Peroxidase Mimic Research Articles

Modulating Catalytic Activity and Stability of Atomically Precise Gold Nanoclusters as Peroxidase Mimics via Ligand Engineering.

Metal nanoclusters (NCs), composed of a metal core and protecting ligands, show promising potentials as enzyme mimics for producing fuels, pharmaceuticals, and valuable chemicals, etc. Herein, we explore the critical role of ligands in modulating the peroxidase mimic activity and stability of Au NCs. A series of Au15(SR)13 NCs with various thiolate ligands [SR = N-acetyl-l-cysteine (NAC), 3-mercaptopropionic acid (MPA), or 3-mercapto-2-methylpropanoic acid (MMPA)] are utilized as model catalysts. It is found that Au15(NAC)13 shows higher structural stability than Au15(MMPA)13 and Au15(MPA)13 against external stimuli (e.g., pH, oxidants, and temperature) because of the intramolecular hydrogen bonds. More importantly, detailed enzymatic kinetics data show that the catalytic activity of Au15(NAC)13 is about 4.3 and 2.7 times higher than the catalytic activity of Au15(MMPA)13 and Au15(MPA)13, respectively. Density functional theory (DFT) calculations reveal that the Au atoms on the motif of Au NCs should be the active centers, whereas the superior peroxidase mimic activity of Au15(NAC)13 should originate from the emptier orbitals of Au atoms because of the electron-withdrawing effect of acetyl amino group in NAC. This work demonstrates the ligand-engineered electronic structure and functionality of atomically precise metal NCs, which afford molecular and atomic level insights for artificial enzyme design.

A new spectrophotometric method for uric acid detection based on copper doped mimic peroxidase

Cascade reactions catalyzed by natural uricase and mimic peroxidase (MPOD) have been applied for uric acid (UA) detection. However, the optimal catalytic activity of MPOD is mostly in acidic conditions (pH 2–5), mismatching the optimal catalytic alkaline environment of uricase. In this paper, using CuSO4 and urea as raw materials, a MPOD with high catalytic activity in alkaline environment was synthesized by hydrothermal method. Then, based on coupling reaction of uricase/UA/MPOD/guaiacol (GA) system, a novel spectrophotometric method was established to detect 5–60 μmol/L UA (limit of detection = 3.14 μmol/L (S/N = 3)) and accurately quantified serum UA (275.6 ± 39.9 μmol/L, n = 5) with 95–105% of standard addition recovery. The results were consistent with commercial UA kit (p > 0.05). The MPOD could replace natural POD to reduce the cost of UA detection due to simple preparation and cheap raw materials, and is expected to achieve the specific detection of some substances, like glucose and cholesterol, combined with glucose oxidase and cholesterol oxidase.

Enzyme assisted magnetic hybrids as self-activated cascade reagent with synergistic activity for antimicrobial application

The abuse of antibiotics seriously threatens the public health, which leads to the urgent need to develop novel antimicrobial strategies. Reactive oxygen species (ROS) have been considered as an alternative approach for bacterial inactivation. At present, magnetic materials, especially iron oxide (Fe3O4) nanoparticles have gained widely application in biomedical field. Owing to the intrinsic peroxidase-like activity, Fe3O4 can catalyze the decomposition of hydrogen peroxide (H2O2) to generate hydroxyl radical (⋅OH) under acidic conditions. Recently, Fe3O4-based peroxidase mimics have been developed for wound infection therapy. However, the direct introduction of H2O2 with relative high concentration may induce undesired effect to normal tissues, and the infection site with near neutral pH still limits the antibacterial efficiency. Herein, carboxyl modified Fe3O4 immobilized silver nanoparticles (Ag NPs) and glucose oxidase (GOx) were synthesized as a self-activated cascade regent. GOx endowed the hybrids with catalytic activity, which could convert glucose into gluconic acid, which activated the peroxidase-like activity of Fe3O4 by decreasing the pH value. Moreover, the self-supplied H2O2 was subsequently catalyzed by Fe3O4 to generate ⋅OH. By combining the excellent antimicrobial performance of Ag NPs, the enzyme assisted magnetic hybrids exhibited robust antibacterial activity, which exhibited a great potential in bacteria-infected wound therapy.

