A Review on Metal- and Metal Oxide-Based Nanozymes: Properties, Mechanisms, and Applications

The characteristics of metal- and metal oxide-based nanozymes with diverse construction are dissertated. The intrinsic properties and catalytic mechanism of metal- and metal oxide-based nanozymes are discussed. The recent applications of metal- and metal oxide-based nanozymes in biological analysis, relieving inflammation, antibacterial, and cancer therapy are reviewed.


Introduction
Enzymes are environmentally friendly biomaterials with remarkable catalytic efficiency and substrate specificity produced by living cells [1,2]. Most of the natural enzymes are proteins, while a small part are RNA. The past decades have witnessed the extensive progress of biological enzymes in biology, medicine, chemistry, and industrial science [3]. Nevertheless, the complicated preparation procedure, unstable catalytical activity and intrinsic environmental sensitivity have restricted the scalable utilization of natural enzymes [4,5]. Therefore, the exploration of alternative artificial enzymes to overcome shortcomings of natural catalysts has become an issue of increasing concern. The evolution of nanotechnology and biology provides a bridge toward novel artificial enzymes. After the pioneering work of Gao et al. [6] reporting ferromagnetic (Fe 3 O 4 ) nanoparticles (NPs) with enzyme-mimicking property in 2007, a bunch of nanozymes have been demonstrated as natural catalysts mimics. For instance, Au@Co-Fe hybrid NPs [7], CuCo 2 S 4 NPs [8], MnO 2 nanowires (NWs) [9], Pt nanoclusters (NCs) [10], Au@Pt nanorods (NRs) [11], and carboxyl-modified graphene oxide (GO-COOH) [12] have been reported as peroxidase (POD) mimics. Nanozymes with multi-enzyme-type activities (e.g., Co(OH) 2 /FeOOH/ WO 3 ternary nanoflowers [13], AuNPs [14,15], Co 3 O 4 NPs 1 3 [16], AgPt NPs [17], N-doped sponge-like carbon spheres [18], Mn 3 O 4 NPs [19]) have been exploited in diverse investigation. Up to date, more than 540 types of nanozymes have been synthesized by over 350 research laboratories from 30 countries [20]. Generally, existing nanozymes are affiliated with two categories, namely oxidoreductase family and hydrolase family. Carbon-based materials, metal, and transition metal compounds are the most common nanozyme composition materials [21]. Wu et al. reviewed the history of nanozyme and draw a brief timeline for the evolution of artificial enzymes and natural enzymes ( Fig. 1) [22]. With extensive efforts devoted to the investigation of artificial enzymes and nanotechnology, creative breakthroughs have been made steadily on the catalytic mechanisms and intrinsic properties of nanozymes, as well as the application field. In the past two years, the investigation on singleatom nanozyme (SAN) has aroused numerous attention due to their outstanding activity and selectivity [23,24]. In the research of Kim et al. [25], the Fe-N-rGO SAN showed the best catalytic efficiency for different substrates among various classical POD mimics and natural HRP. Niu et al. [26] reported that the Fe-N-C SAN not only possessed excellent enzymatic activities, but also exerted splendid stability and robustness within a broad temperature and pH range.
Since nanozymes are recognized as a class of functional nanomaterials, they possess both the unique nature of nanomaterials and enzyme-like activity [27]. The surfaces of metal and metal oxide nanomaterials are covered with a large amount of charge, which was responsible for their superb electron properties [28]. Consequently, metaland metal oxide-based nanozymes stand out in the area of electrocatalysis, sensing and fuel cells [29,30]. Furthermore, as promising alternatives for natural biocatalysts, they commonly retained better stability and robustness under extreme conditions than natural enzymes [5]. The prominent physicochemical properties (e.g., high surface energy, superior optical, and photothermal conversion properties), as well as simplicity in preparation and storage also broaden their applications [31]. Interestingly, the catalytic performance and physicochemical properties of metal and metal oxide nanomaterials could be easily regulated according to the practical demand [32,33]. For instance, surface modification has been revealed as a promising strategy to increase the biocompatibility of these nanozymes [34][35][36]. The structure design associated with the catalytic efficiency is flexible through suitable control of synthetic conditions [37]. Given the above ascendency, the research fields of metaland metal oxide-related nanozymes have gradually extended from environment to chemical industry, food, agriculture, biomedicine, medicine, and so forth [38][39][40]. Even though tremendous efforts were devoted, further promotion of this kind of nanozymes is still facing difficulties. For example, the enhancement of catalytic activity and selectivity, closely associated with the sensitivity and specificity of nanozymebased biosensors, remains a challenge [25,41]. In addition, the strengthened physiological stability and biological safety is vital for the spread application of nanozymes in clinical medicine [42]. Therefore, novel nanozymes and biotechnology are urgently needed to make up these defects.
Dozens of excellent reviews concerned with nanozymes have been published in recent years. Some of the reviews involved the research progress of nanozymes in a particular field [5,21,27,43,44]. Some researchers organized and revealed the natural activities and working mechanisms of specific nanozymes [45][46][47][48][49]. In 2019, Huang et al. [50] systematically discussed the classification, intrinsic nature, enzymatic mechanisms and potential applications of nanozymes for the first time. However, a thorough overview for metal-and metal oxide-based nanozymes is still lacking. In this review, we firstly elucidate the characteristics and synthetic methods of metal-and metal oxide-based nanozymes. Then, we will dig into the catalytic mechanisms and property regulation of these nanozymes. After introducing their appliance in biological analysis, relieving inflammation, antibacterial and cancer therapy, we finally discuss the present challenges and give a future perspective for the research of nanozymes constituted of metal and metal oxide.

Preparing Diverse Nanozymes with Constructive Feature
Generally speaking, the existing metal-and metal oxidebased nanozymes can be roughly assorted into monometal [51], metal alloy [52][53][54], metal oxide [6,55,56], metallic core/shell nanostructure [57][58][59], and hybrid [60] nanomaterials in terms of constructive feature. Monometal nanozymes are usually noble metal nanomaterials possessing prominent chemical stability under natural conditions. They commonly possess facile conjugation sites to diverse biomolecule ligands and antibodies, remarkable surface plasmon resonance (SPR) properties, superior optical, and photothermal conversion properties [61][62][63]. However, bare monometal nanoparticles (e.g., Ag, Pt) tend to aggregate into nanoclusters, resulting in decreasing of catalytic activity [64]. What is worse, most bare noble metal nanozymes (except Au) have biological toxicity, thus limiting their application in clinical medicine. The structure, size, and morphology have been proved to influence the catalytic properties of these nanozymes [65][66][67]. Monometal nanozymes could be prepared through preformed-seed-mediated growth [68], hightemperature reduction method [2,[69][70][71], electrochemical synthesis, photochemical method, biosynthesis [72,73], and spatially confined medium/template approach [74]. With different methods, various forms of noble metal nanomaterials (e.g., nanoparticles [14,15], nanoclusters [10], nanorods [75], nanosheets [76], nanocubes) could be obtained. The preformed-seed-mediated growth is feasible for size control by changing the concentration and nature of seeds in the growth solution [77]. A variety of small molecules (e.g., tannic acid [71], citrate [78]) and macromolecular templates including DNA [79], dendrimers [80], and proteins (e.g., bovine serum albumin, human serum albumin, lactoferrin, pepsin, insulin) [2,70] have been employed for monometal nanozymes synthesis via the high-temperature reduction procedure. The electrochemical strategy could modulate the size and morphology of noble metal nanomaterials through controlling electrodeposition parameters during the deposition process. Metal alloy nanozymes, containing bimetal alloys and multimetallic alloys, could be obtained via common chemical synthesis such as the one-pot strategy [81], galvanic replacement reaction [82,83], co-reduction method [84,85], hydrothermal growth [86], and electrodeposition method [87,88]. Besides, biological strategy [89] and bimetallic nanomaterials printing [90] have been present as favorable synthesis method as well. The biological strategy is widely known as a green synthesis method with biological elements as the reducing agents or growth template (e.g., leaf extract, plant extract, DNA) [91,92]. Along with the preparation of diverse nanoalloys, researchers found that the composition as well as structure affected the enzymatic characteristic of metal alloy nanozymes [93,94]. Therefore, adjusting the proportion of various metals, enlarging porosity and specific surface area of alloy nanomaterials have been recognized as effective approaches to regulate activity. Generally, the cost of metal alloy nanozymes is much lower than that of monometal nanomaterials as the incorporation of nonprecious metals. Owing to the synergistic effect of the two components, bimetal nanoalloys tend to exhibit superiorly optical and chemical properties, as well as better catalytic performance compared with noble metal nanomaterials [95]. Furthermore, the introduction of magnetic metal (e.g., Co, Fe, and Ni) could endow alloys with magnetism besides optimizing their enzymatic properties [83,84].
Possessing high surface energy and surface-to-volume ratio, metal oxide nanozymes have been considered as promising artificial enzymes for decades [96]. The most common metal oxide nanozymes like CeO 2 , Fe 2 O 3 , Fe 3 O 4 , Co 3 O 4 , Mn 2 O 3 , and Mn 3 O 4 nanomaterials have all been reported to possess multi-enzyme-like activities [97]. In addition, they exhibit plenty of unique properties such as magnetic, fluorescence quenching and dielectric properties [98]. Compared with precious metal nanomaterials, metal oxide nanozymes commonly exert lower price and concise synthesis process [99]. Furthermore, the low biological toxicity and favorable accumulation in biological tissues have broadened their application toward biopharmaceutical [100]. Nevertheless, there are some disadvantages of unmodified metal oxide nanozymes in terms of biology. For instance, they might show awful stability and accelerate the generation of harmful free radicals under physiological conditions [101]. Additionally, the improper surface ligands coating would lead to the failure control of drug release [102]. In recent, diverse methods have been employed for metal oxide nanozymes preparation, including the hydrothermal [103,104], solvothermal [105,106], pulsed laser ablation [107], co-precipitation [108,109], sol-gel [110], and thermal decomposition method [111].
The metallic core/shell (inorganic/inorganic) nanostructure-based nanozymes could be prepared through the hydrothermal reaction [112], solvothermal method [113], sol-gel approach [114], and atomic layer deposition [115]. By combining different materials and modifying structure, researchers could regulate the stability and functionality of core/shell structure-based nanozymes conveniently [116]. For example, the introduction of SiO 2 as coating significantly realized good stability and reduced bulk conductivity of the core particles [117]. The dispersion and biological safety of magnetite NPs encapsulated by silica could also be improved when existed in physiological environment [118]. In addition, the Au-coated nanostructure-based nanozymes have demonstrated to show enhanced chemical stability, biocompatibility, and optical properties [119,120]. However, the accessibility between substrate and the active phase of nanozymes could be affected by coating materials [121]. Therefore, regulating coatings' thickness, porosity, and synthesis procedure was demanded to modulate enzyme-like capacity and other chemical properties of nanozymes.
The metal-and metal oxide-based hybrid nanozymes could be prepared with organic molecules or polymers modified on the surface of metal or metal oxide nanomaterials [122,123]. The modifications on the surface of hybrid nanozymes are used to optimize the catalytic performance, instead of acting as stabilizer during the synthesis process [124]. Generally, the intrinsic properties of hybrid nanozymes might be ascribed to size, content, and components structure [125,126]. For instance, polymer/metal nanozymes have been revealed to show stable catalytic capacity in which metal nanoparticles are evenly dispersed in polymer [127,128]. In parallel with enhancing catalytic activity and selectivity, the incorporation of polymer or organic molecule endows hybrid nanozymes with amazing physical, chemical, and mechanical properties (e.g., adsorption [129], water solubility [130], biodegradability [131]), thereby expanding their application in miscellaneous fields [124].
The catalytic activities and efficiency of metal-and metal oxide-based nanozymes involved in the recent reports are listed in Table 1. These nanozymes mainly imitate four kinds of natural enzymes, namely POD, oxidase (OXD), catalase (CAT), and superoxide dismutase (SOD). The Michaelis-Menten constant (K m ) and maximal velocity (V max ) reflects the enzyme affinity with its substrate and maximal reaction velocity respectively [132]. And the K cat is the maximum number of substrate molecules converted to product per enzyme molecule per second. The lower value of K m and the higher value of V max indicate the stronger catalytic activity of nanozymes.

