Introduction

Metal nanomaterials including tiny nanoparticles [1], well-defined nanocrystals [2], and core–shell nanostructures [3] have encouraged extensive research interests in mimicking intrinsic functions of native enzymes, because of their excellent catalytic performance, good stability as well as facile synthesis process [4, 5]. Up to now, considerable research activities have been devoted to boost the enzyme mimicking activities of metal nanomaterials, mainly relying on various modulators such as secondary metallic elements toward electronic and geometric structures [6, 7], organic ligands or inorganic ions toward surface properties [8,9,10,11]. For example, to overcome the pH lacunae, adenosine triphosphate disodium salt (ATP) has been employed to enhance the enzymatic activity of citrate-coated Fe3O4 NPs, which acts as a synergistic agent to accelerate the OH radical production [12]. Moreover, fluoride ion (F−) has been reported to significantly enhance the oxidase-like activity of nanoceria, which inverses the surface charges and increases the content of oxygen vacancies [13]. These studies provide effective approaches to adjust the physicochemical properties of metal nanozymes.

Reduced coenzyme I (NADH) and reduced coenzyme II (NADPH) are ubiquitous electron sources involved in cellular energy metabolism and biosynthetic reaction [14, 15]. Inspired by nature, oxidation-redox mechanisms between NAD(P)H and metal ions have attracted considerable attention in the fields of energy chemistry, biosensing, green synthesis and nanotechnology [16, 17]. For example, Fe3+ ions can be easily reduced into Fe2+ by NADH under mild conditions since the association of Fe3+ with the adenine ring facilitates its of NADH [18]. In the presence of NAD(P)H, the reduction of Au3+ to Au+ occurs, thereby contributing to the catalyzed growth of Au nanoparticles [19]. Recently, NADH was utilized in one-pot synthesis of ultrasmall AuPd nanoclusters, showing good performance in the determination of acid phosphatase [20]. Hence, it is promising to develop desirable metal nanozymes under the assistance of native coenzymes.

Iodide ion (I−) acts as a particular part in the fields of biology, food and environment. Various diseases originate from altered concentration of I−, including goiter, cretinism, hypothyroidism and hyperthyroidism [21, 22]. Nevertheless, a few reported nanozymes (Au, Pt) exhibit stronger affinity toward S2− than I− [23,24,25,26]. Hence, it is still a great challenge to explore iodide-responsive nanozymes with desirable selectivity. Recently, He et al. utilized the I−-induced inhibition of peroxidase-like activity of chitosan-coated Pd nanozyme, to establish a sensitive assay for I− ions [27]. Besides, I− ions have also been reported to exhibit strong affinity to Ag-based nanomaterials [28]. This inspires us to develop new iodide-selective nanozyme based on the electronic adjustment between Ag and Pd.

In this study, NADPH was chosen as both the reductant and the stabilizing ligand to construct highly active peroxidase mimetics in one-step (illustrated in Scheme 1). Both the reduction process and the interaction mechanism between NADPH and metal precursors were deeply investigated through time-dependent UV–vis spectra, ITC, TEM, XRD, XPS, ICP-MS and enzyme kinetics. AgPd nanozymes exhibit superior peroxidase mimicking activities over their monometallic counterparts because of the synergy between Pd and Ag. Owing to the affinity between I− ions and AgPd NCs, the catalytic activity of nanozyme significantly decreases in the presence of I−. Intriguingly, AgPd nanozyme possesses a desirable selectivity toward I− with respect to S2−.