Nanozymes from Cu(II) Metal–Organic Gel and Melamine for Highly Active Peroxidase-Like Activity to Detect Alkaline Phosphatase

Enzyme-like catalytic activity and efficiency of carbon-based nanomaterials are closely linked to their size, heteroatom composition, and structure, and hence the heteroatom regulation needs to be further explored. In this work, a simple and efficient strategy was proposed to develop Cu-doped 2D carbon material C3N4 (Cu-C3N4-550) with excellent catalytic performance by pyrolyzing precursors of Cu(II) metal–organic gel (MOG) and melamine directly. Due to its sufficient metal active sites and adequate specific surface areas, the as-prepared Cu-C3N4-550 was endowed with excellent peroxidase-like activity to promote the oxidation of 3,3′,5,5′-tetramethylbenzidine owing to the generation of •OH in the catalytic reaction. It was amazing to find that the peroxidase mimic activity of the prepared Cu-C3N4-550 has enhanced 32.3-fold compared with bare C3N4. High peroxidase-like activity of Cu-C3N4-550 was influenced severely by the addition of antioxidant ascorbic acid (AA), alkaline phosphatase (ALP) as a typical hydrolase could catalyze substrate 2-phospho-l-ascorbic acid into AA, while AA was capable of capturing •OH generated from the catalytic reaction of Cu-C3N4-550. Hence, a sensitive, selective, and colorimetric method for the detection of ALP was established, the linear concentration of ALP in this colorimetric sensor from 0.4 to 20 U/L was acquired with a low detection limit of 0.32 U/L. This work not only provides ideas for designing enhanced peroxidase-like activity nanozymes in practical biological analysis but also broadens the MOG derivatives and carbon-based nanomaterials in colorimetric applications.

Gelatin-based smart film incorporated with nano cerium oxide for rapid detection of shrimp freshness

A novel and convenient nano cerium oxide peroxidase mimic smart film for rapid detection of hypoxanthine (HX) concentrations to predict shrimp freshness was reported. Xanthine oxidase was incorporated into the smart film to react with HX and produce H2O2. Cerium oxide nanoparticles (CeNPs), as redox amplifiers and chromogenic agents, were oxidized by H2O2 to produce regular color changes to quantify HX concentrations. All components entrapped in a three-dimensional gelatin matrix crosslinked with glutaraldehyde. The nanostructure of CeNPs and the feasibility of their use as multifunctional nanozymes for HX detection were explored, as were the effects of CeNPs addition on the physical and colorimetric properties of the film. The morphology and HX sensitivity were also characterized, and their feasibility in monitoring shrimp freshness was further verified. The results showed the gelatin/glutaraldehyde film with 25% CeNPs had good performance. This film had a high sensitivity, with a linear range of HX detection of 6.2–200 μM and the lower limit of detection of 35 μM. Furthermore, the film could determine the shrimp freshness level with a predictive accuracy of approximately 0.9223. This method can be applied for packaging food as a freshness indicator because of its convenience, high sensitivity, and low cost.

Peroxidase mimics of platinum-group metals for in vitro diagnostics: opportunities and challenges.

Platinum-group metal (PGM) nanostructures with peroxidase-like catalytic activities (i.e., peroxidase mimics) have been actively developed and applied to in vitro diagnostics in recent years. This article provides our viewpoints on this emerging field from the perspectives of materials science and solid-state chemistry angles. We start with an introduction to PGM peroxidase mimics, their catalytic efficiencies, and insights into catalysis from computational simulations. We then discuss chemical approaches to the synthesis of PGM peroxidase mimics with desired physicochemical parameters and catalytic properties. Then, we elaborate on general methods for functionalizing the surfaces of PGM mimics with bioreceptors. Thereafter, we highlight the applications of PGM mimics in in vitro diagnostics, emphasizing the interactions of PGM mimics with other components of a diagnostic system. We conclude this article with our opinions on the challenges and opportunities in this field.

Thermostable protein-stabilized gold nanoclusters as a peroxidase mimic

By using a genetically modified thermostable protein (KTQ5C), we have synthesized protein-stabilized goldnanoclusters (AuNC@KTQ5C) with advantageous properties, such as heat stable fluorescent emission and heat resistant peroxidase-like activity.

Nitrocellulose membranes in situ grown with Prussian blue nanoparticles as stable nanozyme pads for colorimetric detection of dopamine.