Properties of Metal-and Metal
Oxide-Based Nanozymes

Catalase-Like Activity
CAT is a kind of binding enzyme with iron porphyrin as its prosthetic group [161]. CAT presents in the living tissues could catalyze hydrogen peroxide (H 2 O 2 ) into oxygen and water, hence protecting tissues from excessive H 2 O 2 [162].
Up to now, a series of metal-associated nanozymes, such as platinum (Pt) [51], gold (Au) [163], CeO 2 [164], Mn 3 O 4 [19], have been demonstrated to show CAT-like activity. Although promising in anti-inflammatory, tumor treatment, biological detection and many other fields, considerable CAT mimics still constrained by the obscure mechanism [165,166]. Li et al. [167] verified that the pre-adsorbed OH group on the surface of noble metal served as the active site for CAT-like catalytic reaction. Although most reported nanomaterial-based CAT mimics showed favorable catalysis ability in neutral and alkaline environment, Liu et al. [80] firstly reported that amine-terminated PAMAM dendrimer encapsulated gold nanoclusters (AuNCs-NH 2 ) displayed CAT-mimicking property not only in acidic environment but also over physiological pH range (i.e., pH 4.8-7.4). They speculated that the protonation of tertiary amines from dendrimers in acidic solution could stimulate pre-adsorbing OH, thus providing active sites for H 2 O 2 decomposition to generate oxygen and water.
In terms of metal oxide nanozymes, Celardo et al. put forward a possible catalytic model of CeO 2 NPs with CATmimicking properties in 2011 [168]. In the system, H 2 O 2 was firstly bind to the 2Ce 4+ binding site presented by the oxygen vacancy site of CeO 2 NPs (Fig. 2a➀, ➁). Then, the fully reduced oxygen vacancy site was formed as the protons released and two electrons transferred to the two Ce 4+ (Fig. 2a➂). The oxygen was generated from the reduced oxygen vacancy site (Fig. 2a➃). Afterwards, another H 2 O 2 molecule was bind to the 2Ce 3+ site (Fig. 2a➄). The homolysis of O-O bond happened with the transfer of two electrons and a uptake of two protons (Fig. 2a➅). After H 2 O molecules released, the initial Ce 4+ sites were regenerated on nanoceria surface. Interestingly, Mu et al. reported that a larger concentration of the perhydroxyl anion (OOH − ) contained in H 2 O 2 molecule were existed in the neutral and alkaline solution [103]. The OOH − then might interact with metal centres of Co 3 O 4 and form the · O 2 H due to its prominent nucleophilic ability compared with H 2 O 2 . With terephthalic acid as the fluorescent probe, it could be found that the production efficiency of the hydroxyl radical ( · OH) depended on the Co 3 O 4 concentration, indicating that the CAT-type property of Co 3 O 4 NPs would influence the decomposition of H 2 O 2 to · OH. Moreover, thermodynamic and kinetic analysis revealed that there might be more "active sites" on the surface of Co 3 O 4 NPs than natural CAT owing to the stronger affinity between H 2 O 2 and Co 3 O 4 compared with natural CAT.
The existing hypothetical mechanisms for the CAT-like property of CeO 2 NPs and Co 3 O 4 NPs mentioned above still BSA bovine serum albumin, PVP polyvinylpyrrolidone, PNPP p-nitrophenyl phosphate, LDH layered double hydroxides, HS heparin sodium, RGO reduced graphene oxide, His histidine, GOx glucose oxidase, HRP horseradish peroxidase, OPD o-phenylenediamine, NSs nanosheets, HSA human serum albumin  [160] show certain limitations due to the neglect of the real structural features discussion [169]. Therefore, Guo et al. [170] investigated the possible catalytic mechanism of CAT-type activity at atomic or molecular level, involving the baselike dissociative, acid-like dissociative, and bi-hydrogen peroxide associative mechanisms. Based on the calculation of thermochemical energies and associated activation barriers, they reported that the bi-hydrogen peroxide associative mechanism was most viable for the CAT-mimicking catalytic recycle for Co 3 (111) surface and H 2 O 2 was also exoenergetic and kinetically favorable as well (Fig. 2e).
The catalytic mechanisms of various nanomaterial-based POD mimics could generally be concluded as Fenton or Fenton-like reaction or the electron transfer process [117]. Wang (Fig. 3b). What is more, the production effect of the catalytic centers on the surface of γ-Fe 2 O 3 was demonstrated to be at least 50-fold higher than that of the dissolved metal ions.
The nanocrystalline structure of nanozymes was also considered to make contribution to the H 2 O 2 -activating ability. André et al. reported that the intrinsic POD-like activity of V 2 O 5 nanowires was attributed to surficial properties of the nanozymes instead of free orthovanadate anions [189]. They proposed a likely reaction mechanism based on analyzing the layered V 2 O 5 orthorhombic structure (Fig. 3c). The (001) surface and the (110) surface were predominantly connected to the selective oxidation of hydrocarbons and total oxidation, respectively. The surface sites on the exposed (010) lattice planes of V 2 O 5 NWs was assumed to be related to their enzyme-like property. The V atoms in the (010) plane and the electron lone pairs of the bridging oxygen atoms, respectively, acted as Lewis acid and base sites. Consequently, an intermediate peroxo species was produced after the reaction between V 2 O 5 NWs and H 2 O 2 (Fig. 3d). Afterward, the ABTS was bind to the vanadium peroxo species via a nucleophilic attack and then oxidized into ABTS* + species. The regeneration of the V 2 O 5 NWs required another ABTS molecule since H 2 O 2 is a two-electron oxidant.
In recent years, the electron transfer-related mechanism was applied to a bunch of POD mimics such as IrO 2 / rGO nanocomposites [123], FePt-Au hybrid NPs [190], Co 3 O 4 NPs [191], and AuNPs@CDs nanocomposites [122]. Han et al. [9] obtained recyclable biotemplate-based MnO 2 nanowires with genetically engineered filamentous phages M13 as template. As illustrated in Fig. 3e, an electron transfer model was proposed for the reaction mechanism. With an electron transferred to MnO 2 NWs, the first substrate ABTS was oxidized. Then, another electron would transfer from MnO 2 to H 2 O 2 and hence produced H 2 O molecules. According to the chromogenic reaction and a series of control experiments, the enhanced POD-mimetic capacity of 1D M13-E4@MnO 2 nanozymes could be attributed to the surface effect, the small size effect and the homogeneous distribution of nanocrystals. When it comes to noble metal nanozymes, Cui et al. [71] speculated that Ir NPs could serve as the electron transfer mediators between H 2 O 2 and 3,3′,5,5′-tetramethylbenzidine (TMB) (Fig. 3f). TMB adsorbed on the Ir surface provided lone-pair electrons from amino group to the Ir NPs, whose electron density was consequently increased. The electrons that transferred from the Ir NPs to peroxides would accelerate the oxidation of TMB and the reduction of H 2 O 2 .

