague-Dawley (SD) rat (Experimental Animal Research Center of Xinjiang, Shihezi, China) and


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Adr, 5-HT and Trp in rat serum. The brain sample treatment follows behavioral experiment requirement. First, SD rat was beheaded to collect brain and blood samples and stored at 4 °C when not in use. Brain sample was homogenized with a homogenizer (RH Instruments FSH-2) while blood samples were centrifugated and the upper serum was collected. Then both specimens were mixed with four-fold volume of methanol to precipitate protein, and then the supernatant was diluted with a volumetric ratio of 1:1. The final supernatant was used for analyte detection25.

3. Discussion 3.1 Preparation and characterization of self-cleaning electrode Surface morphology and constituent characterization of the prepared nanocomposites were investigated by SEM and EDS. SEM images (Fig. 2A) demonstrated that ZIF-67@ZIF-8 exhibits a core–shell structure with a distinct rhombohedral shape and uniform size (350 nm). The topological characteristics of the typical core–shell architecture were verified by the corresponding elementary mapping (Fig. 2B-2D). The distribution of Co and Zn indicated that the Co-containing ZIF-67 is located inside the rhombohedral dodecahedron and is encapsulated by the Zn-containing ZIF-8 shell26. Besides, the successful formation of ZIF-67@ZIF-8 is supported by EDS data (Fig. 2G). After high-temperature carbonization, the parent ZIF-67@ZIF-8 converted to cZIF with a gauzy carbon layer (Fig. 2E), and the EDS data also shows decrease of Zn level (Fig. 2H). By heating the mixture of cZIF with PDMS at an equal proportion, cZIF gets agminated and its thickness is increased (Fig. 2F). The presence of Si element was observed by EDS spectra (Fig. 2I).



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Fig. 2 SEM pictures of ZIF-67@ZIF-8 (A, B), cZIF (E), PDMS@cZIF (F), elemental mapping of ZIF-67@ZIF-8 (C, D), EDS spectrum of ZIF-67@ZIF-8 (G), cZIF (H) and PDMS@cZIF (I). Insets are either magnified images of the materials (A, E, F) or atom content ratio (G-I). XPS measurements were used to further insight into element content of cZIF and PDMS@cZIF (Fig. 3A). After the PDMS layer formed, the C 1s, O 1s, Si 2p level were increased distinctly when compared with cZIF. As displayed in Fig. 3B, the O 1s peak high resolution spectroscopy of cZIF has two peaks (529.6 eV and 530.3 eV), which are ascribed to O=C-O and C-O-C, respectively27. As illustrated in Fig. 3C, except that the peaks centered at 529.6 eV and 530.3 eV were essentially the same as cZIF, PDMS@cZIF showed a pronounced C-O-Si peak (at 531.9 eV), which can indicate that there is a chemical bonding between PDMS and cZIF27-29; and the one at 533.1 eV is from the Si–O–Si in PDMS29.

Fig. 3 XPS spectra of cZIF and PDMS@ cZIF (A) and the O 1s peak high resolution spectroscopy of


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cZIF (B) and PDMS@cZIF (C). X-ray powder diffraction (XRD) was used to study the crystal structure of ZIF-67@ZIF-8, cZIF and PDMS@cZIF (Fig. S1). As previously reported, for ZIF-8, seven main characteristic peaks were observed at 2θ of 10.4, 12.7, 14.7, 16.4, 18.0, 24.5 and 26.7; for ZIF-67, characteristic peaks were present at 2θ of 19.4, 22.0, 29.6, 30.6, 31.5, 32.3, 34.9, 36.6, which confirm the successful synthesis of ZIF-67@ZIF-826, 30 (Fig. S1A). Fig. S1B shows that the initial ZIF-67@ZIF-8 diffraction peaks completely disappeared and ZIF-67@ZIF-8 was converted to cZIF. After co-heating treatment with PDMS, the position of the diffraction peak of the other diffraction angle did not change, but the intensity thereof was significantly weakened31, demonstrating that the PDMS modification was successful. FT-IR spectra further verify the structure of ZIF-67@ZIF-8, cZIF and PDMS@cZIF. As shown in Fig. 4A, the bands at 2905 cm−1 and 620 cm−1 are ascribed to the stretching vibration of C-H and C=N32, respectively, both of which indicate that the primary ligand in these ZIFs is 2-methylimidazole. The band at about 1258 cm−1 corresponds to the symmetric band of Si-CH3, and the band at 1078 cm−1 pertains to the stretching vibration absorption of the Si-O and the band at 814 cm−1 is attributed to CH3 shaking vibration of Si-CH324 (Fig. 4B). These bands confirm the presence of PDMS in the PDMS@cZIF composite.