Scheme 1
scheme 1

The preparation of monometallic and bimetallic NCs through the reduction and stabilization of NADPH

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Experimental section

Chemicals

β-Nicotinamide adenine dinucleotide phosphate disodium salt (NADP, 98%) was obtained from Shanghai Yuanye Bio-Technology Co., Ltd (http://www.shyuanye.com/). Reduced coenzyme II tetrasodium salt (NADPH, 98%) was obtained from Coolaber Co., Ltd (http://www.coolaber.com/). NaCl (99.99%), Na2SO4 (99%) and AgNO3 (99%) were obtained from Aladdin Industrial Corporation (https://aladdin.company.lookchem.cn/). K2Pd(NO2)4 (99%), H2O2 (ACS, 29–32 wt %) and citric acid (99.5%) were obtained from Alfa Aesar (Shanghai, www.alfa.com/zh-cn/). CH3COONa (99%) was obtained from Sangon Biotech (Shanghai, https://www.sangon.com/) Co., Ltd. KI (98.5%) was obtained from Tianjin Guangfu Technology Development Co., Ltd (http://www.guangfubiaowu.com/). Sodium citrate tribasic dehydrate (99%) and 3,3′,5,5′-tetramethylbenzidine (TMB, 98%) were obtained from Tianjin Heowns Biochemical Technology Co., Ltd (Tianjin, www.heowns.cn/). Na2CO3 (99.8%) was obtained from Tianjin third Chemical Reagent Factory (http://m.sp7804794.zjbiz.net/). NaNO3 (99%) was purchased from Tianjin Damao Chemical Reagent Factory (http://dmchem.b2bvip.com/). KF (99%) was purchased from Tianjin Fengchuan Chemical Reagent Technology Co., Ltd. Na2SO3 was purchased from Tianjin Guangfu Fine Chemical Research Institute (https://tjguangfu.company.lookchem.cn/). Na2S was purchased from J&K Scientific Ltd (https://jkchemical.company.lookchem.cn/). Human serum was purchased from EpureSino.

Characterizations

UV–vis absorption spectra were recorded on a Varian Cary 300 spectrophotometer, and all the samples were measured in a 1 cm path quartz cuvette. Transmission electron microscopy (TEM) images were collected on JEM-2100F (JEOL, Japan). Typically, a 10 μL droplet of the sample was placed on a carbon-coated grid, and stood for 10 min before removing the liquid with filter paper. The binding energies of the surface species of NCs were measured by X-ray photoelectron spectroscopy (XPS, PHI5000 Versaprobe), and all the specimens were prepared on silicon wafers under the protection of N2. Inductively coupled plasma-mass spectrometry (Thermo Fisher Scientific, USA: ICPA-MS-Qc) was adopted to determine the elemental contents for AgPd NCs. To determine atomic ratios, the purified samples were dissolved in aqua regia at 40 °C for 2 h. X-ray diffraction (XRD) patterns were recorded on X-ray diffractometer (D/Max-2500, Rigaku, Japan) equipped with Cu–Ka radiation (λ = 0.154178 nm). All the samples were prepared on glass slides under N2 atmosphere. Isothermal titration calorimetry (ITC) experiments were conducted by using MicroCal VP-ITC instrument with the sample cell of 1.45 mL. Under stirring at 307 rpm at 25 °C, 10 μL aqueous solution of 20 mM K2Pd(NO2)4 or AgNO3 was injected into 50 μM NADP solution for total 28 injections with the interval of 360 s. Zeta potential of as-prepared NCs was collected on Zetasizer Nano ZS90 instrument (Malvern Panalytical Ltd.).

Synthesis of AgPd NCs by using NADPH

NADPH was employed as both stabilizing and reducing agents to prepare AgPd NCs. 300 μM of AgNO3, K2Pd(NO2)4 or two precursors with different ratios of [K2Pd(NO2)4]/[AgNO3], was mixed with 600 μM NADPH in 5 mL water. The mixture was incubated at 25 °C for different reaction time, and then purified by ultrafiltration tube (molecular weight cutoff: 10 kDa) at the centrifugal speed of 7500 r/min for 0.5 h. The concentrated NCs on the membrane were further washed more than three times by using water. The collected NCs was diluted to 5 mL by water.

