Transition metal doping boosting paper-based photoelectrochemical immunosensing for neurofilament light chain protein detection

Abstract

Early diagnosis of neurodegenerative diseases relies on highly sensitive detection of their specific biomarkers. However, existing photoelectrochemical (PEC) sensing technologies, despite the promising prospects, exhibit significant limitations in detection stability, photoelectric conversion efficiency, and portability. This work reported an innovative approach integrating transition metal doping modification with paper-based electrodes to construct a highly sensitive and low-cost paper-based sensing platform for detecting neurofilament light chain protein (NEFL). Fe-doped TiO₂ nanomaterials were synthesized via a homogeneous thermal treatment method. Fe substitution within the TiO₂ lattice constructed an intermediate energy level, optimizing the bandgap to 2.17 eV. This expanded the photoresponse range into the visible spectrum, enhancing the photocurrent intensity by 31-fold compared to pure TiO₂. A printed carbon paper triple-electrode system was fabricated via a template method, demonstrating excellent stability and reproducibility. Based on an immuno-sandwich model, the presence of target NEFL triggered glucose oxidase conjugated Au nanoparticle (GOx@Au NP)-labeled signal antibodies to catalyze glucose into hydrogen peroxide, further sensitizing the photocurrent response of Fe-TiO₂ to achieve wide range (0.01–200 ng mL⁻¹) detection of NEFL. This work enhances the PEC efficiency via Fe-doped TiO₂ and solves portability with paper-based electrodes, enabling sensitive NEFL detection, which supports early diagnosis of neurodegenerative diseases and provides an innovative technical solution for rapid point-of-care diagnosis of neurodegenerative diseases, while also offering theoretical and experimental references for the modification of wide band gap semiconductor materials and the development of paper-based sensing platforms.

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Introduction

Neurodegenerative diseases as major challenges in the field of global public health, their early diagnosis and intervention are of great significance for slowing down disease progression and improving patients’ quality of life.^1,2 Neurodegenerative disease biomarkers, reflecting neuronal damage and pathological processes, require highly sensitive and specific detection for early screening, disease monitoring, and treatment efficacy assessment.^3,4 Photoelectrochemical (PEC) technology, with its high sensitivity, rapid response, and simplicity, has shown great potential in biomarker detection.^5–13 However, current PEC-based methods for detecting biomarkers still face numerous challenges: on one hand, the stability of the detection system is insufficient, making it susceptible to environmental interference and resulting in poor reproducibility of detection results; on the other hand, traditional photoactive materials have high photogenerated carrier recombination rates, limiting the photoconversion efficiency and affecting the detection sensitivity; additionally, the specific recognition capability of PEC immunosensors needs improvement to reduce interference from complex matrices in biological samples with detection results.^14–16

Titanium dioxide (TiO₂) has been widely used in the field of photocatalysis due to its high chemical stability, low cost, non-toxicity, and strong catalytic activity, such as in photocatalytic hydrogen production, pollutant degradation, and sensors.^17–19 However, as a wide-bandgap semiconductor (bandgap width 3.0–3.2 eV), it only responds to ultraviolet light with wavelengths < 400 nm, resulting in a solar energy utilization rate of less than 5%.²⁰ Additionally, the photogenerated electron–hole pairs tend to recombine, leading to low photoconversion efficiency. Therefore, performance optimization through doping modification is urgently needed.²¹

Introducing transition metal ions (Fe, Mn, Co) into the oxide lattice through ion substitution or interstitial doping is an effective strategy for regulating the bandgap.^22–25 Their multivalency and unpaired d electrons can form impurity levels within the lattice, altering electronic transition pathways and energy thresholds, thereby enhancing the photon capture efficiency.^26,27 Using TiO₂ as an example, Fe³⁺ isotopic substitution of Ti⁴⁺ constructs intermediate energy levels between the valence band top and conduction band bottom, extending light absorption into the visible light region.²⁸ Compared to traditional physical modification, transition metal doping offers a simpler process, lower cost, and controllable concentration, while stabilizing the crystal structure and enhancing the optoelectronic performance, making it highly valuable in the development of optoelectronic sensors.²⁹

In addition, the portability of PEC biosensors urgently needs to be improved. Existing PEC detection systems rely on large equipment such as xenon lamps and electrochemical workstations, whose size and power consumption hinder the portability of sensors.³⁰ Complex circuit connections and signal processing modules also limit the flexibility of the system. Screen-printed electrodes based on paper substrates, with their advantages of simple manufacturing processes and low costs, have emerged as a key solution.³¹ Integrating these electrodes with a smartphone can create an intelligent detection platform: leveraging paper-based materials to reduce costs and improve efficiency, while utilizing the widespread adoption and data processing capabilities of smartphones to enable real-time data collection, analysis, and remote transmission, thereby opening new avenues for the miniaturization of sensors.