Prussian blue (PB) is a typical peroxidase mimic with simple preparation, low cost and high eco-friendliness, but it still has drawbacks of poor stability (e.g., decomposition in aqueous dispersions) and intrinsic optical interference (e.g., high extinction coefficient over a wide wavelength range) in colorimetric assays. Herein, we used nitrocellulose (NC) membranes as synthesis hosts of PB nanoparticles (NPs) to develop a new type of three-dimensional (3D) porous nanozyme pad. By means of an in situ synthesis route, PB NPs were uniformly grown on the surfaces of the fiber scaffolds with desirable stability, which also avoided signal interference from PB NPs owing to the easy handling of the pads in a quantitative solid state. The pads showed significant peroxidase-mimicking activity toward the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) with the output of colorimetric signals. Based on the reduction of oxidized TMB (oxTMB) by dopamine (DA), the pads were exploited for simple and quantitative colorimetric detection of DA with a limit of detection (LOD) of 0.59 μM and a satisfactory accuracy for analysis of real human urine samples. This is the first attempt at exploiting NC membranes as the synthesis hosts to develop nanozyme pads, which solves the above drawbacks of traditional PB-based peroxidase mimics and has promise for various colorimetric bioanalyses, given the structural benefits of NC membranes and their broad applications in biosensors.

Ultrafast colorimetric detection of Cr(VI) using Fe3O4@polydopamine/Prussian blue composites as a highly efficient peroxidase mimic.

A recyclable peroxidase mimic Fe3O4@polydopamine/Prussian blue (Fe3O4@PDA/PB) composite was facilely prepared by coating PDA on an Fe3O4 nanoparticle core and in situ growth of PB nanoparticles on a PDA shell. The prepared Fe3O4@PDA/PB composite exhibited excellent peroxidase-like activity and can catalytically oxidize the colorless colorimetric substrate 3,3',5,5'-tetramethylbenzidine (TMB) into a blue colored product in the presence of H2O2 at 30 °C in 1 min. The catalytic mechanism was deduced to be the nanozyme-promoted generation of a hydroxyl radical (·OH), and the catalytic behavior followed the typical Michaelis-Menten kinetics. Based on Cr(VI)-boosted peroxidase-like activity of Fe3O4@PDA/PB, a simple and fast colorimetric method for detection of Cr(VI) was developed. Under the optimum conditions, the colorimetric method exhibited wider linear range (100 nM to 140 μM), low LOD (51.1 nM), good selectivity and short detection time (1 min). Moreover, the feasibility of the proposed colorimetric method was evaluated by determination of Cr(VI) in spiked tap water and lake water samples. Good recoveries (95.2-102.9%) and low relative standard deviations (RSDs) (1.6-4.4%) were obtained, showing great promise for practical use.

Fe(III)-incorporated porphyrin-based conjugated organic polymer as a peroxidase mimic for the sensitive determination of glucose and H2O2.

Nanozymes, i.e., nanomaterials that possess intrinsic enzyme-like behaviour, have thrived over the past few decades owing to their advantages of superior stability and effortless storage. Such artificial enzymes can be a perfect alternative to naturally occurring enzymes, which have disadvantages of high cost and limited functionality. In this work, we present the fabrication of an Fe(III)-incorporated porphyrin-based conjugated organic polymer as a nanozyme for the efficient detection of glucose through its intrinsic peroxidase activity and the amperometric detection of hydrogen peroxide. The iron-incorporated porphyrin-based conjugated organic polymer (Fe-DMP-POR) possesses a spherical morphology with high chemical and thermal stability. Exploiting the peroxidase-mimicking activity of the material for the determination of glucose, a detection limit of 4.84 μM is achieved with a linear range of 0-0.15 mM. The Fe-DMP-POR also exhibits a reasonable recovery range for the detection of human blood glucose. The as-synthesized material can also act as an H2O2 sensor, with a sensitivity of 947.67 μA cm-2 mM-1 and a limit of detection of 3.16 μM.