Contact angle tests were carried out to evaluate the wetting abilty of ZIF-67@ZIF-8, cZIF and PDMS@cZIF nanocomposites (Fig. 4C-4E). The contact angles of the three nanocompsites are 66.02°, 24.32° and 153.76°, respectively, which could be attributed to the hydrophobic nature of ZIF-67@ZIF-8 and the hydrophilic carbonized surface of cZIF. As for PDMS@cZIF, the water



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droplet exhibits an obviously large contact angle, which indicates that PDMS@cZIF contribute to a stable super-hydrophobic surface33-34. Detailed description toward the superhydrophobic surface model of PDMS@cZIF was displayed in Supporting Information. The results of the above experiments all confirm the successful preparation of PDMS@cZIF.

Fig. 4 FT-IR spectra of ZIF-67@ZIF-8 (A), cZIF/GCE and PDMS@cZIF/GCE (B); contact angle photographs of ZIF-67@ZIF-8 (C), cZIF (D) and PDMS@cZIF (E).

3.2 Electrochemical characteristics CV and EIS measurements were implemented to trail the conductivity and resistivity of different electrodes during preparation, the results of which are presented in the Supporting Information file. As shown in Fig. S2A, after ZIF-67@ZIF-8 modification, the characteristic redox peak currents of probe got decreased, which was ascribed to the poor conductivity of ZIF-67@ ZIF-8. In contrast, decoration of cZIF increased the peak current of GCE, which was very likely to do with the enlarged carbonized surface area with good conductivity which elevates electron transfer rate. Introduction of PDMS slightly reduced current response, which could be ascribed to the poor conductive PDMS. The EIS results in Fig. S2B are in consistent with CV, and the calculated electron transfer resistance


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(Rct) value of cZIF/GCE, PDMS@cZIF/GCE, bare GCE and ZIF-67@ZIF-8/GCE are 2.4 Ω (curve c), 24.7 Ω (curve d), 52.2 Ω (curve a) and 143.5 Ω (curve b), respectively.

3.3 Fabrication of self-cleaning electrodes and R-ECS This work have proposed a novel sensing platform that combines self-cleaning electrodes with ratiometric electrochemical detection. In order to verify feasibility of this platform, we conducted a series of experiments. First, electrochemical catalytic oxidation of three analytes was studied. Fig. S2C illustrates the DPV diagrams of different electrodes in 0.1 M PB containing 4 μM MB, 30 μM Adr,

5-HT

and

Trp.

The

electrodes

include

bare

GCE,

ZIF-67@ZIF-8,

cZIF/GCE,

PDMS@cZIF/GCE. It can be observed that each analyte provides a proper redox potential which doesn’t interfere the electrochemical reactions of others, and the peak currents obtained at PDMS@cZIF/GCE and cZIF/GCE are higher than the other two electrodes. These results indicate that the sensors are able to take effective measurement of the three analytes (Adr, 5-HT and Trp), and MB is an ideal internal reference with well separated oxidation peak at −0.28 V. The electro-oxidation mechanism of Adr, 5-HT and Trp is displayed in Fig. S2D. Then the robustness of the developed R-ECSs was further demonstrated by conducting the following experiment. After each assay, PDMS@cZIF/GCE was taken out and thoroughly rinsed with water. The electrodes were then re-immersed into 0.1 M PB containing 30 μM Adr, 5-HT and Trp with or without MB for the next testing of the targets. As shown in Fig. 5A and 5B, repeated measurements lead to successive decrease of electrical signals (IAnalyte and IMB), which could be attributed to irreversible oxidation reactions occurring at electrode interface. Fig. 5C is the plots of IAnalyte versus measurement times, the data of which are derived from Fig. 5A. Apparent signal decrease (IAnalyte) is observed, generating large deviation among repeated measurements. By contrast,



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Fig. 5D displays the quantification results with ratio signals originated from Fig. 5B. It is obvious that introduction of ratiometric assay contributes a lot to the repeatability among continuous tests, with relative standard deviations (RSDs) reduced from ~50% to about 2%. These results suggest the remarkably robustness and dependability of R-ECS compared to traditional ones. Stability and reproducibility of the fabricated sensor were also estimated. Herein, the self-cleaning electrode was placed in the air for up to 45 days to evaluate time stability, while reproducibility was assessed by continuously polishing and remanufacturing one electrode for five times. The experimental results display that the current responses of modified electrode are stable with an average RSD of 6.67% (Fig. 5E-5F). Based on the above results, one can conclude that PDMS@cZIF nanocomposite can be employed as a self-cleaning material for building R-ECS by using the oxidation peak of MB as the internal reference.