Oxidase-Like Activity
Oxidases catalytically oxidize substrate (electron donor) and produce H 2 O or H 2 O 2 in the presence of oxygen, which is served as the electron acceptor. The oxidase family is classified according to the acting group of donors, including amino groups, CH-OH group (GOx), Ph-OH group (polyphenol oxidase), sulfur group (sulfite oxidase, SuOx), and ferrous ions (ferroxidase and cytochrome c oxidase) [192]. Among them, the OXD-mimetic nanozymes that acting on amino groups were widely investigated. Up to date, a large amount of metal-based and metal oxide-based oxidase mimics have been uncovered, such as CuO [193], MnFe 2 O 4 [194], and Pt@MnO 2 [58]. The formation of intermediates (e.g., singlet oxygen, oxygen superoxide anion) and electron transfer process have been demonstrated to have important impacts on the OXD-type properties of these nanozymes [195].  ABTS (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)). Adapted from a Ref. [187], b Ref. [188], c, d Ref. [189], e Ref. [9], f Ref. [71] with permission 1 3 [196].  [197]. In the reaction mechanism, the O 2 molecules were adsorbed onto defect sites of nanoceria and converted into O 2 ·− under acidic conditions (Eq. 4). As the surface Ce 4+ reduced to Ce 3+ , the TMB was oxidized into TMB ox (Eq. 6). As the main intermediate, the in situ produced O 2 ·− finally regenerated Ce 4+ via the oxidation of Ce 3+ , accompanied by the generation of water (Eq. 7). Alternatively, the oxidation of TMB could be directly initiated by O 2 ·− as well (Eq. 5).  6 ]. c Catalytic mechanism of Au NPs as GOx mimics. Adapted from a Ref. [196], b Ref. [198], c Ref. [199] with permission Mechanism study on the nanozymes mimicking the other members of the oxidase family has made great progress as well. Following the exploration on the MoO 3 NPs as SuOx mimics [198], Chen et al. synthesized PEGylated (polyethylene glycol)-MoO 3−x nanoparticles (P-MoO 3−x NPs) that could catalytically oxidize sulfite. As shown in Fig. 4b, the sulfite was oxidized into sulfate with the two electron oxidative hydroxylation. Following the reduction of [Fe(CN) 6 ] 3− , one electron then transferred in succession to the Mo V intermediate for the stabilization of the inactive Mo IV state. In terms of nanozymes with GOx-like acticity, Comotti et al. put forward a two-electron mechanism to explain the intrinsic catalytic activity of the Au NPs ( Fig. 4c) [199]. In their model, the hydrated glucose anions that formed in the presence of alkali were adsorbed on the surface of AuNPs. The gold surface atoms on the hydrated glucose then activated molecular oxygen and formed the dioxogold intermediate, for the electron transfer. After two electrons transferring from glucose to dioxygen, the gluconic acid and H 2 O 2 were finally generated. Zhang et al. [200] prepared crown-jewelstructured Au/Pd nanoclusters with high reactivity. The anionic charge on the top Au atoms may directly contribute to the high GOx-like activity since a hydroperoxo-like species was formed during the electron transfer progress form the anionic top Au atoms to O 2 . In addition, the PtCu NPs were reported to possess ferroxidase-like activity isolated from the impact of other ions based on the Fenton-like reaction [201]. Despite the obscure mechanism, the Pt NPs (as catechol oxidase mimics) [202], Au nanorod/ Pt nanodot structures (as ferroxidase mimics) [203], Cu 2 O NPs (as cytochrome c oxidase mimics) [204] and many other metaland metal oxide-based nanozymes have broaden the way toward the prosperity of OXD mimics.

Superoxide Dismutase-Like Activity
Superoxide dismutase is a kind of metalloenzyme that mainly distributed in microorganisms, plants and animals. Oxidative stress, involving the increasing concentration of reactive oxygen species (ROS), is considered to be an important factor in aging and disease [205]. ROS refers to the reduction products of oxygen in the body, including oxygen radicals (e.g., O 2 ·− , · OH, HO 2 · ) and certain nonradical oxidizing agents (e.g., ozone, H 2 O 2 , hypochlorous acid) [206]. SOD is selected as a favorable tool to anti-oxidation and anti-aging since it could transform superoxide anion radicals into H 2 O 2 and O 2 [207]. Numerous nanomaterials have been proven as SOD mimics, such as Mn 3 O 4 [208], Au [63], MnO 2 [209], and CeO 2 [210]. The coupled electron-transfers model was once accepted as a rational mechanism to explain the SOD mimetic property of CeO 2 NPs as shown in Fig. 5a [168]. Following the oxidative half-reaction ( Fig. 5a➀-➃, same as that in Fig. 2a), a O 2 ·− molecule would bind to the reduced oxygen vacancy site (Fig. 5a➄). Then, H 2 O 2 was released with the absorption of two protons and the transfer of electron from one Ce 3+ (Fig. 5a➅). The original nanoceria state would be regenerated by repeating this reaction with a second O 2 ·− molecule (Fig. 5a➆). However, this model was questioned since Cafun et al. demonstrated the absence of spin-unpaired Ce 3+ sites in colloidal nanoceria via means of high-energy resolution hard X-ray spectroscopy [211].
Given profound consideration about the true structure and electronic characteristics of cerium oxide, Wang et al. proposed a polished catalytic cycle mechanism for nanoceria as SOD mimics [171]. The surface defect states were critical to the enzyme-like activity in this model. After the coadsorption of HO 2 · onto the surface of CeO 2 , the intermediate was formed as shown in Fig. 5b. Then, the reaction between the intermediate and another HO 2 · radicals could release H 2 O 2 and O 2 , with the nanoceria restored to the initial state.
With the assistance of rigorous density functional theory and microkinetic modeling, Guo et al. investigated the Langmuir-Hinshelwood (LH) and Eley-Rideal (ER) mechanisms to describe the SOD-like activity of Co 3 O 4 [170] and Fe 3 O 4 [212] respectively. As illustrated in Fig. 5c, the ER mechanism is more viable for Co 3 O 4 as the barriers involved through ER mechanism was lower than those along LH mechanism [170].

Others
Compared with oxidoreductive family, the reports about metal-and metal oxide-based nanomaterials with hydrolase mimetic activities are relatively rare. The peptide-functionalized monolayer protected gold clusters (Au MPCs) have been demonstrated as mimics of nuclease, esterase and silicatein [213][214][215][216]. The functional groups present on the protecting shells of Au MPCs were fundamental to their catalytic activities [217]. In addition, the CeNPs have been uncovered to show phosphatase-like property since they could cleave the phosphate ester bond of ATP, pNPP, and o-phosphol-tyrosine [218][219][220]. The key to their catalytic phosphate ester bond cleavage lied on the availability of cerium(III) sites. Dhall et al. prepared CeNPs with phosphatase and CAT-mimetic activities via the wet chemical method [147]. The kinetic studies using pNPP as the substrate indicated that their phosphatase-type catalytic mechanism followed the saturation-based kinetics with V max and K m values of 0.44 nmol min −1 and 0.74 mM, respectively. In their study, the tungstate and molybdate tend to inhibit the phosphatase mimetic activity of CeNPs owing to the interaction of anions with the CeNPs surface.

Morphology
Previous studies have demonstrated that the morphology control would affect the catalytic activity of nanozymes to a large extent [146]. Exploration on the relevance between morphology and catalytic activity mainly involved surface area, pore size and volume. Tian et al. prepared VO 2 NPs in three kinds of morphologies (fibers, sheets and rods) as POD mimics [221]. The VO 2 nanofibers performed best in the H 2 O 2 and glucose colorimetric assay due to their largest specific surface area. Singh et al. [222] compared Mn 3 O 4 NPs in cube-, polyhedron-, hexagonal plates-, lakes-and flower-like morphology (Mnf). The larger size and higher surface area seemed to create higher catalytic activity of Mnf. Moreover, the multi-enzyme property of Mnf could be ascribed to the larger pore size, which would hold the substrates and cofactor for the catalytic reactions.
The effect of surface facets has gradually become a focus in morphology control as it determines surface energy or surface reactivity [223]. Huang et al. [55] found the OXD-type activity of CeO 2 nanorods with unique {110} planes was more ingenious than that of nanopolyhedra and nanocubes. In the research of Mu et al. [224], the catalytic activities of Co 3 O 4 materials were in the order of nanoplates > nanorods > nanocubes. The difference in lowering energy barrier and electron transfer ability might be related to distinct POD-like properties of three kinds of Co 3 O 4 nanozymes. Ge et al. [67] reported that the Pd octahedrons enclosed by {111} facet structure showed lower surface energy, which were more sensitive to CAT-type property and ROS-eliminating capacity than the Pd nanocubes enclosed by {100} facet structure. As shown in Fig. 6a [225]. The binding between O 2 and Pd {100} facet (an adsorption energy of − 1.40 eV) was much stronger than that between O 2 and Pd {111} facet due to the higher adsorption energy at Pd {100} facet (Fig. 6b). Also, the activation energy of surficial O 2 dissociation for {100} facets (0.31 eV) was lower than that for the {111} facets (0.67 eV). Thus, the energetically more favorable dissociative adsorption of the O 2 molecule on the Pd {100} facet explained its higher OXD-like activity. In terms of POD capacity, the homolytic dissociation reaction on the Pd {100} facet was more feasible than on the Pd {111} facet considering the reaction energy (Fig. 6c).

Size
Generally speaking, size sheds significant influence on the properties of diverse nanomaterials [226]. In most cases, the nanozymes with smaller size tend to be more active in catalytic reactions ascribed to the larger specific surface area. For example, Xi et al. [32] reported the size-dependent POD-type properties of Pd-Ir NPs within the size range from 3.3 to 13.0 nm. With an enzyme-linked immunosorbent assay (ELISA) as a model platform, they attributed the higher catalytic properties of the smaller nanoparticles to their diffusivities and reduced steric effect. Luo et al. considered that the amount of surficial Au atoms was the key point to control the catalytic reaction rate, thus explaining the size-related GOx mimics activities of AuNPs [133]. They prepared CeO 2 NPs with SOD-and CAT-mimetic capacities in four kinds of sizes (4.5, 7.8, 23, and 28 nm) [227]. The decreased particle sizes could increase the Ce 3+ fraction along with enhancing catalytic efficiency. Interestingly, Liu et al. [228] discovered that the catalytic activity of β-Casein-AuNPs (β-casein functionalized AuNPs) was increased in the order of 4.2, 2.8, and 8.7 nm. Obviously, the smallest β-Casein-AuNPs did not possess the best POD-like activity. They deduced that the coated protein might affect the proximity between substrates and the nanozyme core, which also determined the enzyme-like property.