Hollow Co-CeO2@PEG nanospheres: Ultrasound enhanced cascade-nanozyme for synergetic anticancer

The limited H2O2 level in tumor microenvironment (TEM) constrains the effect of chemodynamic therapy (CDT). Herein, Co-CeO2@PEG hollow nanospheres are synthesized for two strategies of H2O2-supply. By varying hydrothermal times, hollow Co-CeO2 nanospheres (∼55 nm) are prepared derived from Ostwald ripening. Under ultrasound (US) irradiation, reactive oxygen species (ROS) are generated due to cavitation effect. ESR and capturing experiments reveal the US energy transfer (1O2) and US-generated electrons/dissolved O2 (H2O2) to determine ROS production. The doping of Co increases oxygen vacancy, which is in favor of O2 adsorption and ROS generation. Both density functional theory (DFT) calculation and experimental measurement reveal the decreased band gap of Co-CeO2. The emerging new energy band improves the separation/transfer of electron-hole and ROS production. Besides, hollow structure is significant to introduce extra cavitation nucleus, promoting cavitation effect and ROS generation. Furthermore, Co-CeO2@PEG nanospheres reveal glucose oxidase (GOx) activity, which not only can consume endogenous glucose to introduce starvation therapy but also to produce H2O2. The above two ways can realize intracellular H2O2 replenishment. In addition, Co-CeO2@PEG also show mimic peroxidase (POD) and catalase (CAT) behavior to introduce CDT and O2-supplementation. In comparison with pure CeO2@PEG, the greater nanozyme activity of Co-CeO2@PEG is ascribed to the lower resistance of charge transfer that is further testified by DFT and electrochemical analysis. Synergetic CDT/starvation/sonodynamic therapy (SDT) reveals noteworthy anticancer efficiency and also arouses immune response for metastasis inhibition.

Light-Mediated Modulation of Enzyme-Mimetic Activity of CuMnO2 Nanosheets.

As typical copper-based-manganese delafossites, CuMnO2 nanoflakes (CuMnO2 NFs) display excellent light absorption capacity, showing great promise for facile regulation of catalysis reactions by light stimulation. Here, we report that the as-prepared CuMnO2 NFs possess intrinsic peroxidase- and oxidase-like activities, which are capable of catalyzing the oxidation of the enzymatic substrate 3,3',5,5'-tetramethylbenzidine (TMB) in the presence of H2O2 or O2, respectively. Additionally, CuMnO2 NFs exhibit significant light-enhanced dual enzyme-like performance (peroxidase and oxidase) for accelerating the oxidation of TMB, owing to a much greater light-induced reactive oxygen species (ROS) yield. The light-induced hydroxyl radicals (·OH) and superoxide radicals (O2•-) hold the key to rapidly catalyzing the oxidation of TMB into the two-electron charge transfer complex diamine, subsequently improving the oxidation capacity and enzyme-like activity of CuMnO2 NFs as peroxidase and oxidase mimics. These findings confirm an efficient way to improve the enzyme-like performance of nanozymes and regulate the reactive oxygen species (ROS) level by simply employing irradiated light. This work also provides a general method for constructing external field-modulated smart nanozymes and an insightful approach to the corresponding enzyme-mimetic reaction mechanism.

Platinum-Group Metal Nanoparticles as Peroxidase Mimics: Implications for Biosensing

Over the past few decades, peroxidase mimics made of platinum-group metal nanoparticles (PGM NPs) have been actively developed and applied to various biosensing platforms. Nevertheless, there is a lack of a comprehensive study that compares the peroxidase-like activities of PGM NPs and their performance in biosensing. Here, we report a systematic study of PGM NPs as peroxidase mimics, including Pd, Pt, Rh, and Ir NPs. NPs of these elements were uniformly synthesized and their nanoscale features were probed to ensure a consistent size, shape, and chemical ligand on the surface. Our measurements indicate that the Ir NP is the most active one with a catalytic constant as high as 6.27 × 105 s–1, followed by Pt, Rh, and Pd NPs. The binding affinities of NPs to peroxidase substrates during catalysis were also quantitively analyzed and compared. Using enzyme-linked immunosorbent assay as a model biosensing platform, the performance of PGM NPs in detecting carcinoembryonic antigen (a cancer biomarker) was evaluated. The results showed that the detection sensitivity was correlated to the catalytic activity of PGM NPs, where Ir NPs achieved the highest sensitivity with a limit of detection at the level of low picogram per milliliter.

Recent Advances in the Immunoassays Based on Nanozymes.