Evaluation of peroxidase mimicking activity

To start the reaction, 1.2 µM (total concentration of metal precursors) of NCs was added into the reaction system containing 0.125 mM TMB and 125 mM H2O2 in 10 mM citrate buffer (pH 4.0). The kinetic curves were monitored at 652 nm. The initial velocities (ν) were obtained using Eqs. (1) and (2):

$$\mathop C\nolimits_{p} = \frac{{\mathop A\nolimits_{652} }}{\varepsilon L}$$
(1)
$$\nu = \frac{{\mathop {{\text{d}}C}\nolimits_{p} }}{{{\text{d}}t}}$$
(2)

Cp The concentration of oxidized product; ε The extinction coefficient of oxidized product, which equals to 3.9 × 104 M−1 cm−1; L The optical path length of quartz cell, which equals to 1 cm.

The enzyme parameters were obtained via Eq. (3):

$$\frac{1}{v} = \frac{{\mathop K\nolimits_{{\text{m}}} }}{{\mathop \nu \nolimits_{{{\text{max}}}} \left[ {\text{S}} \right]}} + \frac{{1}}{{\mathop \nu \nolimits_{{{\text{max}}}} }}$$
(3)

[S] The concentration of substrate; Km The Michaelis–Menten constant; Vmax The maximal velocity.

Stability of Ag1Pd1 NCs

The stability of NCs was assessed as follows: 1.2 µM of Ag1Pd1 NCs was added into 10 mM citrate buffer (pH 4.0) that contains 0.05 mM TMB and 63.7 mM H2O2, respectively. After 25 min-reaction, the absorption intensity of the product was read at the wavelength of 652 nm by UV spectrophotometer. To start the reaction again, concentrated solution of TMB and H2O2 were added into the above reaction system. After 25 min, the increment in absorption intensity (ΔA) was calculated. Eight cycles were repeated as the above procedure.

Determination of I− ions in aqueous solution

To study the influence of I−, the reaction system was comprised of 125 mM H2O2, 0.125 mM TMB, 1.2 µM of Ag1Pd1 NCs (total concentration of metal precursors) as well as different concentration of KI in 10 mM citrate buffer (pH 4.0). Firstly, 10 μL solution of Ag1Pd1 was added into 3 mL citrate buffer (pH 4.0). Then the stock solution of KI was added into the above mixture. Finally, 15 μL of TMB solution (25 mM in ethanol) and 38.5 μL of H2O2 solution (30 wt% aqueous solution) were added successively to start the reaction. The A652 signals were recorded after reaction for 30 min at 25 °C. The LOD was calculated according to 3σ/slope, where σ and slope represent the standard deviation of the blank sample and the slope of the standard curve, respectively [29]. In control experiments, other anions including CO32−, Cl−, NO3−, SO42−, SO32−, F−, Br−, CH3COO−, HCO3− and S2− were individually investigated under the same conditions.

Determination of I− ions in cooking salt and human serum

To determine the concentration of I− in cooking salt, the reaction system contains 1.2 mg/L cooking salt, 125 mM H2O2, 0.125 mM TMB and 1.2 µM of Ag1Pd1 NCs (total concentration of metal precursors) in 10 mM citrate buffer (pH 4.0). Then different concentrations of I− were spiked into the above solution. After incubation for 30 min at 25 °C, the A652 signals were recorded by UV–vis absorption spectroscopy.

Human serum was diluted by 1000-times in 10 mM citrate buffer (pH 4.0) containing 125 mM H2O2, 0.125 mM TMB and 1.2 µM of Ag1Pd1 NCs (total concentration of metal precursors). Next, different concentrations of I− were spiked into the above solution. After incubation for 30 min at 25 °C, the A652 signals were recorded by UV–vis absorption spectroscopy.