Surface Valence State
The controls of the surface valence state and oxygen vacancies are considered as essential factors to modulate catalytic properties. Researches have demonstrated that the surface oxidation state of nanoceria played a considerable role in tuning the enzyme-like properties of CeO 2 due to the association between Ce 3+ and oxygen vacancies. Pirmohamed et al. verified that the H 2 O 2 decomposition rate of nanoceria increased with the decreasing of Ce 3+ /Ce 4+ redox state ratios [229]. In contrast, the reduced Ce 3+ /Ce 4+ ratio was responsible for the decay of SOD mimetic capacity [230]. Besides CeO 2 nanozymes, Wang et al. reported that the POD mimicking activity in Ni-based nanozymes was associated with the oxidation state of Ni [231]. In their study, the catalytic performance of porous LaNiO 3 perovskite was about 58and 22-fold higher than that of NiO and Ni NPs, indicating the Ni oxidation state-dependent POD-like properties of Nibased nanomaterials. Moreover, they proved the significance of Ni 3+ in regulating catalytic activities via the comparison between LaNiO 3 -H 2 and LaNiO 3 nanocubes, in which the ratios of Ni 3+ were different. With tuning copper states from Cu 0 to Cu 2+ , Xi et al. found that the multi-enzyme-like activities (POD, CAT and SOD) of copper/carbon nanozymes were closely related to the Cu state [232]. Fan

Composition
The composition control of nanozymes provides possibility to tune their catalytic activity [33]. Some studies demonstrated that the catalytic performance and Raman scattering (SERS) activities of AgAu, AgPd, and AgPt NPs are more obvious than that of Ag NPs [234][235][236]. Similarly, alloying with other metals (e.g., Pd, Au, Cu, and Co) has also been regarded as feasible solutions to catalytic ability regulation of Pt NPs [237]. In fact, adjusting the proportion of components and designing metallic core/shell structurebased nanomaterials are both feasible solutions modulate the enzyme-like properties [154,238]. Liu et al. speculated that the Pt/Ru molar ratio would affect electronic variation and electronic charge transfer effects of PtRu nanoalloy, thereby tuning their POD-and OXD-like activity [239]. In their work, the enzyme-type property was enhanced in the order of Pt 40 Ru 60 , Pt, Pt 75 Ru 25 , and Pt 90 Ru 10 . He et al. reported that the change of Au/Pt molar ratio not only influenced structure of AuPt alloy NPs, but also improved the catalytic reaction rates when increasing Pt/Au ratio [85]. To investigate the metallic core/shell structure-based nanomaterials, Xia et al. adjusted the amount of Ir precursor to obtain Pd-Ir cubes with different Ir shells [240]. In this work, the Ir shells at certain thicknesses would effectively increase the surface reactivity of Pd and reduce the dissociation difficulty of H 2 O 2 molecules. Moreover, the thickness of Ir shells could enhance or weaken the ligand effect stemming from the interaction of Ir monolayer with Pd substrate, in which the Pd(100) surface with single Ir layer was more active than that with three Ir layers during the oxidation process of TMB.
Owing to the synergetic effects between ceria and heteroatoms, doping CeO 2 with suitable foreign atoms is favorable to boost the catalytic activity [241]. By replacing Ce 4+ ion in the CeO 2 lattice, the incorporation of heteroatoms tends to strengthen surface defects in the CeO 2 lattice via generating more oxygen vacancies for oxygen migration and diffusion [242,243]. Among diverse heteroatoms, the introduction of one-dimensional nanowires achieved the best catalytic activity enhancement effect [244]. Zhang et al. synthetized CeO 2 nanozymes doped with different metal elements (such as Ag, Cr, Co, Rh, Pd, Mn, and Ni) and possessed multi-enzyme-like activities, herein the Cr/CeO 2 nanozymes owned best catalytic performance. The Cr 3+ incorporation could improve surficial Ce 3+ /Ce 4+ ratio, thus reinforcing the catalytic capacity of CeO 2 NPs [245]. In addition to the types of doped atoms, the amounts are critical to regulate activity of nanozymes as well. Jampaiah et al. revealed that the catalytic efficiency toward TMB oxidation of 6% Fe 3+ -doped CeO 2 NRs was the best among the CeO 2 NRs incorporated with 3, 6, 9, and 12% Fe respectively [246]. The Raman and X-ray photoelectron spectroscopy (XPS) results indicated the higher amount of surface defects including Ce 3+ ions and oxygen vacancies in the 6% Fe 3+ -doped CeO 2 nanozymes.

Surface Modification
Surface modification ranging from functional group, inorganic ions and small molecules to macromolecules has been revealed as a promising strategy to regulate the mimetic enzyme properties of metal-and metal oxide-based nanozymes by affecting their surface chemistry [247][248][249]. For instance, ligands such as glutathione (GSH), dendrimer, DNA, and protein tend to protect metal nanoclusters from aggregation, thence reinforcing the stability, biocompatibility and catalytic activity of nanozymes [250,251]. Liu et al. reported that the catalytic efficiency of the DNAcapped iron oxide NPs as POD mimics was about tenfold higher than that of naked NPs [252]. The DNA coatings not only strengthened combining capacity with the amino groups of TMB via hydrogen bonding, but also provided the π-π stacking for nucleobase interacting with the benzene rings of TMB, which effectively enhanced the affinity of and CeO 2 , along with the carboxyl groups served as substrate binding sites, was critical to the promotional effect on the enzymatic activity. The addition of F − into nanoceria obviously caused the generation of more oxygen vacancies, facilitating electron transfer between the Ce 4+ /Ce 3+ redox couple as well as the stimulating product desorption, thereby enhancing OXD-mimetic capacity of nanoceria by fluoride capping [255].

External Triggers
(1) pH and temperature Up to date, the enzyme-like activities of numerous metaland metal oxide-based nanozymes have been verified to be sensitive to pH and temperature [17,[256][257][258]. The POD-type property of Fe@PCN-224 NPs was optimal in pH 3.5 with the temperature of 45 °C [259]. And the activity could remain 80% and 90% of the highest activity at 25 and 37 °C, respectively. Although an increasing number of novel nanomaterials have shown high enzyme-like property within a wide temperature range, the catalytic activity of nanozymes would slightly decrease when the temperature was not at optimal [260]. Liu et al. [261] found that the ROS eliminating activity of Pt NPs was strengthened with the increment of environment pH by the assistance of electron spin resonance (ESR) spectroscopy and spin traps. It has been reported that Pt NPs [261], Ag NPs [262] functioned as POD mimics in acidic conditions while exhibited CATlike activities in neutral and alkaline environment. What is more, Pt and Au NPs were demonstrated to show SOD mimetic capacity under neutral conditions [63,261]. Li et al. [167] dug into the pH-switchable enzyme-like properties of Au, Ag, Pt, and Pd nanozymes. The adsorption of H + and OH − ions on the metal surface was feasible under acidic and basic conditions, respectively. The base-like decompositions of H 2 O 2 in low-pH conditions was fundamental to the PODlike activities of Au, Ag, Pt and Pd nanozymes while their CAT-type activity was related to the acid-like decompositions of H 2 O 2 in high-pH conditions.
(2) Hydronium The catalytic activity of nanozymes could also be affected by metal ions (e.g., Fe 3+ , Hg 2+ , Ni 2+ , Cd 2+ , and Al 3+ ) and anions (e.g., S 2-, F -, Cl -, Br -, and I -) [136,263,264]. For example, heavy metal ions might inhibit catalytic activities of metal-and metal oxide-based nanozymes, which could be ascribed to the metallophilic interaction between nanozymes and heavy metal ions, including the deposition of metal ions [265], the formation of alloy on the surface of nanomaterials [266], and the leaching of surface atoms [267]. The integration between heavy metal ions and the surface ligands also affected the catalytic performance of nanocomposites by deposing of ligands or decreasing affinity toward substrate [268,269]. Han et al. conjectured that the promotional or block effects of Ca 2+ , Fe 3+ , Hg 2+ , and Mn 2+ toward the CAT-type property of Co 3 O 4 NPs were related to their influence on the electron transfer rate in Co 3 O 4 [270]. In the report of Liu et al., the S 2at low ion concentration tended to inhibit the POD-mimetic catalytic reactions of β-casein stabilized Pt NPs (CM-PtNPs) toward TMB while switch on their enzyme-like activity toward ABTS [264]. Besides, the sulfide-mediated activity switching efficiency decreased with the increment of S 2concentration. Fluorescence spectra and X-ray photoelectron spectroscopy (XPS) data revealed that the key of S 2--mediated activity switching mechanism lied in the structure change of protein molecule and ratio change of Pt 2+ /Pt 0 with the introduction of sulfide ions.
(2) Light The photothermal effect and light-induced electron transfer have been demonstrated to be involved with the photoenhanced enzyme-like activity of nanozymes [271][272][273]. With AuNPs and α-FeOOH microcrystals grown on porous carbons, Zhang et al. obtained Au/α-FeOOH-FPC catalysts with visible-light-driven enzymatic property [274]. Herein, the system temperature was raised to accelerate the process of glucose oxidation when the Au NPs converted the absorbed light energy into heat. And the generated gluconic acid could lower surrounding pH to stimulate the enzymatic reaction. Furthermore, hot electrons from plasmon-excited AuNPs promoted charge separation at the interface of Au/α-FeOOH, resulting in efficient cycling of Fe 3+ /Fe 2+ to produce Fenton reaction. The introduction of visible light has increased the POD-type activity of Fe 2 O 3 NPs by at least 1.2 times in the research of Zhu et al. [275]. They found that the light-related catalytic property of Fe 2 O 3 nanozymes was concerned with the bandgap and light absorption range, which were responsible for the barrier density generation and the light energy absorption. In addition, the influence on the enzyme mimetic properties changed according to the type of light excitation. Wang et al. discovered that the catalytic activity of Au/Si/Azo (AuNPs encapsulated and dispersed by the azobenzene-modified expanded mesoporous silica) was activated under UV illumination while inhibited under visible light [276]. The control of the host-guest interaction between Azo and cyclodextrin (CD) via the isomerization between trans and cis conformations of Azo was significant to the activity regulation by UV or visible light.

(4) Others
Nucleoside triphosphates (NTPs) including adenosine triphosphate (ATP), guanosine triphosphate (GTP), cytidine triphosphate (CTP) and uridine triphosphate (UTP) have been considered as promoters for nanozymes owing to the coupling of their hydrolysis with oxidative reaction [220]. Vallabani et al. discovered that the employment of ATP could reinforce the affinity between Fe 3 O 4 NPs and their substrate, thus maintaining the POD mimetic capacity of Fe 3 O 4 nanozymes within a wide range of pH and temperature [277]. Interestingly, Cheng et al. [197] found that the introduction of ATP might restrain the enzymetic reaction of nanoceria in prolonged reactions despite its initial enhancing effect. They attributed the inhibition to Ce-PO 4 complexes formation in the presence of ATP, which could interact with nanoceria and shield active centers. Furthermore, Jia et al. [278] reported that the antioxidants possessed inhibitory effect on the POD-type property of Co 3 O 4 NPs. The addition of gallic acid (GA), tannic acid (TA) and ascorbic acid (AA) would slow the catalytic reaction toward the TMB or OPD, among which the influence of TA was the highest because of its numerous phenolic groups.