Neurofilament light chain protein (NEFL) is a specific marker of neuronal damage. Its detection is of indispensable clinical value for the early diagnosis, disease progression monitoring, and treatment efficacy assessment of neurodegenerative diseases.³² Inspired by relevant strategies, this study focused on the synergistic innovation of photoconversion and printed screen-printed paper-based electrodes to develop highly sensitive, low-cost photoelectrochemical immunosensors (Scheme 1A). By regulating Fe doping to optimize the photoconductive properties of TiO₂ nanomaterials and combining the advantages of paper-based electrodes, a novel sensing platform was constructed. Fe-doped TiO₂ (TiO₂-Fe) nanoparticles prepared using a homogeneous thermal treatment method exhibit high photoconversion efficiency and excellent stability. Band structure analysis confirmed that Fe doping introduced intermediate energy levels, altering the electron transition pathways. A three-electrode system was constructed on a paper substrate using the template method, and electrochemical performance characterization was completed (Scheme 1B). Based on the immunological sandwich model, glucose catalyzed the generation of hydrogen peroxide (H₂O₂) in the presence of the target, sensitizing the TiO₂-Fe photocurrent response, providing innovative strategies and theoretical support for the development of high-performance PEC sensors.

Scheme 1 (A) Schematic diagram of the enzymatic reaction triggered by NEFL captured in microplates; (B) fabrication process of printed screen electrodes and photocurrent conversion using TiO₂-Fe triggered by NEFL.

Experimental section

Materials and reagents

The antibodies used in this study are as follows: purified rabbit anti-human NEFL capture antibody (280029, Abcam), NEFL polyclonal antibody (12998-1-AP, Proteintech). Bovine serum albumin (BSA) and Tween 20 were purchased from Sigma-Aldrich (Shanghai, China). Iron nitrate (Fe(NO₃)₃), tetrabutyl titanate, and polyvinylpyrrolidone were ordered from Aladdin (Shanghai, China). All other materials and chemicals not mentioned were purchased from Sinopharm Chemical Reagent Co., Ltd (Shanghai, China). High-binding microplates were purchased from Sangon Institute (Shanghai, China). All experiments were performed using ultra-pure water (>18.0 MΩ, Milli-Q).

Protocol for the synthesis of TiO₂-Fe nanoparticles

The synthesis of Fe-doped lychee-shaped TiO₂ was achieved via a one-step hydrothermal method. Firstly, 25 mg of hydroxypropyl cellulose was precisely weighed and dispersed in 50 mL of an ethanol–acetonitrile mixed solvent with a volume ratio of 3 : 1. After thorough dispersion, 0.6 mL of ammonia water was added to the system and stirred at an appropriate speed (600 rpm) on a magnetic stirrer for 5 min to obtain a uniform and stable solution A. Subsequently, 70 mg of iron nitrate nonahydrate (Fe(NO₃)₃·9H₂O) was weighed and dissolved in 8 mL of the same volume ratio of ethanol–acetonitrile mixed solvent, followed by 2 mL of tetrabutyl titanate (TBOT), and stirred continuously for 5 min to successfully prepare solution B with a Fe doping concentration of 3%. Finally, under vigorous stirring conditions, solution B was rapidly injected into solution A. The system color gradually changed from clear to turbid, forming a yellow suspension. After stirring the suspension for 2 h, solid–liquid separation was performed using a high-speed centrifuge (10 000 rpm, 5 min). The solid product was washed multiple times with anhydrous ethanol to thoroughly remove impurities, and then dried in a vacuum drying oven at 60 °C for 6 h. The dried powder was placed in a muffle furnace and heated at a rate of 5 °C min⁻¹ to 500 °C (3 h), ultimately yielding lychee-shaped TiO₂-Fe microspheres.