Results and discussion

Synergy between Ag and Pd in the growth of AgPd NCs

The growth process of NCs was monitored by time-dependent UV spectroscopy. The typical band at 340 nm corresponding to NADPH quickly decreases upon addition of K2Pd(NO2)4 (Fig. S1a in supplementary material), which could be ascribed to the loss of electrons of NADPH. As shown in Fig. 1a, an inducible peak at 470 nm occurs upon addition of NADPH into K2Pd(NO2)4. The absorption intensity of this band gradually increases from 2 to 19 h. As a control, the mixture of K2Pd(NO2)4 and NADP does not show any absorption peak in the region of 400 ~ 800 nm (Fig. S2). Ligand-stabilized Pd NCs have been widely reported to display multiple UV–vis absorption bands [30, 31]. Therefore, such UV–vis absorption features (Fig. 1a) might be assigned to the charge transfer in NADPH-generated Pd NCs [32]. As shown in Fig. 1b, the reduced Pd species form highly disperse NCs with the average size of 1.8 nm.

Figure 1
figure 1

UV–vis spectra for the reduction process of a K2Pd(NO2)4, c AgNO3 and e the mixture of K2Pd(NO2)4 and AgNO3. TEM images of b Pd NCs, d Ag NPs and f Ag1Pd1 NCs

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Upon addition of AgNO3, the oxidation of NADPH undergoes a slower process with respect to K2Pd(NO2)4 (Fig. S1b). According to the SPR band (~ 410 nm), Ag NPs gradually grows within 36 h, to generate the final diameter of 15.3 nm (Fig. 1c and 1d). ITC experiments show that NADP can associate with both K2Pd(NO2)4 and AgNO3, with the binding constants of 525 M−1 and 3380 M−1, respectively (Fig. S3). So, it is suggested that NADP potentially protects Pd NCs/Ag NPs against aggregation.

In the presence of two precursors ([K2Pd(NO2)4]/[AgNO3] = 1), the absorption intensity in long wavelength significantly increases within 2 h, and then it changes slightly with time evolution (Fig. 1e). It is worth noting that the growth of AgPd involves much faster kinetics relative to their monometallic counterparts. Importantly, no characteristic band was observed for either Ag or Pd, which is indicative of a fused structure.

As the ratio of [K2Pd(NO2)4]/[AgNO3] is less than 1/2, the SPR band of Ag is distinguishable (Fig. S4). Moreover, the distinctive band related to Pd NCs appears at the [K2Pd(NO2)4]/[AgNO3] of 2, and its intensity greatly enhances with further increasing the [K2Pd(NO2)4]/[AgNO3]. Ag1Pd1 and Ag2Pd1 NCs show the mean size of 2.0 and 2.2 nm, respectively (Fig. 1f and Fig. S5). The increasing order of the average size is Pd < Ag1Pd1 < Ag2Pd1 < Ag, showing the same order with the relative Ag content.

According to XRD results, the crystal structures for Ag, Pd, Ag1Pd1 and Ag2Pd1 were analyzed, respectively. The peaks at 37.95°, 43.64°, 64.32° and 77.36° match with the (111), (200), (220) and (311) planes of Ag NPs [33] (Fig. S6). The typical peaks at 40.54° and 47.03° correspond to the (111) and (200) planes of Pd [34]. For Ag1Pd1 NCs, the peaks observed at 39.51° and 45.04° locate between the peaks for Pd NCs and Ag NPs. Hence, it is demonstrated that either Ag1Pd1 or Ag2Pd1 exists in a fused state.

The valence states of different NPs and NCs were further studied by XPS. As shown in Fig. 2a, the binding energy (B.E.) of Ag 3d for Ag NPs locates at 367.6 eV with no obvious Ag+ species, demonstrating that Ag species on the surface of Ag NPs exist in an atomic state. For Pd NCs, the B.E. value of Pd 3d locates at 338.2 eV, concomitant with a shoulder peak around 335.7 eV (Fig. 2b). It indicates that Pd NCs are mainly comprised of positively charged Pd2+ species, accompanied by a small proportion of low coordination number Pd species. The deconstruction spectra show two typical bands with the B.E. values of 338.2 eV (77.4%) and 335.8 eV (22.6%), which are in accordance with Pd2+ and Pd0, respectively [35].