Applications of Metal-and Metal
Oxide-Based Nanozymes

Applications in Analytical Field
As mentioned above, metal-and metal oxide-based nanozymes normally come along with unique physicochemical properties including high surface-to-volume ratio, enzymatic activity and good biocompatibility. These capabilities endow them with promising applications in target substances detection following the extensive exploration of biosensing schemes [279].
The integration of nanozymes and conventional determination technologies containing colorimetric, electrochemical, and fluorescence has gradually become optimal candidate for biological analysis. The past decade has witnessed the inclusive utilization of novel nanozyme-based sensors in detecting proteins, glucose, heavy metal ions, pathogen microorganisms and many other substances.

Heavy Metal Ions
Previous studies have illustrated that excessive heavy metal ions are one of the culprits of environmental pollution [280]. Furthermore, heavy metal ions could invade human body through water and food, resulting in permanent chronic poisoning [281]. Therefore, detecting heavy metal ions is of great significance to protect ecology and human health. Nevertheless, most analytical platforms (e.g., atomic absorption spectrometry, energy-dispersive X-ray, and inductively coupled plasma mass spectrometry) for heavy metal ion analysis relied on expensive instruments and professional technicians [282]. Nanozymes provided a potential to simultaneously improve the performance of metal ion detection with low cost. For instance, Han et al. designed a portable paper chip based on AuNPs (AuNZ-PAD) to investigate Hg 2+ in distilled and tap water samples, in which Au-Hg 2+ integration could influence enzyme-like catalytic activity of AuNPs and caused paper discoloration (Fig. 7a) [226]. This ultrasensitive AuNZ-PAD further cooperated with mobile phone camera, effectively reducing the cost of assay and simplifying the operation. Among the classical analytical assays basing nanozymes, colorimetric stood out for the operation convenience. Some references concluded that heavy metal ions might enhance or inhibit the POD-like property of nanozymes [283][284][285]. Hence, histidine(His)-Pd [268], MMoO 4 (M = Co, Ni) [286], DNA-Ag/Pt [287], MnO 2 [288] have been synthesized for Ag + [268], Cu 2+ [286], Hg 2+ [287,288] monitoring by colorimetric assay. In addition, Pb 2+ ions would accelerate the AuNPs leaching in presence of S 2 O 3 2− and lead to less oxidation of TMB, expanding the Pb 2+ determination with the assistance of nanozymes [289,290]. Xie et al. [291] fabricated a colorimetric probe by using metallic nanozyme to determine Pb 2+ . The Au@Pt NPs served as POD mimics were introduced, which could detect Pb 2+ ions in the lake water samples within a linear range from 20 to 800 nM.
As high electrolyte has an adverse effect on the catalytic performance and stability of nanozymes, analyzing heavy metal ions in seawater is much more difficult than other liquid samples such as lake water and drinking water [292]. Logan Fig. 7 a Detection progress of the Hg 2+ in distilled and tap water samples with AuNZ-PAD based on the TMB-H 2 O 2 catalytic reaction. b Principle of quantitative detection of Hg 2+ ions in seawater (3.5% NaCl) using OEG-AuNPs compared with that using bare AuNPs. c Detection principle of TAC based on the reaction between antioxidants and H 2 O 2 in the presence of Pt nanozymes as POD mimics. d Synthesis of cDNA@ Pt@P-MOF(Fe) as the signal probe for the analysis of telomerase activity. e Preparation of Ag-Au/AgCl nanohybrid with OXD-like and PODlike activity and the working mechanism of Spm detection. GCE Glassy carbon electrode, TP telomerase primer. Adapted from a Ref. [226], b Ref. [293]. c Ref. [301], d Ref. [302], e Ref. [303] with permission functionalizing AuNPs with oligo-ethylene glycol (OEG) [293]. In this proposal, OEG-AuNPs exhibited enhanced stability and weakened catalytic properties in a wide pH range under high NaCl concentration, which effectively ameliorated the poor stability of bare-AuNPs (Fig. 7b). The Hg 2+ detection limit of coastal seawater by this platform was 13 ppb in only 45 min.

Biomarkers
Biomarkers refer to biochemical indicators that mark the structure or functional changes of biosystems including organ, tissue and cell. The exploration of biomarkers is beneficial to clinical diagnosis, drug analysis and ecosystem protection. Enormous effort has been made in nanozyme-based biomarker detecting, including biological macromolecules (e.g., acid phosphatase (ACP) for prostate cancer [149]; human epidermal growth factor receptor-2 (HER2) for breast cancer [294,295]; carcinoembryonic antigen (CEA) for rectal cancer [296,297] and benzo[a] pyrene-7,8-diol 9,10-epoxide-DNA (BPDE-DNA) for woodsmoke exposure [298]) and small molecule biomarkers (e.g., sarcosine for prostate cancer [299] and uric acid [300]). Pedone et al. [301] developed a colorimetric approach to determine the total antioxidant capacity (TAC) in saliva on basis of the reaction between antioxidants and H 2 O 2 in the presence of Pt nanozymes, which was acted as POD mimics. TAC acted as an important biological indicator closely associated with oxidative stress. It reflected the total effects of enzymes and non-enzymatic analytes in the body. The combination of Pt nanozymes and · OH radical substrates allowed the detection scheme sensitive to both single electron transfer (SET) and hydrogen atom transfer (HAT) reactions (Fig. 7c).
The improvement in signal transduction rate is a breakthrough to raise the sensitivity of biomarker detection [298]. Thence, metal-and metal oxide-based nanozymes functioned as signal amplification has boosted biomarkers analysis in sundry assays involving electrochemical, fluorescent and so on [300]. Ling et al. obtained Pt@ P-MOF (Fe) nanozymes by growing ultra-small Pt nanoparticles on metalloporphyrin metal organic frameworks [302]. The novel artificial nanozymes were employed as signal probe, allosteric switch of DNA and Exo III recycling amplification in their electrochemical template for telomerase detection (Fig. 7d). The catalytic property of Pt NPs on P-MOF (Fe) could decompose H 2 O 2 , and hence strengthened the electrochemical signal. Kuo et al. [303] synthesized Ag-Au/ AgCl nanohybrid with OXD-and POD-type capacities for spermine (Spm) analysis in urine, which could act as the diagnostic indicators for liver cancer and stroke. As is shown in Fig. 7e, Spm inhibited fluorescent molecules generation of H 2 O 2 -Amplex Red (AR) system when in the presence of Ag-Au/AgCl, thereby realizing highly selective and ingenious determination of Spm.

Pathogen Microorganisms
The analysis of pathogenic microorganisms, ranging from viruses, bacteria, parasites to prions, is crucial to prevention and control of infectious diseases [304]. The nanozymes have become powerful competitors for natural enzymes in field of pathogen detection due to their low-cost (especially for foodborne bacteria), timesaving operation and sensitivity [305][306][307]. For instance, Cheng et al. employed Pd@Pt NPs as a signal amplifier in the lateral flow immunoassay (LFIA) assays for Salmonella Enteritidis (S. enteritidis) and Escherichia coli (E. coli) O157:H7 [57]. The integration of Pd@Pt NPs and smartphone-based device offered a portable platform for fast detection of foodborne pathogens. The studies involving nanozyme-based pathogen analysis in the past 5 years are listed in Table 2. All the metal-and metal oxide nanozymes mentioned in this table were functioned as POD mimics.
In contrast to POD mimics, other enzyme-like activities of nanozymes are waiting for further development in biological sensing. Yao et al. [308] designed a colorimetric immunoassay scheme to investigate Staphylococcus aureus (S. aureus) with the assistance of magnetic carbon dots (Mag-CDs) and AgNCs. AgNCs with OXD-mimicking properties could accelerate oxidatiing o-phenylenediamine (OPD) to produce yellow products. And the Mag-CDs were introduced to capture bacteria in their system. Bu et al. [309] built a point-of-care (POC) platform to analyze Salmonella sp. and E. coli O157:H7 by using MnO 2 nanoflowers with CATtype activity. Besides, MnO 2 possessed bacteria recognition ability via the binding between Con A and O-antigen on the bacterial surface.

Antibiotic
The dose control of antibiotics, which sheds significant influence on antibacterial and anti-cancer treatment, has been a hot topic in the medical field. It has been demonstrated that overdose causes serious side effects, while insufficient antibiotics are unconducive for clinical therapy [61,325]. While, the pioneering works of antibiotic determination, including liquid chromatography-mass spectrometry (LC-MS) [326], electrochemical [327], high performance liquid chromatography (HPLC) [328], etc. suffer from time consuming, high cost, complicated operations and poor sensitivity. The prosper of Au nanozymes with intrinsic POD-like activity provided possibility to tune the functionalization of existing methods in analyzing multiple antibiotics (e.g., doxycycline [325], kanamycin [61], tetracycline [329]). Kong et al. [330] designed a novel photoelectrochemistry (PEC) biosensor for bleomycin (BLM) detection, which was natural antibiotics for Hodgkin's disease, cervical cancer therapy. The biosensor reached a detection limit to 0.18 nM in which Ag/ZnMOF nanozymes acted as a signal amplifier and Au NPs/tungsten sulfide nanorod array (Au/WS 2 ) photoelectrode used as a PEC matrix (Fig. 8a). When the Au/WS 2 photoelectrode generated PEC signals under light, the Ag/ZnMOF nanozymes with mimetic POD properties reduced the background signal via the catalyzing reaction between H 2 O 2 and 3,3-diaminobenzidine (DAB), thus greatly improving the sensitivity and specificity of BLM analysis.