As a rapid and simple method for the detection of multiple targets, immunoassay has attracted extensive attention due to the merits of high specificity and sensitivity. Notably, enzyme-linked immunosorbent assay (ELISA) is a widely used immunoassay, which can provide high detection sensitivity since the enzyme labels can promote the generation of catalytically amplified readouts. However, the natural enzyme labels usually suffer from low stability, high cost, and difficult storage. Inspired by the advantages of superior and tunable catalytic activities, easy preparation, low cost, and high stability, nanozymes have arisen to replace the natural enzymes in immunoassay; they also possess equivalent sensitivity and selectivity, as well as robustness. Up to now, various kinds of nanozymes, including mimic peroxidase, oxidase, and phosphatase, have been incorporated to construct immunosensors. Herein, the development of immunoassays based on nanozymes with various types of detection signals are highlighted and discussed in detail. Furthermore, the challenges and perspectives of the design of novel nanozymes for widespread applications are discussed.

Silver nanoparticles@metal-organic framework as peroxidase mimics for colorimetric determination of hydrogen peroxide and blood glucose

• A colorimetric strategy for sensitive detection of H 2 O 2 and glucose has been developed. • A broad dynamic response range for glucose in the range from 0.5 to 50 μM with a limit of detection of 0.23 μM. • The proposed method can be used to monitor glucose in human serum samples. As a major origin of energy for cellular activity and intermediates product of metabolism, glucose plays a significant role in biological activity. Thus, it is relatively vital to explore a sensitive and accurate approach to monitor blood glucose. Herein, a colorimetric biosensor for sensitive, rapid, and reliable monitoring of blood glucose was successfully established. In this sensing system, a nanocomposite of silver nanoparticles (AgNPs) depositing on the MIL-101 (Fe) surface (AgNPs@MOF) was employed as an excellent peroxidase mimic. Glucose could be effectively catalyzed by glucose oxidase (GOx) to produce hydrogen peroxide (H 2 O 2 ) and gluconic acid. Afterward, the AgNPs@MOF nanozyme catalyzed the oxidation of the colorless material TMB into the blue product oxTMB by H 2 O 2 , causing a noticeable change in absorbance at 652 nm. A colorimetric glucose measurement platform was proposed with preferable simplicity and high sensitivity using GOx and AgNPs@MOF nanozyme that with peroxidase-mimicking activity. The limit of detection of the established biosensor was down to 0.17 μM for H 2 O 2 and 0.23 μM for glucose. More importantly, as a result of the application of this biosensor with human blood samples, glucose levels could be monitored with high accuracy, which indicated that this approach had great potential for diabetes therapy. A colorimetric biosensor for sensitive, rapid, and reliable monitoring of blood glucose was successfully established. In this sensing system, a nanocomposite of silver nanoparticles (AgNPs) depositing on the MIL-101 (Fe) surface (AgNPs@MOF) was employed as an excellent peroxidase mimic. Glucose could be effectively catalyzed by glucose oxidase (GOx) to produce hydrogen peroxide (H 2 O 2 ) and gluconic acid. Afterward, the AgNPs@MOF nanozyme catalyzed the oxidation of the colorless material TMB into the blue product oxTMB by H 2 O 2 , causing a noticeable change in absorbance at 652 nm.

Pt nanoparticle/N-doped graphene nanozymes for colorimetric detection of acetylcholinesterase activity and inhibition

Herein, Pt nanoparticles encapsulated in 2D N-doped graphene (Pt/NG) nanozymes containing abundant active Pt sites is rationally fabricated by a one-pot pyrolysis of EDTA-Pt complex. The two-dimensional sheet structure and the reconstructed electronic structures enable Pt/NG nanozymes exhibiting excellent peroxidase-mimicking activity. Taking advantages of the remarkable catalytic activity of Pt/NG nanozymes, a colorimetric assay is strategically constructed for sensitively determining acetylcholinesterase (AChE) activity and tacrine based on the thiocholine-regulated effect towards the peroxidase-like activity of Pt/NG. The detection limit for AChE activity can achieve as low as 0.0652 mU mL −1 , and the IC 50 value of tacrine for AChE is about 0.954 µM. The robust analytical performance in serum samples test verifies the great application potential of this assay in real matrix. Furthermore, “IMPLICATION-AND” logic gate is constructed based on the proposed colorimetric assay. This work not only provides one sustainable and effective avenue to fabricate Pt-based peroxidase mimic with high catalytic performance, but also gives new impetuses for developing novel colorimetric assay by applying Pt-based enzyme mimics as substitutes for the natural enzymes.