A4-sized cellulose paper was selected as the substrate, the surface was cleaned with anhydrous ethanol, allowed to dry naturally, and then fixed to the printing platform. Next, a 400-mesh nylon screen template with a three-electrode pattern was precisely aligned and secured to the cellulose paper. Conductive carbon ink with a solid content of 30% was uniformly applied to the template surface. A stainless-steel squeegee was used at a 45° angle, maintaining a constant speed during the scraping process, to transfer the conductive carbon ink onto the substrate material surface, thereby forming the electrode pattern. The resulting sample was left to stand at room temperature for 15 min, and then cured in a 60 °C forced-air drying oven for 30 min. Finally, excess conductive layers were removed using laser etching technology, successfully producing a carbon paper three-electrode system. A 300-mesh screen template was selected and fixed onto the prepared carbon-based cellulose paper. Using silver ink with a solid content of 40%, a circular silver electrode pattern with a diameter of 3 mm was formed via the coating and scraping process. The sample was left to stand at room temperature for 20 min, then cured at 80 °C for 40 min, followed by immersion in a 0.1 M FeCl₃ solution for 30 min for chlorination treatment. After treatment, the sample was rinsed three times with ultra-pure water and then air-dried. Finally, the prepared Ag/AgCl reference electrode was immersed in a 0.1M KCl solution, and electrochemical impedance spectroscopy (EIS) testing was conducted to assess its stability. When the electrode impedance value was ≤500 Ω, it was considered to meet the performance requirements, thereby completing the preparation of the Ag/AgCl reference electrode.

Immunoassay of target NEFL

The NEFL antibody was dissolved at a concentration of 10 μg mL⁻¹ in 0.05 M, pH 9.6 carbonate buffer (CB buffer). 30 μL of the antibody solution was added to a high-binding microplate and incubated at 4 °C overnight to immobilize the antibody on the surface of the microplate. The remaining free sites on the electrode surface were blocked using BSA. Firstly, a 1% (w/v) BSA solution was prepared using 0.01 M, pH 7.0 phosphate-buffered saline (PBS) as the solvent. Subsequently, the microplate was incubated with 50 μL of the 1% BSA solution at 37 °C for 2 h. NEFL target solutions of different concentration gradients were prepared using the serial dilution method. Then 50 μL of antigen solutions at each concentration were added to the microplate and incubated at 37 °C for 40 min. After each step, the microplate was washed three times with buffer to remove the unbound material. Then, 50 μL of a 10 µg mL⁻¹ GOx@Au NP-anti-NEFL antibody (GOx@Au NP-pAb₂) solution was added to the microplate and incubated at 37 °C for 30 min. After incubation, the microplate was washed multiple times with washing buffer to thoroughly remove unbound signal antibodies. Then, 200 μL of a 10 mM glucose solution, at the optimized concentration, was added to the treated microplate. After the reaction was complete, the catalytic product mixture was transferred to the working electrode region of the electrode.

Results and discussion

Structural characterization of TiO₂-Fe nanoparticles

The central aim of this investigation is to optimize the electron transfer pathway by introducing novel intermediate energy levels via Fe atom doping, thus augmenting the sensitivity of PEC immunosensors. This is accomplished through the integration of the sensor system onto paper substrates to fabricate a portable photoelectrochemical immunosensing platform. As such, it is imperative to initially verify the successful synthesis of TiO₂-Fe and the manifestation of the anticipated high-efficiency signal conversion capability. Transmission electron microscopy (TEM) was employed to characterize the morphology and size of the synthesized TiO₂ and TiO₂-Fe materials (Fig. S1A). The TEM analysis revealed that the particle sizes of both the synthesized TiO₂ and TiO₂-Fe nanoparticles were predominantly concentrated around 100 nm, exhibiting a relatively homogeneous spherical morphology (Fig. 1A). Notably, Fe doping did not induce significant alterations in the particle size distribution or fundamental morphological features, suggesting that the incorporation of Fe elements had minimal impact on the growth mechanism of TiO₂ nanomaterials. High-resolution transmission electron microscopy (HRTEM) was further utilized to determine the interplanar spacing (Fig. 1B). The HRTEM results indicated that the selected area interplanar spacing of TiO₂ was measured to be 0.349 nm, which corresponded to the (120) crystal plane of anatase TiO₂. Conversely, the selected area interplanar spacing of TiO₂-Fe was found to be 0.236 nm, aligning with the (131) crystal plane of the same anatase crystal structure. Elemental analysis confirmed the uniform distribution of each element within it (Fig. 1B and S1B). Elemental mapping analysis was performed on the synthesized TiO₂-Fe nanoparticles to visualize the spatial distribution of Ti, O, and Fe elements (Fig. 1C). As shown in the mapping images, the green signal (Ti), red signal (Fe), and yellow signal (O) were uniformly distributed throughout the nanoparticle domain, with no local aggregation of the Fe signal observed. The uniform colocalization of iron with titanium and oxygen elements directly confirms the successful synthesis of Fe-TiO₂. The morphology of the synthesized TiO₂-Fe was characterized by scanning electron microscopy. The results revealed that the synthesized nanoparticles exhibited a rough, lychee-like particle structure with dimensions around 100 nm (Fig. S2).