Figure 2
figure 2

XPS analysis of AgPd NCs: a Ag 3d of the Ag NPs, Ag2Pd1 and Ag1Pd1 NCs. Pd 3d of b Pd NCs, c Ag2Pd1 NCs and d Ag1Pd1 NCs. Black and red represent the raw and the fitted curves. Blue and pink lines illustrate the Pd2+ and the Pd0 components

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The B.E. of Ag species for Ag2Pd1 NCs shows a slight downshift relative to that of Ag NPs. It is noticeable that the B.E. of Pd 3d for Ag2Pd1 NCs shows an obvious negative shift to 335.2 eV with respect to Pd NCs (Fig. 2c), which demonstrates that Pd species dominantly exist as atomic or low coordinated states. The Ag2Pd1 consists of 76.3% metallic Pd0 atoms as well as 23.7% high-valent species in total Pd species. It is demonstrated that the percentage of zero-valent species is greatly improved for Pd by employing Ag as the secondary metal.

In comparison with monometallic Ag NPs, the B.E. value of Ag 3d was positively shifted by 0.5 eV for Ag1Pd1 NCs. The observed peak shift was primarily ascribed to the electron donation from Ag to Pd due to different electronegativity between Pd (2.20) and Ag (1.93). The opposite shifts of B.E. values indicate the existence of strong interaction between Ag and Pd. It is also suggested that more Ag atoms are imposed by surrounding Pd atoms in Ag1Pd1 as compared to Ag2Pd1. Moreover, atomic Pd0 species account for 65.1% in total Pd for Ag1Pd1 NCs (Fig. 2d). Therefore, the electronic status of either Ag or Pd was altered in AgPd NCs.

Synergistic effect in AgPd nanozyme for peroxidase mimicking activity

The peroxidase mimicking activities of different NCs and NPs were evaluated in order to study the synergy between Ag and Pd. As shown in Fig. 3a, Ag1Pd1 NCs display considerable activity in the oxidation of TMB by H2O2. For comparison, the mixture of NADP and metal precursors shows extremely low activity under the same condition. In the case of monometallic catalysts, Pd NCs exhibit the reaction rate of 0.48 µM/min, whereas Ag NPs show poor activity (Fig. 3b). Notably, the enzymatic activities are greatly enhanced through increasing the relative proportion of Ag from 10 to 50%. With 50% addition of Ag, Ag1Pd1 NCs display the optimal peroxidase-like activity (v = 1.8 µM/min). The catalytic activity further decreases as the fraction of Ag increases to 90%.

Figure 3
figure 3

a Reaction kinetics of TMB-H2O2 reaction (pH 4.0) in the presence of Ag1Pd1 NCs, the mixture of Ag+ -Pd2+-NADP as well as the mixture of K2Pd(NO2)4 and AgNO3. b The initial velocities (v) of TMB-H2O2 system (pH 4.0) in the presence of two precursors at different ratios reduced by NADPH

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Michaelis–Menten curves were received for Ag1Pd1, Ag2Pd1 and Pd NCs within the appropriate ranges of two substrates (Fig. 4, Fig. S7 and S8). The data fit well to the Lineweaver–Burk equation, giving the affinities of 147 mM for H2O2 and 0.32 mM for TMB at 25 °C. As listed in Table 1, the Km value toward H2O2 are increases in the order of Ag1Pd1 < Ag2Pd1 < Pd, whereas the Km toward TMB decreases in this order. The maximum velocities decrease in the following order: Ag1Pd1 > Ag2Pd1 > Pd. Hence, it can be concluded that the peroxidase mimicking activities predominantly depend on the intrinsic properties of catalysts in the activation of H2O2.