Fig. 5 DPV voltammograms and plots of electrochemical response versus measurement times by using PDMS@cZIF/GCE in 0.1 M PB containing 30 μM Adr, 5-HT and Trp with (A, C) or without MB (B, D). Repeated usage of PDMS@cZIF/GCE at different time points (E) and after several


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treatments of the electrode (F).

3.4 Influence of pH on electrochemical oxidation of Adr, 5-HT and Trp The effect of pH on performance of PDMS@cZIF/GCE towards three analytes was investigated in pH range of 6.0 ~ 8.0. As shown in Fig. S3, the oxidation peak currents of MB, Adr, 5-HT and Trp first increase and then decrease with increasing pH, and the maximal response of the three substances was obtained at pH 7.0. What’s more, the oxidation peaks of MB, Adr, 5-HT and Trp shift to more negative potentials with increased pH, suggesting that occurance of their oxidation gets easier. The linear relationships between peak potentials and pH values are: Ep (MB)= 0.233–0.071 pH (R2 = 0.994) for MB , Ep (Adr)= 0.489–0.052 pH (R2 = 0.973) for Adr, Ep (5-HT)= 0.724–0.067 pH (R2= 0.990) for 5-HT, and Ep (Trp)= 1.043–0.055 pH (R 2= 0.999) for Trp. The calculated slopes for these four substances are close to the theoretical value of −59.1 mV/pH according to the Nernst equation, implying that the ratio between electrons and protons involved in charge transfer is 225, 35. The decrease of pH value in the electrolyte environment led to an increase in the concentration of H+, which suppressed the oxidation reaction of Adr, 5-HT and Trp. In order to guarantee the utility of sensor in actual physiological environment, we chose pH 7.0 in the subsequent experiments.

3.5 Determination of Adr, 5-HT and Trp Both individual and simultaneous determination of Adr, 5-HT and Trp was carried out by using the developed R-ECS based on PDMS@cZIF/GCE. In all the measurements, MB, added as the internal reference, was at the concentration of 4 μM. For individual tests, other analytes remained constant at 30 μM, only changed the concentration of the target analyte. As for simultaneous detection, concurrent change in the concentration of the three analytes were conducted in the range of 1~60 μM.



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The results of individual measurements were displayed in Supporting Information (Fig. S4). A comparison was established to contrast the main features of some chemically-modified electrochemical sensors reported for Adr, 5-HT and Trp detection (Table S2). It can be seen that PDMS@cZIF/GCE exhibits relatively low LOD and wider linear range, which are comparative or better than its counterparts. The subsequent simultaneous detection of the three analytes using PDMS@cZIF/GCE further confirms the decent electrochemical performance of PDMS@cZIF/GCE (Fig. 6). The corresponding linear ranges for Adr, 5-HT and Trp are all from 1.0 to 60 μM with LOD of 0.13 μM for Adr, 0.03 μM for 5-HT and 0.5 μM for Trp (S/N = 3).

Fig. 6 DPV voltammograms at PDMS@cZIF/GCE in 0.1 M PB (containing 4 μM MB, pH 7.0) for simultaneous determination of Adr, 5-HT and Trp (A); calibration plots of Ianalyte/IMB versus concentration of the three analytes from three parallel measurements (B).

3.6 Interference study Interference study of PDMS@cZIF/GCE was performed by determining the mixture of analytes (30 μM Adr, 5-HT and Trp) and several potential interfering ions and molecules (Na+, K+, NH4+, Mg2+, Ca2+, Cl−, SO42−, NO3−, Guanine, Adenine, AA, Glu and L-cysteine at the concentration of 5 mM). Experimental results shown in Table S3 illustrate that the impact caused by the interferents on determination of Adr, 5-HT and Trp using PDMS@cZIF/GCE are negligible with the relative signal change less than 3%. The good anti-interference capability is very beneficial when used in complicated ACS Paragon Plus Environment

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matrix such as biological fluid samples.