Antioxidant
Antioxidants, substance to scavenge ROS or free radicals, could prevent human body from cell apoptosis and nerve damage induced by oxidative stress [331]. Nevertheless, inappropriate supplementation of antioxidants may result in diseases and increase risk of death. Therefore, quantitatively analyzing antioxidants is of great significance. The nanozyme-related antioxidant detection is based on the inhibition of antioxidants on the nanozymes' catalytic activities [260,278]. Following the evolvement of nanozymes and biosensing technology, the sensitive colorimetric determination for antioxidants has been extensively discussed, including ascorbic acid (AA, based on CoMn/NF@C [332], Pt/CeO 2 [333], Fe 3 O 4 / CoFe-LDH [158], Mn-CDs [334], etc.,), GSH (based on SPB-MnO 2 [335], Mn 3 O 4 [336], Ir [337], V 2 O 5 [338], etc.,) and l-Cysteine (l-Cys, based on Fe 3 O 4 [339], etc.,). Most existing biosensors were designed for specific antioxidant analysis, while approaches for multiple antioxidants detection are scarce. Huang et al. [340] designed a MnO 2 nanosheets triggered colorimetric sensor array for simultaneous discrimination of UA, GSH, AA, l-Cys, and melatonin (Mel) in serum (Fig. 8b) [340]. The inhibitory effects on the catalytic performance of MnO 2 nanosheets vary according to the kind of antioxidants, resulting in different degrees of TMB oxidation and generating multicolors. Since the absorbance values at 370, 650, and 450 nm would change, the corresponding absorbance values A 370 , A 450 , and A 650 were employed as three crossreactive sensing elements in the visual colorimetric sensor array. The detection results revealed that the sensor could precisely and rapidly identify the five antioxidants and their mixture at a low concentration.

Other Substances
As a byproduct of respiratory metabolism, H 2 O 2 is one of most common molecule in biological tissues [341]. When the concentration is at an abnormal status, H 2 O 2 would cause damage to health and might induce oxidative stress related diseases [342]. Besides, hydrogen peroxide was widely used in biopharmaceuticals, environmental management, food manufacturing and some other fields due to its strong oxidant properties [343]. A bunch of methods have been designed to monitor H 2 O 2 in various matrices considering its significant roles in biological metabolisms and broad utilization in industrial production [341,344]. Among these assays, colorimetry and electrochemistry have gradually became main technologies for H 2 O 2 determination owing to low cost, high   Adapted from a Ref. [330], b Ref. [340], c Ref. [349], d Ref. [362] with permission sensitivity and selectivity [345]. Up to now, a variety of metaland metal oxide-based nanozymes (e.g., CuO-g-C 3 N 4 [346], MnO 2 [347], V 2 O 5 -CeO 2 [348]) have been exploited for electrochemical analysis. Zhang et al. fixed ZIF-8 on graphene oxide (ZIF-8-rGO) and further synthesized AuPt/ZIF-8-rGO with POD-like activity to practically track H 2 O 2 in human serum samples (Fig. 8c) [349]. The AuPt/ZIF-8-rGO-based electrochemical scheme showed remarkable electroanalysis performance along with excellent sensitivity and selectivity. This work reached the detection limit of 19 nM (S/N = 3), which obtained the lowest detection limit compared with previously reported electrochemical sensors. The color change of peroxidase substrate (e.g., TMB) triggered by hydrogen peroxide is the foundation in colorimetric detection of H 2 O 2 . Diverse POD mimics (e.g., Cu 2 O-Au [350], Fe-N-C [351], Cu(II)-coated Fe 3 O 4 [352], PtCu [353], V 2 O 5 [341], C-dots/Fe 3 O 4 [130], and Rh [354,355]) have been developed to manufacture colorimetric sensors. To our knowledge, the currently lowest detection limit of H 2 O 2 based on colorimetry is 0.0625 µM reported by Tripathi et al. [356], and the palladium nanoclusters (Pd NCs) were designed by biological methods firstly in their study, in which Pd NCs were served as POD mimics.

(2) Glucose
Glucose is an indispensable nutrient for metabolism in organisms. The heat released during its oxidation reaction is a considerable energy source required by life events [357]. However, a surfeit of glucose might cause various diseases, including hyperlipidemia, arteriosclerosis, hypertension, diabetes and so on [358]. The concentration of glucose in blood or urine is a crucial indicator of physical condition [357,359]. By combining the catalytic performance of glucose oxidase (GOD) and nanozymes with POD-type activity (e.g., Zn-CuO [331], Au@Ag [360], MoO 3 /C [331], Ag [361], and Pt [135]), numerous optical technologies have described for glucose analysis in serum [135], beverage [279], and urine [331,361] samples. Blood pretreatment and serum extraction were often demanded in conventional blood glucose detecting programs. To simplify determination steps, Lee et al. [362] designed a protocol that could directly monitor glucose in whole blood and avert pretreatment. They prepared a GOD-conjugated graphene oxide/MnO 2 (GOD-GO/ MnO 2 ) sensor platform for quantitatively analyzing blood glucose with a detection limit of 3.1 mg dL −1 (Fig. 8d). The results indicated that this colorimetric sensor possessed clinical potential for blood glucose monitoring of diabetic patients.

Application in Antibacterial
The lack of non-antibiotic therapies and multiple drug resistance caused by bacteria diseases become one of the most serious problem, which threatens human health [363][364][365]. In the process of developing optimal antibacterial strategies, nanometallic materials have been discovered to exert antimicrobial nature [366,367]. In addition, POD and OXD mimics were verified to catalyze producing harmful ROS, ranging from H 2 O 2 , superoxide, hydroxyl radicals to other small reactive molecules [27]. Hence, metal-and metal oxide-based nanozymes (e.g., V 2 O 5 [368], CuO [369], CeO 2 [370], Au/MOF [371], and Tb 4 O 7 [372]) have been gradually regarded as promising bactericides. For example, Fe 3 O 4 NPs with POD-like properties could decompose H 2 O 2 to generate toxic · OH for bacterial infections treatment [373]. Evidence has emerged that enzyme mimic abilities of nanomaterials are closely associated with their composition and structure, which would affect antibacterial capacity [374]. Xi et al. [232] designed two types of copper/carbon nanozymes including two Cu states (Cu 0 and Cu 2+ ). The copper/carbon nanozymes displayed multi-enzyme activities and antibacterial mechanism dependent on Cu states. In the study, Xi et al. concluded that hollow carbon spheres (HCSs) modified with CuO (CuO-HCSs) nanozymes could induce Gram-negative bacteria death (E. coli and P. aeruginosa) when releasing Cu 2+ . While the key of Cu-HCSs nanozymes to resist Gram-positive (Salmonella typhimurium, S. typhimurium) and Gram-negative bacteria (E. coli and P. aeruginosa) was based on POD-type activity, which was responsible for ROS generation (Fig. 9a).
The pH-dependent catalytic activity of nanozymes has been demonstrated that would limit their antimicrobial application under neutral pH, and was beneficial to grow bacteria like Escherichia coli, Staphylococcus aureus and so on [375,376]. Fortunately, ATP served as modulators has been reported to improve the POD-like property of nanozymes, and it could interact with iron ions to produce · OH under neutral pH [128,377]. Therefore, Vallabani et al. [378]     Adapted from a Ref. [232], b Ref. [378], c Ref. [379], d Ref. [382], e Ref. [369] with permission showed that Fe 3 O 4 NPs exhibited superior antibacterial performance against E. coli and Bacillus subtilis (B. subtilis, gram positive) in presence of H 2 O 2 under a neutral pH environment with the assistance of ATP (Fig. 9b). Chishti et al. discovered that fluorite-structured CeO 2 nanocrystals with ~ 23.04% Ce 3+ had recyclable POD-like activity [379]. Mechanism investigation indicated that the reduction of substrate affinity caused by ATP is the key to improve the low enzyme-like activity of nanoyzymes in a neutral environment (pH 7.4), further strengthening the sterilization sequel against both gram-positive (S. aureus) and gram-negative (E. coli) bacteria (Fig. 9c).
Besides optimizing the catalytic capacity, applying external triggers to control their antibacterial activity is essential to develop nanozyme-based antibacterial agents. Otherwise, the sustained action of nanozymes might induce bacteria to yield drug resistance. Karim et al. firstly reported that light could act as an external spark to control nanomaterials' catalysis [369]. A highly basic tertiary amine could produce visible light to excite CuO NRs. The increment of light intensity enhanced the affinity of CuO NRs and H 2 O 2 , thereby improving the POD-like activity and antimicrobial properties (Fig. 9e). Results showed that CuO NRs catalyzed H 2 O 2 under visible light irradiation to output · OH with 20 times higher than that under no light.
The exaltation of H 2 O 2 sterilization efficiency has become an issue of increasing concern as H 2 O 2 is a crucial and easily available ROS. Although numerous studies were devoted to this issue, applications of these systems were still restricted by the health hazard from high concentration of H 2 O 2 (greatly higher than biologically relevant concentration) [380,381]. Wang et al. integrated Au NPS with graphitized carbon nitride (g-C 3 N 4 ) to synthesize non-toxic ultra-thin g-C 3 N 4 @ AuNPs (CNA) nanozymes with high POD catalytic activity [382]. CNA nanozymes were firstly reported to possess excellent bactericidal properties under biosafety level of H 2 O 2 , and could efficiently decompose DR-biofilms to inhibit bacteria growth (Fig. 9d). In vitro experiments proved that CNA system provided significant advantages in preventing bacterial infections and accelerating wound healing.