Fig. 1 (A) TEM image of the synthesized Fe-doped TiO₂ nanoparticles. (B) High-resolution TEM images of selected areas of Fe-TiO₂ and the corresponding inverse Fourier transform images. (C) Elemental mapping surface scan analysis image of TiO₂-Fe, where green, red, and yellow represent Ti, Fe, and O, respectively.

To comprehensively elucidate the crystallinity and phase structure of the synthesized TiO₂ nanomaterials, X-ray diffraction (XRD) analysis was meticulously performed (Fig. S3). By juxtaposing the obtained diffraction patterns with the standard PDF card (PDF #29-1360), prominent diffraction peaks were discerned at 2θ angles of 25.3°, 37.3°, 37.8°, and 48.0°, which could be distinctly indexed to the (120), (201), (231), and (231) planes of anatase-type TiO₂.²⁴ This observation unequivocally validates the successful synthesis of TiO₂ nanomaterials featuring an anatase crystal structure. Undoped TiO₂ nanomaterials manifested pronounced peak intensities. Specifically, pristine TiO₂ exhibited sharp and intense diffraction peaks corresponding to the characteristic (120) and (231) crystal planes, which were indicative of high crystallinity. Conversely, upon the incorporation of Fe atoms into the TiO₂ lattice, the disparity in ionic radii between Fe³⁺ (ca. 0.64 Å) and Ti⁴⁺ (ca. 0.61 Å) instigated lattice distortion. This distortion not only diminished the crystallinity, but also exhibited the potential to modulate the migration pathways and recombination dynamics of photo-generated carriers. To further authenticate the surface chemical state of the TiO₂-based nanomaterials, Fourier-transform infrared spectroscopy (FT-IR) characterization was conducted. Rigorous spectral analysis revealed the conspicuous absence of typical vibration absorption peaks associated with organic functional groups within the fingerprint region and characteristic frequency range (Fig. S4). This absence strongly suggested that the surface of the nanomaterials was devoid of organic contaminants, thereby attesting to their high degree of cleanliness and purity. The binding energy of the TiO₂-based nanomaterial was also determined via X-ray photoelectron spectroscopy (XPS) as shown in Fig. S5. The results indicate that the introduction of Fe caused Ti and O to shift toward lower binding energies, thereby promoting the formation of an internal electric field near Fe (Fig. S6). The Fe X-ray fine satellite peak resolution similarly confirmed the coordination environments of Fe and Ti–O (Fig. S7). Collectively, these findings provided conclusive evidence for the successful synthesis of TiO₂-Fe nanomaterials.