Figure 4
figure 4

Enzymatic parameters evaluated for Ag1Pd1 NCs in TMB-H2O2 reaction at 25 °C: a Plot of v with the concentration of H2O2. c Plot of v with the concentration of TMB. b and d were double-reciprocal plots of (a) and (c), respectively

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Table 1 Enzymatic parameters of nanozymes in this study
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Regarding the stability of Ag1Pd1 NCs, the increment in absorbance decreases by approximately 23% after 8 cycles in TMB-H2O2 system (Fig. S9). The electrostatic attraction of oxTMB on the negatively charged Ag1Pd1 NCs might have influence on their catalytic activities. Overall, Ag1Pd1 NCs exhibit good stability in TMB-H2O2 reaction.

According to ICP-MS, the atomic ratios of [Pd]/[Ag] are 0.62 and 1.18 for Ag2Pd1 and Ag1Pd1, respectively. Ag2Pd1 NCs contains 76% Pd0 in total Pd species, while Ag1Pd1 NCs has 65% Pd0 in total Pd. It means that Ag1Pd1 has a higher proportion of metallic Pd0 species than that of Ag2Pd1. So, the peroxidase-like activities are closely related to the amount of metallic Pd0 species.

Further, the synthesis of NCs was optimized by varying the concentration and the reduction time of NADPH. Firstly, the effect of NADPH concentration was investigated at a constant [NADPH]/[precursor] of 2. The peroxidase-like activity of Ag1Pd1 significantly grows until it reaches to a plateau with 600 µM NADPH (Fig. S10a). Next, the influence of reduction time was also considered. According to the catalytic performance, Ag1Pd1 achieves a steady reaction rate after 2 h-reduction (Fig. S10b). The most active Ag1Pd1 nanozyme was synthesized with 600 µM NADPH through 2 h-reduction. It has the optimal pH of 4 (Fig. S11).

Effects of anions on the peroxidase-like activities of Ag1Pd1 nanozyme

The potential interactions between metal and anions play a crucial role on catalysis, environment, medicine, and material science. Thus, the influence of various anions on the activity of AgPd NCs was studied. Comparative studies were performed between I− and S2−. As Ag NPs have low enzymatic activity, UV–vis titrations were performed to compare the affinity of Ag NPs with I− and S2−. As a result, both I− and S2− have strong interactions with Ag NPs (Fig. S12).

For comparison, addition of 200 nM I− inhibits the intrinsic activities by approximately 57%, 72% and 94% for Pd, Ag2Pd1 and Ag1Pd1 NCs, respectively (Fig. S13a). The adsorption of nucleophilic I− on the surface of NCs blocks the active sites of NCs, which results in the considerable inhibition of the intrinsic activity. As compared to I−, S2− causes slight decrease of their activities at the same concentration (Fig. S13b). As described above, Pd NCs exhibit higher affinity toward I− against S2−, which mainly originate from the interactions between metal and anions. Such iodide-affinity was greatly enhanced through the adjustment of Pd species by secondary Ag, e.g., Ag1Pd1 displays the most sensitive response to I−.

TEM, XPS and zeta potential were adopted to characterize the morphology, surface charge and electronic property of Ag1Pd1 NCs upon addition of I− ions. The NCs are highly dispersive without aggregation, and the zeta potential slightly decreases after I− added (Fig. S14). Negative shifts of B.E. values were observed for both Ag 3d and Pd 3d after Ag1Pd1 NCs were incubated with I− (Fig. 5), corresponding well to the previously reported changes for Ag NPs [33]. Therefore, it is reasonable to deduce that the adsorption of I− changes the electronic properties of NC surface owing to the metal-iodide bonding [36]. Combining with enzyme kinetics and ICP-MS, Ag1Pd1 has a higher proportion of metallic Pd0 species than that of Ag2Pd1. So, it is suggested that the peroxidase-like activities are closely related to the content of metallic Pd0 species. Moreover, the binding affinity between Pd and I− mainly contributes to the inhibition of the activities of AgPd NCs. Therefore, it can be deduced that the fraction of active Pd0 species plays a predominant role in the peroxidase-like activity along with the sensitivity to I−.