Application in Relieving Inflammation
Inflammation, including acute and chronic inflammation, is regarded as a precursor to certain diseases [383]. An obvious feature of inflammatory tissue is the increasing of reactive oxygen or nitrogen species (RONS) content [384,385]. Owing to the ROS scavenging ability, favorable stability in extreme environments and excellent biocompatibility, nanozymes have been indicated to be potential substitutes for broad-spectrum antioxidants in terms of inflammation treatment [386][387][388]. So far, a variety of metal-based and metal oxide-based nanozymes (such as Mn 3 O 4 [56], CeO 2 [389], Pt/CeO 2 [390], and Cu-TCPP MOF [391]) have been reported for anti-inflammatory therapy. The main challenge to realize clinical transformation of nanozymes is to enhance the ROS eliminating performance and simplify nanomaterials' structure. Liu et al. synthesized ultra-small Cu 5.4 O nanoparticles (Cu 5.4 O USNPs) with mimic enzyme properties of CAT, SOD and GPx (Fig. 10a1) [392]. The ultra-micro size of Cu 5.4 O USNPs ensured their biocompatibility via the rapid removal of nanomaterials in the kidney (Fig. 10a2). Cu 5.4 O USNPs were confirmed to protect healthy cells from ROS at extremely low dosage. They also showed the promoting effect on the treatment of acute kidney injury, acute liver injury and wound healing in animal experiments. Wu et al. introduced RuO 2 -PVP NPs to set up a therapeutic nanoplatform for inflammation alleviation and neuroprotection [393]. In this work, RuO 2 -PVP NPs with multi-enzymatic properties effectively protected lipid, DNA and protein from oxidative stress in parallel with the broad-spectrum ROS elimination performance against inflammation and Parkinson's disease in vivo. Yao et al. [56] expanded the use of Mn SOD in anti-inflammatory. Their team demonstrated the multiple enzyme mimics activities of Mn 3 O 4 NPs, which could scavenge superoxide free radicals, H 2 O 2 and hydroxyl free radicals. In in-vitro experiments, the ROS-eliminating level of Mn 3 O 4 NPs was much higher than traditional CeO 2 nanozymes. The experimental results indicated the obvious prospects of Mn-based nanozymes in treating and preventing ROS-mediated neuroinflammation.
The combination of nanozymes and other kinds of antiinflammatory agents could bring a turning point for refractory inflammatory diseases. For example, the lack of targeting strategies and the risk of side effects with increasing dosage increased the difficulty in treating inflammatory bowel disease (IBD) [397]. By growing CeO 2 NPs in situ on montmorillonite (MMT) sheets, Zhao et al. designed CeO 2 @ MMT nanozymes with SOD-type, CAT-type and · OH scavenging properties to directly target the inflammatory colon for IBD therapy [394]. In this system, MMT alleviated the potential nanotoxicity of CeO 2 NPs via reducing their Adapted from a Ref. [392], b Ref. [394], c [395] d Ref. [396], e Ref [52] with permission systemic absorption, which in turn endowed MMT sheets with ROS eliminating activity. Animal experiments have also proved that the nanozyme-based drugs were suitable for oral delivery and stable in gastrointestinal environment (Fig. 10b). CeO 2 @MMT exhibited good targeting for colon disease sites, effectively treating IBD induced by dextran sulfate sodium in mice model. Although carbon monoxide (CO) gas therapy was recently revealed as a novel antiinflammatory strategy, it still suffered from the low tissue specificity and troublesome amount control [398][399][400]. By Despite the tremendous attention that paid to nanozymerelated anti-inflammatory therapies, there are still few reports about metal-and metal oxide-based nanozymes with reactive nitrogen species (RNS) scavenging ability. RNS including nitric oxide ( · NO), nitrogen dioxide ( · NO 2 ) and peroxynitrite ( · ONOO − ) etc. are a major culprit in aggravating neuroinflammation induced by traumatic brain injury (TBI) [401]. Miao et al. prepared polyethylene glycol (PEG) coated (PEGylated) ultra-small rhodium nanodots (Rh-PEG NDs) showing excellent multi-enzyme-like activity and high photothermal conversion efficiency [396]. On the one hand, Rh-PEG NDs possessed similar RONS removal capacity as natural CAT, thereby alleviating the inflammation of colon disease. On the other hand, they could be used for photoacoustic imaging and photothermal therapy (Fig. 10d). Mu et al. [52] prepared PtPdMo trimetallic (triM) nanozymes for neuroinflammation treatment through multi-enzyme mimetics reaction-based RONS elimination. In addition, triM nanozymes displayed highly catalytic selectivity in neutral environments, which provided possible application of nanozymes in brain science (Fig. 10e). Zhang et al. doped Cr 3+ ions into CeO 2 to prepare Cr/CeO 2 nanozymes by increasing Ce 3+ states [402]. The higher Ce 3+ /Ce 4+ ratio contributed to strengthening enzyme-like activity of nanozymes with 3-5 times higher than undoped CeO 2 . The Cr/CeO 2 -based catalytic patch has been demonstrated as a promising choice for non-invasive TBI treatment and neuroinflammation relief owing to the satisfactory RONS (including · OH, ONOO − and H 2 O 2 ) scavenging ability.

Application in Cancer Treatment
According to the latest global cancer statistics from the World Health Organization/International Cancer Center team, cancer is expected to become the main cause of death in countries around the world in twenty-first century [403]. Compared with traditional tumor treatment methods (surgery, chemotherapy, radiotherapy, etc.), external minimally invasive or non-invasive strategies containing photodynamic therapy (PDT), chemodynamic therapy (CDT), sonodynamic therapy (SDT), immunotherapy etc. show a favorable development prospect due to their accurate tumor specificity, space/time controllability and biosafety [404,405]. However, the complex tumor microenvironment (TME) limited the therapeutic effects of many methods. TME not only refers to structure, function and metabolism of tumor tissue, but is also related to the internal environment of tumor cell (nuclear and cytoplasm) possessing the characteristics of hypoxia, acidity, glutathione and overexpression of H 2 O 2 [406,407]. The intrinsic catalytic activity enables nanozymes to regulate TME via changing RONS content or eliminating hypoxia [43,[408][409][410]. The biological safety, photothermal performance and some other physicochemical properties of nanozymes also indicated their potential in cancer therapy [411]. Given these reasons, nanozymes have been regarded as the prospective standalone agents or synergist for the progress of tumor treatment [43].

Photodynamic Therapy
PDT relied on ROS generated by photosensitizers (PSs) under light irradiation to induce cancer cell apoptosis [412]. Nevertheless, most PSs still face disadvantages of low selectivity, poor water solubility and high selfdestruction [413]. In order to reinforce the stability of loading PSs, various nanozymes such as MnO 2 [414], Pt [51] and so on were utilized. In the research of Xu et al. [415], Pt/C nanozymes not only served as chlorin e6 (Ce6) nanocarriers, but also promoted the conversion of H 2 O 2 and O 2 into ROS with anti-tumor property (Fig. 11a). They compared the nanozymes with various structures and found that HCS@Pt NPs (Pt NPs decorated with hollow carbon spheres) showed favorable POD-and OXD-like activity, thereby further firming the therapeutic efficacy of PDT for cancer.
In addition to PSs transportation, studies have also confirmed that tumor hypoxia would weaken PDT efficiency [417]. Hence, nanozymes (e.g., Pt [418], Mn 3 O 4 [419]) as CAT mimics were employed to consume intratumoral H 2 O 2 and generate oxygen in parallel with photosensitizer carriage. However, tumor hypoxia was difficult to be continuously suppressed due to the respiration of intratumoral mitochondria [420]. Yang et al. integrated IR780 PSs into mesoporous silica NPs (MSNs) and then covered with Mn 3 O 4 NPs to produce Mn 3 O 4 @MSNs@IR780 nanocomposites [419]. Mn 3 O 4 nanozymes that accumulated in the tumor sites could decompose H 2 O 2 and trigger switch to release IR780, which specifically targeted to mitochondria and produced ROS to inhibit cancer cells respiration after destroying mitochondria. In vitro experiments proved that oxygen supplementation and mitochondrial destruction were vital to PDT enhancement. Hu et al. [416] employed exogenous oxygen-generating materials (CaO 2 NPs) to alleviate tumor hypoxia. In this report, MnO 2 nanozymes with CATmimicking activity not only catalyzed CaO 2 NPs to generate O 2 , but also allowed image-guided PDT as a promising MR T1 nanoprobe (Fig. 11b).

Chemodynamic Therapy
Chemodynamic therapy generates · OH by catalyzing intratumoral H 2 O 2 via Fenton or Fenton-like reactions, thereby killing tumor cells [421]. Nanozymes with POD-like activity (e.g., Fe 3 O 4 NPs [422], AFeNPs [423]) have been recognized as Fenton reaction catalysts for CDT in acidic environments. Since existing reports revealed the pH-dependence of CDT, the pH-independent nanozymes (e.g., Fe/Al-GNE [424], Au 2 Pt [142]) were designed to provide efficient Fenton reactions in neutral TME. What's worse, high concentration of GSH and low H 2 O 2 in TME have also been demonstrated to restricted CDT effect [425]. Therefore, conquering the above-mentioned TME is a challenge to optimize CDT reaction efficiency.
Fu et al. synthesized CoO@AuPt nanocatalyst with high biocompatibility and stability under physiological environment, which regulated responsive CDT by lowering pH, increasing H 2 O 2 level and consuming GSH content [426]. In the work, CoO template could degrade and generate Co 2+ in acidic and high-level H 2 O 2 environment, which was further acted as a useful Fenton-like reagent. The released Au/ Pt nanozymes as multi-enzyme (GPx, CAT, POD, and GOx) mimics were responsible for decreasing GSH concentration and catalyzing H 2 O 2 into O 2 and · OH (Fig. 12a). Moreover, the nanosatellites consumed intratumoral glucose to generate numerous H 2 O 2 and induced starvation therapy, thereby enhancing the effect of CDT.
Another challenge to achieve augmented CDT is to increase the generation rate of · OH. Ma et al. [131] introduced microwave (MW) as an external stimulus to regulate CDT and realize controllable tumor therapy, named as microwave enhancing dynamic-therapy (MEDT). By coupling gold nanoclusters (BSA-Au NCs) with Fe-metal organic frameworks (MIL-101(Fe)), IL@MIL-101@ BSA-AuNCs NPs were prepared after loading methylimidazolium hexafluorophosphate (IL) on MIL-101(Fe) NPs. Under microwave irradiation, MIL-101(Fe) enzymes owned MEDT by catalyzing H 2 O 2 to produce toxic · OH in tumor. The dynamic distribution of MIL-101 (Fe) NPs in vivo and tumor site could be real-time monitored by magnetic resonance imaging (MRI) and fluorescence imaging (FI) (Fig. 12b).