Exploration of photoelectrical activity

To further investigate the application prospects of the synthesized TiO₂-Fe nanomaterials in the realm of photocatalysis, a 300 W Xe lamp-based photocatalytic reaction system was utilized to conduct a series of experiments aimed at assessing the photoconversion efficiency. In these experimental setups, a 0.1 M Na₂SO₄ solution was employed as the electrolyte, and a three-electrode configuration was established to construct the testing environment. Specifically, the working electrode was a screen-printed electrode modified with TiO₂-Fe, the counter electrode was a carbon-ink electrode, and the reference electrode was an Ag/AgCl electrode. Under intermittent illumination, the photoresponse characteristics of TiO₂-Fe nanoparticles and pristine TiO₂ nanoparticles were systematically characterized via the chronoamperometric technique. The experimental findings revealed that the photocurrent of TiO₂-Fe nanoparticles reached approximately 9.4 μA, whereas that of pure TiO₂ nanoparticles was merely 300 nA (Fig. 2A). The peak-like current was attributed to the reversible transformation of the redox states of Fe species in TiO₂ during the photoexcitation process. Compared with pristine TiO₂ nanoparticles, the photocurrent response of TiO₂-Fe nanoparticles was enhanced by a factor of 31. This notable enhancement could be primarily attributed to the fact that the incorporation of Fe elements substantially modified the electronic structure and light absorption properties of TiO₂ nanomaterials. On one hand, the introduction of Fe effectively suppressed the recombination probability of photo-generated carriers and prolongs their lifetime; on the other hand, Fe doping reduced the bandgap of TiO₂, thereby expanding the light response range and significantly improving the material's utilization efficiency of visible light. In addition, changes in the open-circuit voltage under light excitation were recorded, revealing that the photovoltage of TiO₂-Fe was 74.1 mV, while that of the TiO₂ electrode was only 39.3 mV (Fig. 2B). In a continuous 12 open-closed cycling experiment, the photocurrent response showed no significant decay. Under standard laboratory storage conditions, the batch-produced printed screen-printed paper-based electrodes also exhibited good stability (Fig. 2C). EIS testing results showed that undoped TiO₂ nanomaterials exhibited significant charge transfer resistance characteristics, appearing as large semicircles in the Nyquist plot, indicating high impedance during electron transfer (Fig. 2D). When Fe element was introduced into the TiO₂ lattice through doping, the diameter of the semicircle in the EIS spectrum was significantly reduced, intuitively reflecting that Fe doping could effectively reduce the charge transfer resistance of the material and promote rapid electron transfer between the material surface and the electrolyte solution.

Fig. 2 (A) Photocurrent experiment image of the paper-based three-electrode system. (B) Open-circuit voltage image of the paper-based three-electrode system under light excitation. (C) Stability analysis. (D) EIS of TiO₂ and TiO₂-Fe. (E) UV-Vis absorption spectra of the synthesized TiO₂ and TiO₂-Fe powders. (F) XPS valence band spectra and the corresponding valence band edge positions of the synthesized TiO₂ and TiO₂-Fe.

Additionally, the light absorption properties of the nanomaterials were characterized via powder UV-Vis spectroscopy, as shown in Fig. 2E. The results indicated that the incorporation of Fe broadened the light absorption spectrum of TiO₂ from the ultraviolet region to the visible region (∼600 nm). Combined XPS valence band spectrum analysis revealed that the valence bands of TiO₂-Fe and TiO₂ were 2.21 eV and 1.30 eV, respectively, with Fe doping causing a significant decrease in the valence band of TiO₂ (Fig. 2F). This was because the Fe 3d orbitals matched the valence band top energy level of TiO₂, and when Fe replaced Ti and formed a vacancy state, it hybridized with the O 2p orbitals, jointly leading to a reduction in the valence band top energy level. This alteration in the valence band structure enhanced the efficiency of photogenerated charge separation, providing a theoretical basis for improving the sensitivity of photoelectrochemical immunosensors. The specific band positions are shown in Fig. S8.

Analytical performance of the paper-based sensing platform

To assess the paper-based sensing platform's performance, especially its stability in mass production, a systematic study was carried out. Bending performance tests on the prepared paper-based electrodes examined their electrochemical responses under various bending angles and cycle counts, validating the current signal stability. Results showed no significant photocurrent decline over 10 consecutive tests, proving the platform's durability (Fig. S9). For paper-based electrode arrays made by mass printing, the photocurrent fluctuation among 16 electrodes in each batch was within 5%, indicating high process stability and reproducibility for large-scale production (Fig. S10).

The integration of paper-based electrodes and TiO₂-Fe nanomaterials presents a promising avenue for the development of a portable and efficient PEC sensing platform. The widespread availability and cost-effectiveness of TiO₂, coupled with the enhanced performance achieved through minimal Fe doping, contribute significantly to the control of raw material expenditures. The screen-printing technique employed herein is inherently straightforward, obviating the requirement for sophisticated vacuum or lithography apparatus. This enables the large-scale fabrication of electrodes under ambient conditions, thereby reducing both equipment and energy consumption costs. Leveraging biodegradable paper substrates, paper-based electrodes offer a more economical alternative to conductive glass counterparts. Additionally, their roll-to-roll manufacturing process streamlines production, minimizing time and labor costs, and aligning well with the cost-efficient demands of industrial applications. Collectively, these features endow the developed PEC sensing platform with distinct competitive advantages.