Cascade reaction biosensor based on gold nanocluster decorated iron-cobalt oxide nanosheets as a superior peroxidase mimic for dual-mode detection of α-glucosidase and its inhibitor

Herein, we have synthesized a novel kind of gold nanoclusters decorated iron-cobalt oxide nanosheets (His-AuNCs@FeCo-ONSs) assembled by electrostatic interaction, which possessed both outstanding peroxidase-like activity and fluorescence property. Taking advantage of our bifunctional hybrid nanozyme and enzyme cascade reactions, a sensitive dual-mode (colorimetric/fluorescent) detection method for α-glucosidase was constructed. The detection limits for α-glucosidase were 2.2 U/L and 3.3 U/L in fluorometric and colorimetric mode, respectively. This method not only provides high sensitivity, but also can correct itself to improve the accuracy of analysis due to the dual-response signals. Furthermore, it was employed for α-glucosidase determination in real samples and screening of α-glucosidase inhibitors.

Usnea antarctica (James Ross Island, Antarctica) and Usnea subfloridana (Uludağ, Turkey) Incorporated Hybrid Nanoflowers with Their Intrinsic Catalytic and Antimicrobial Activities

AbstractHerein, we used lichen extracts, for the first time, in formation of organic‐inorganic nanoflowers and we utilized them as novel nanobiocatalyst and antimicrobial agents. Two lichen species, Usnea Antarctica (U. Antarctica) and Usnea subfloridana (U. subfloridana) were collected from James Ross Island (Artarctica Peninsula, Antarctica) and from Uludağ National park (Bursa, Turkey), respectively and both species were separately extracted in methanol and water. In terms of nanoflower synthesis, the extracts were used as organic components and copper (II) ions (Cu2+) acted as inorganic components for formation of U. antarctica and U. subfloridana based hybrid nanoflowers (hNFs). The peroxidase mimic catalytic and antibacterial activities of the hNFs were systematically examined towards model substrate and bacteria strains, respectively via Fenton reaction mechanism. We demonstrated how the types of lichens and their extracts prepared in methanol and water influence morphologies, catalytic and antibacterial activities of the hNFs. We found that no 2 hNFs formed from U. subfloridana extract prepared in water exhibited highly enhanced catalytic activity owing to its uniform, more compact and porous flower‐like structures with high surface‐to‐volume ratio. We expect that lichens extracts based hNFs with great catalytic activities can be promisingly alternative against enzymes and used in various scientific and technical fields.

Colorimetric Test Paper for H2O2 Determination Based on Peroxidase‐Like Activity of an AuFe/ZIF‐8‐Graphene Composite

AbstractHerein, we prepared a bisuppport‐based bimetallic nanoparticle composite AuFe/ZIF‐8‐graphene. It exhibited high peroxidase activity compared to those hybrids without graphene support, which demonstrated the regulatory effect of graphene on the catalytic activity. SEM, TEM, EDX, XRD, FT‐IR and XPS characterization showed that Au and Fe3O4 NPs were successfully immobilized on ZIF‐8‐rGO carriers. Kinetic analysis confirmed that the fabricated composites showed better Km and Vmax for H2O2 than HRP and some other reported peroxidase mimics. Under the optimal conditions, a sensing assay for H2O2 determination was constructed with a linear detection range of 0.01–2.5 mM and a detection limit of 0.63 μM. On this basis, a colorimetric test paper with its standard card was prepared for rapid visualization detection. It realized H2O2 detection in the range of 0.01‐500 mM and was further successfully applicated in the semi‐quantitative detection of H2O2 in actual spiked water samples. This achievement was expected to encourage the development of rapid visual detection for other pollutants in environmental and medical field.

Food-borne melanoidin based peroxidase mimic for the precise detection of total antioxidant capacity

Drawing on the structure of the natural horseradish peroxidase (HRP) active site, in this study, food-borne Melanoidin and Fe3+ were used to construct a peroxide mimic enzyme, which possessed excellent peroxidase (POD)-like activity with the Km values of 0.088 mM and 0.358 mM toward 3,3′,5,5′-tetramethylbenzidine (TMB) and hydrogen peroxide (H2O2), respectively. Given the good POD-like activity of Melanoidin/Fe3+, a convenient, selective, and sensitive method for the determination of ascorbic acid (AA) was defined. The method covered the 10–100 μM AA concentration range with a 1.19 μM detection limit. With favorable anti-interference and strong stability, Melanoidin/Fe3+ was successfully applied to the determination of AA in foods and medicines. The high activity of Melanoidin/Fe3+ provides a new strategy for designing high-performance nanozymes, which has great potential in exogenous dietary and healthcare applications.