3.7 Repeatability, reproducibility and stability of PDMS@cZIF/GCE Measurements of a mixture of 4 μM MB, 30 μM Adr, 5-HT and Trp were carried out for 20 times using PDMS@cZIF/GCE, and RSDs of the oxidation peak currents of the four species were less than 3.5%, confirming admirable repeatability of the prepared sensor. Reproducibility was evaluated by preparing five independent PDMS@cZIF/GCEs in the same condition for detection. The results prove that the sensor contains good reproducibility and the RSDs of their current signals from 1.7%~3.1%. The long-term stability of PDMS@cZIF/GCE was investigated by comparing the response of electrode after storage at 4 °C for 30 days. The results showed that the peak currents of MB, Adr, 5-HT and Trp were 97.6%, 94.7%, 95.3%, and 95.5% of the initial peak currents, respectively. These results indicate that reproducibility and stability of PDMS@cZIF/GCE are satisfactory.

3.8 Determiantion of Adr, 5-HT and Trp in biological sample Feasibility of the established sensing tactic in analyzing real biological samples was investigated by assaying 5-HT in rat brain tissue and Adr, 5-HT and Trp in rat blood samples via standard addition method. Before analysis, the samples were pretreated as described in Experimental Section 2.6, and the oxidation peak current ratios of analytes to MB were measured. After that, a certain amount of analytes were added into the solution and the ratios were recorded again. Recoveries ranging from 93.5% to 101.8% (Table S4) suggest good accuracy of PDMS@cZIF/GCE in real biosample analysis.

4. Conclusion Herein, we have manufactured a novel R-ECS and associated it with a self-cleaning electrode for sensitive and reliable detection of Adr, 5-HT and Trp. This new ratiometric electrochemical strategy was ACS Paragon Plus Environment


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carried out by adding the internal reference probes directly into the electrolyte solution, which is different from the traditional approach by modifying the probes on electrode surface and free from discreet electrode modification process. Self-cleaning electrode was prepared by modifying a dispersion solution of nanoparticles obtained by co-heating a cZIF material with PDMS onto a GCE. Such treatment eliminates electrode passivation and provides a refreshable sensing platform. Quantification of these three targets in biological samples exhibits decent sensitivity, reproducibility and stability, with satisfactory detection recoveries. In addition, thanks to the ratiometric electrochemical strategy and the reproducible electrode, long-term storage and successive measurement experiments demonstrated that the as-proposed sensor possesses excellent robustness and repeatability. However, It should be noted that this novel R-ECS suffers from limitation in the aspect of in vivo detection, but it will serve as a flexible and robust analytical tool in most in vitro assay environments. Nevertheless, our work offers an original R-ECS combined with renewable electrode facilitating electrochemical sensor development in terms of practicability, multifunctionality and reliability.

ASSOCIATED CONTENT Supporting Information. Comparison of the major characteristics of self-cleaning electrodes; XRD peaks of ZIF-67@ZIF-8, XRD patterns of ZIF-67@ZIF-8, cZIF and PDMS@cZIF; Detailed description of the superhydrophobic surface model of PDMS@cZIF; Cyclic voltammograms, Nyquist diagrams and differential pulse voltammograms of differently modified electrodes, and the proposed electro-oxidation mechanism of Adr, 5-HT and Trp; Differential pulse voltammograms of PDMS@cZIF/GCE at different pH values; Individual determination of Adr, 5-HT and Trp; Comparison of the major characteristics for detecting Adr, 5-HT and Trp using different electrochemical sensors; Sensing responses towards Adr, 5-HT and Trp in the presence of interfering substances; Determination of Adr, 5-HT and Trp in biological samples ACS Paragon Plus Environment

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using PDMS@cZIF/GCE.

AUTHOR INFORMATION Corresponding author *E-mail address: [email protected]. Tel/Fax: +86-755-86239466

Author Contributions The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. 1

These authors contributed equally to this work.

Notes Any additional relevant notes should be placed here.

Acknowledgements The work financially supported by National Natural Science Foundation of China (81773680) and free exploration project from the Natural Science Foundation of Shenzhen (JCYJ20170811152540640).

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Ratiometric Electrochemical Sensors Associated with Self-Cleaning

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