Figure 5
figure 5

XPS spectra of Ag 3d a and Pd 3d, b for Ag1Pd1 before and after incubation with I−

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Furthermore, Ag1Pd1 NCs were employed to quantitatively determine the concentration of I− ions in aqueous solution. As shown in Fig. 6a, a color progression from dark blue to colorless was observed with a gradual increase of I− concentration from 0 to 180 nM. The activities of Ag1Pd1 NCs, represented by the absorbance at 652 nm, show a decreasing tendency as the concentration of I− increases (Fig. 6b). The linear relationship lies in the range from 0.5 to 180 nM, giving the limit of detection (LOD) of 1.5 nM (S/N = 3). At the upper concentration of this linear range (180 nM), Ag1Pd1 NCs almost lose the peroxidase-like activity, which implies the efficient coverage of active sites by I− ions. A recent study reported a nanozyme-based assay for I− ions by employing the peroxidase-like activity of Pd NPs [27]. In this assay, monometallic Pd nanozyme lost less than 50% of its activity in the detection range. Regarding I− ion-responsive materials in studies (listed in Table S1), Ag1Pd1 NCs possess higher sensitivity relative to other Au-, Ag- and Cu-based nanomaterials.

Figure 6
figure 6

a Photographs of the color progression in Ag1Pd1-TMB-H2O2 reaction system at pH 4.0 with different concentration of I−. b The relationship between the A652 signals and the concentration of I−. c The A652 signals in the TMB-H2O2 reaction system catalyzed by Ag1Pd1 in the presence of various anions

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Next, the effects of various anions on the activity of Ag1Pd1 NCs were assessed by CO32−, Cl−, NO3−, SO42−, SO32−, F−, CH3COO−, HCO3− and S2−, respectively. In comparison with I−, other anions cause insignificant changes of the activities at the same concentration (Fig. 6c). Nevertheless, Br− ions have considerable influence on the activities of NCs (Fig. S15). Therefore, the potential interference should be considered for practical applications.

To further assess the practical application of this assay, the determination of I− was done in the samples that contains cooking salt and human serum, respectively. As listed in Table 2, the recovery rates were calculated to be 99.9% for 30 nM, 95.8% for 80 nM and 99.0% for 120 nM spiked I− ions in cooking salt. For diluted human serum, the recovery rates were determined to be 92.4% for 25 nM, 107.0% for 50 nM, 102.8% for 75 nM, and 92.2% for 100 nM I− (Table 3). It can be seen that our developed AgPd nanozyme has potential prospect in practical applications. Iodide is not only essential to daily nutrition for human, but also widely used in organic synthesis and nuclear areas. Thus, the sensitive and selective response of Ag1Pd1 nanozyme should provide guidance for developing I− sensors as well as adsorption materials. Metal nanozymes have been reported to display extensive applications in biosensing, immunoassay, diagnostics, therapy and so on [37,38,39]. Future studies will focus on the applicability of NADPH-generated AgPd nanozyme in other fields.

Table 2 Recovery experiments in cooking salt
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Table 3 Recovery experiments in diluted human serum
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Conclusion

NADPH was adopted as a bifunctional template for one-pot synthesis of AgPd NCs, showing peroxidase mimicking activity. Synergistic effects between Ag and Pd exist in both nanocluster growth and enzymatic activity. The growth of AgPd NCs undergoes faster kinetics in comparison with the formation of monometallic Pd NCs or Ag NPs. The electron transfer from Ag to Pd contributes to the marked enhancement of metallic Pd0 species in NCs. Ag1Pd1 NCs (~ 2.0 nm) possess the affinity of 147 mM toward H2O2 and 0.32 mM toward TMB. Interestingly, I− significantly inhibits the peroxidase-like activity of Ag1Pd1 NCs, accompanied by a desirable selectivity toward I− against S2−. This study not only paves a promising pathway to design biomimetic nanocatalysts, but also provides guidance for developing iodide-responsive nanomaterials and analytical methods for iodide ions.