By further integrating a split-type immunosensing strategy, a highly sensitive and specific PEC detection system for biomarkers was successfully established. The study systematically investigated the influence of antigen–antibody specific binding at various stages on the photocurrent response. Further optimization of specific experimental parameters, including catalytic time and substrate concentration, was performed (Fig. S11 and S12). Under optimal reaction conditions, the photocurrent increased with the target concentration across a wide range of 0.01–200 ng mL⁻¹. Linear regression analysis revealed a good linear correlation between current intensity (I) and target concentration (C), with the fitted curve conforming to the equation I = 10.26 + 14.05 × lg C (R² = 0.9717), and the detection system exhibited an extremely low detection limit (LOD = 0.007 ng mL⁻¹) (Fig. 3A and B). Therefore, a systematic study was conducted on the anti-interference performance of the paper-based photodetection platform constructed based on the antigen–antibody specific binding mechanism. Additionally, the linear range and LOD values of the developed paper-based PEC sensing platform were compared with those of previously reported sensing platforms (Table S1). In simulated real-sample environments, the effects of various interference substances, including ions (Na⁺, K⁺, Cl⁻, SO₄²⁻), small molecules (uric acid, urea, creatinine), and large-molecule proteins (carcinoembryonic antigen, alpha-fetoprotein, prostate-specific antigen), on the sensor response were examined at different concentrations.^33,34 The results showed that, except for the target substances and mixed samples, the aforementioned interfering substances did not cause significant changes in the detection signals of the paper-based photoelectronic sensing platform, fully validating the sensor's excellent specific recognition capability (Fig. 3C).

Fig. 3 (A) Photocurrent data images of standard NEFL samples with gradient dilution, evaluated three times for each sample (0.01–200 ng mL⁻¹). (B) Regression analysis curve of photocurrent data from standard NEFL samples with gradient dilution, sampled from a stable photocurrent data platform. (C) Selective analysis, in which interference substances mainly include ions (1–4: Na⁺, K⁺, Cl⁻, SO₄²⁻), small molecules (5–8: uric acid, urea, creatinine, glucose), and interference proteins (9–11: carcinoembryonic antigen, alpha-fetoprotein, prostate-specific antigen), and mixture simple (12 and 13).

Conclusions

In summary, this study reports a portable paper-based PEC sensing platform based on a TiO₂-Fe photoresponsive system. The introduction of Fe significantly expanded the inherent light absorption range of TiO₂, thereby exhibiting excellent photoelectric conversion behavior. The paper-based PEC immunosensing platform constructed in this study demonstrates significant technical advantages: (i) a paper-based signal conversion system based on TiO₂-Fe nanomaterials can generate specific photoelectric response signals for target analytes under 10 s of xenon lamp irradiation, providing an efficient detection method for the rapid clinical diagnosis of neurodegenerative diseases; (ii) breaking through the equipment dependency limitations of traditional spectrometers, this sensor platform has excellent portability and scene adaptability, enabling widespread deployment in the point-of-care testing field. This study not only proposes a general-purpose, highly sensitive paper-based PEC bioanalysis strategy, but also provides innovative research ideas for the design and development of functional PEC conversion materials.

Author contributions

Cheng-Hua Xu: writing – original draft, visualization, software, methodology, investigation, and data curation. An-Yang Tao: Writing – original draft, investigation, and visualization. Mei-Qing Meng: visualization, investigation, formal analysis, and data curation. Xi-Le Zhang: investigation and formal analysis. Cheng Fang: investigation and formal analysis. Feng-Zao Chen: writing – review & editing, validation, supervision, project administration, funding acquisition, and conceptualization. De-Man Han: validation, supervision, and project administration.

Conflicts of interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Compliance with ethical standards

All the experiments were performed in accordance with the Guidelines of Taizhou University (China) and approved by the ethics committee at Taizhou University (China). Informed consent was obtained from human participants of this study.

Data availability

The data that support the findings of this study are available from the corresponding author upon reasonable request.

Supplementary information (SI): experimental section and optimization of experimental conditions. See DOI: https://doi.org/10.1039/d5an01030f.

Acknowledgements

This work was supported by the National Natural Science Foundation of China (No. 22204118).

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