Sonodynamic Therapy
PDT is commonly suitable for relatively small superficial tumors due to the limited depth of light penetration through tissues [427]. In contrast, ultrasound (US) owns a higher tissue penetration depth than light waves. Thus, US-triggered sonodynamic therapy is promising to treat deep or large tumors by activating sonosensitizers to generate ROS [428,429]. Resemble to PSs in PDT, the performance of sonosensitizers plays a fundamental role in SDT [430]. The past 5 years witnessed the development of novel marvelous sonosensitizers [429,431]. The stability and catalytic activity allowed some metal-and metal oxide-based nanozymes to function as sonosensitizers and Fenton reagents simultaneously to achieve CDT-enhanced SDT. For instance, Wang et al. designed polyethylene glycol (PEG)-modified nanozymes with ultrafine rod-like structure, named PEG-TiO 1+x NRs for tumor ablation [432]. Compared with traditional inorganic sonosensitizers, the sensitivity of PEG-TiO 1+x NRs was more prominent due to hypoxic structure. Furthermore, PEG-TiO 1+x NRs with HRP-type activity showed Fenton-like catalytic property. As SDT reagent possessing CDT function, the intravenously injected PEG-TiO 1+x NRs were significantly more effective in inhibiting tumors than traditional TiO 2 NPs under US irradiation (Fig. 13a) Fig. 13 a Schematic illustration of the working mechanism of TiO 1 + x NRs with HRP-like activity for SDT/CDT-combined tumor therapy. b Preparation procedure and working mechanism of PtCu 3 -PEG nanocages with HRP-and GPx-type property for PA/CT dual-modal imagingguided CDT-enhanced SDT. Adapted from a Ref. [432], b Ref. [433] with permission toxicity and inhibited tumor growth by generating ROS by decomposing H 2 O 2 into · OH and depleting GSH under US, in which PtCu 3 could obviously optimize the reaction environment of CDT. Meanwhile, owing to high light absorption and strong X-ray attenuation in near-infrared region, PtCu 3 could be employed for photoacoustic (PA)/computed tomography (CT) imaging-guided CDT-enhanced SDT (Fig. 13b).

Photothermal Therapy
Materials with high photothermal conversion efficiency are exploited in photothermal therapy (PTT), which could convert light energy into heat energy for the death of cancer cells under external light irradiating [434]. Numerous metal-based and metal oxide-based nanozymes (e.g., MnO 2 [435], Ru-Te [436], Ru@CeO 2 [437]) have been reported as photothermal agents (PTAs). In these studies, nanozymes ameliorated PTT efficacy due to their enzyme-like abilities and other superior properties at the same time. Wang et al. [435] synthesized 2D MnO 2 nanosheets (M-NSs) with controllable protein orientation through a wet chemical method, and then functionalized M-NSs via a sonochemical proposal. As is shown in Fig. 14a, the M-NS served as GOx mimics with highly dispersion and stability, which finally realized starvation therapy by consuming glucose of tumor cells. The nanozymes also presented remarkable photothermal conversion efficiency and PA imaging performance under nearinfrared (NIR) irradiation, thereby achieving PA imagingguided synergistic cancer treatment of starvation therapy and PTT. However, the effect of PTT is stinted by light penetration depth and thermal damage to healthy tissue induced by overexposure [438]. Therefore, a series of studies tried to combine PTT with other treatment methods to achieve synergistic therapy [439]. For example, Au 2 Pt nanozymes as POD and CAT mimics with potent photothermal performance were reported for PDT/CDT/PTT synergistic cancer therapeutics [142]. AgPd NPs with POD-like activity could improve photothermal conversion efficiency, and have been proved to be acted as carriers for chemotherapeutic drugs transmission during a weakly acidic environment (pH 5.5), thus achieving ROS/PTT/chemotherapy guided by NIR laser [440]. Pt-CuS Janus nanozymes were adopted in synergistically enhanced SDT and PTT [441]. In this system, Pt-CuS Janus hollow structure was used as sonosensitizers carrier, showing photothermal conversion capacity under laser irradiation, and could decompose endogenous H 2 O 2 expeditiously. The Pt NPs [442] with CAT-mimicking capacity and Ru-Te hollow nanorods [436] with OXD, POD-, CAT-and SOD-type activity both acted as carriers and relieved TME hypoxia to enhance cancer PDT/PTT effect. Different from most nanozyme-based synergistic therapy, Yang et al. [443] covered Pt-carbon integrated nanozymes as PSs via one-step reduction. Under NIR light lasering, the nanozymes provided brilliant photosensitivity and photothermal effect. And the PDT reinforcement was relied on the CAT-like catalysis activity. In vivo experiments revealed that Pt-carbon nanozymes inhibited mice colon cancer reaching an 90% efficiency (Fig. 14b). Li et al. [411] prepared the H 2 O 2 -responsive PtFe@Fe 3 O 4 , which possessed POD-like activity, CAT-type property and exceptional photothermal performance under acidic TME environment. Experimental results indicated that tumor catalytic therapy based on PtFe@Fe 3 O 4 nanozymes obtained a 99.8% anti-tumor rate for deep pancreatic cancer when cooperating with photothermal therapy What is more, the electron transfer process between PtFe nanorods, Fe 3 O 4 NPs and H 2 O 2 molecules was also firstly described in their study (Fig. 14c).

Immunotherapy
Cancer immunotherapies, regarded as promising strategies for tumor therapy, utilize the immune system of patients to treat cancer [444], and might include cytokine therapy, tumor vaccines, immune checkpoint blockade (ICB) therapy, adoptive cell therapy and so on [445]. Studies have demonstrated that the modulation of TME is conducive to tumor immunotherapy [43]. Yang et al. [446] designed a polyethylene glycol (PEG)-modified hollow manganese dioxide (H-MnO 2 ) nanoshells to load photodynamic agent Ce6 and chemotherapy drug doxorubicin (DOX), forming H-MnO 2 -PEG/C&D complex for cancer combination immunotherapy. The H-MnO 2 could alleviate tumor hypoxia via catalytically decomposing hydrogen peroxide to generate O 2 . A series of immunological responses were discovered with synergistic treatment of H-MnO 2 -PEG/C&D and Chemo-PDT, resulting remarkable decreasing in the secretion of IL-10 (predominant cytokine secreted by M2 macrophages) and the increment in the secretion of IL-12 (predominant cytokine secreted by M1 macrophages). Moreover, the introduction of 1 3 anti-PD-L1 checkpoint blockade showed further enhanced therapeutic efficacy for tumor with by improving TNF-α.
Moreover, it has been reported that tumor-associated macrophages (TAMs) are critical to tumor growth and metastasis, thereby playing an important role in the cancer immunotherapy [447]. Regulating TME could facilitate macrophage polarization from M2 to M1 since the tumor hypoxia is associated with macrophage recruitment and polarization [448]. Xu et al. [449] loaded TGF-β inhibitor (TI) to the PEGylated iron manganese silicate nanoparticles to prepare IMSN-PEG-TI nanoplatform for tumor immunotherapy (Fig. 14d). In this system, IMSN nanozymes with POD-and CAT-like property could decompose H 2 O 2 into · OH and O 2 to kill tumor cells and overcome tumor hypoxia in respective. The interaction of IMSN and TI effectively regulated the tumor immune microenvironment, leading to elevated ratio of M1 to M2 macrophages, CD4 + T to T reg cells, and CD8 + T to T reg cells. Furthermore, the enhanced macrophages polarization would in turn induce the reproduction of H 2 O 2 , thus promoting enzymatic properties of IMSN nanozymes.

S t a r v a t i o n T h e r a p y
Fig. 14 a Working principle of M-NSs as GOx mimics and PTAs with effective PA imaging performance for the synergistic starvationenhanced PTT guided by PA imaging. b Synthesis procedure and the working mechanism of Pt-carbon-integrated nanozymes for synergistic PDT/PTT cancer therapy. c Working mechanism of PtFe@Fe 3 O 4 with POD-, CAT-like activity and excellent photothermal effect under acidic TME environment for tumor catalytic therapy combined with PTT. d Illustration of the cancer immunotherapy using the IMSN-PEG-TI nanoplatform. Adapted from a Ref. [435], b Ref. [443], c Ref. [411] with permission

Conclusion
The prosperity of nanotechnology and biology created a series of novel artificial enzymes. As promising natural enzymes mimics, nanozymes have demonstrated remarkable performance in clinical medicine, biopharmaceuticals, environmental monitoring and many other fields. In this review, we meticulously elaborated the intrinsic activity and catalytic mechanism of the classical metal-and metal oxide-based nanozymes, including monometal-, metal alloy-, metal oxide-, metallic core/shell nanostructure-based and hybrid nanomaterials. The recent research progress of metal-and metal oxide-based nanozymes in analysis, antibacterial, relieving inflammation, and cancer therapy was also involved. Although nanozymes have been revealed to overcome many limitations of natural enzymes such as low stability, complicated preparation and expensive storage, there are still severe challenges for future researches. (1) Compared with most natural enzymes, metal-and metal oxide-based nanozymes seem to lack the substrate specificity. Even though researchers have discovered amounts of inner and external factors that influencing enzymatic properties, the precise control of catalytic performance, especially for the nanozymes with multi-enzyme-like activities, still has a long way to go. (2) The exploration of the internal catalytic mechanism is fundamental for understanding and mastering the catalytic reaction of nanozymes. In contrast to the synthesis and employment of novel nanomaterials, studies that involved the deep comprehension of working mechanism are relatively rare. What's worse, the advanced strategies dedicated to mechanism clarification are also limited. (3) The POD mimics have become an issue of extensive concern in most nanozyme-related applications, especially in the field of analysis and detection. While other component of oxidoreductase family have also been proved to possess unsubstituted function in many circumstances. Therefore, the spread utilization of SOD, CAT, OXD mimics are yet to be developed. (4) Most previous biosensors based on nanozymes could only detect one or two substances. The schemes for simultaneous discrimination and quantification of multiple (≥ 3) substances with high sensitivity are required to be further investigated and simplified. (5) Considering the cost control in large-scale preparation, seeking alternatives for noble metal nanozymes has gradually received increasing attention. Besides, the reduction of their content in nanoalloys and nanocomposites while guaranteeing the performance is also worth more efforts. (6) The longterm in vivo toxicity of nanozymes still remains a challenge for their clinical employment. Although a large amount of studies have involved the discussion about the biocompatibility, the systematic mechanisms of toxicity and corresponding solutions are in urgent need. 81571729 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http:// creat iveco mmons. org/ licen ses/ by/4. 0/.