Peroxidase-like Activity Of Gold Nanoparticles Research Articles

Target-triggered dual signal amplification based on HCR-enhanced nanozyme activity for the sensitive visual detection of Escherichia coli.

The detection of foodborne pathogens is crucial for food hygiene regulation and disease diagnosis. Colorimetry has become one of the main analytical methods in studying foodborne pathogens due to its advantages of visualization, low cost, simple operation, and no complex instrument. However, the low sensitivity limits its applications in early identification and on-site detection for trace analytes. In order to overcome such a limitation, herein we propose a joint strategy featuring dual signal amplification based on the hybridization chain reaction (HCR) and DNA-enhanced peroxidase-like activity of gold nanoparticles (AuNPs) for the sensitive visual detection of Escherichia coli. Target bacteria bound specifically to the aptamer domain in the capture hairpin probe, exposing the trigger domain for HCR and forming the extended double-stranded DNA (dsDNA) structures. The peroxidase-like catalytic capacity of AuNPs can be enhanced significantly by dsDNAs with the sticky ends of dsDNAs being adsorbed on AuNPs and the rigidity of dsDNAs causing the spatial regulation of AuNP concentration. The intensity of the enhancement was linearly related to the number of target bacteria. With the above strategy, the detection limit of our colorimetric method for Escherichia coli was down to 28 CFU mL-1 within a short analytical time (50 min). This study provides a new perspective for the sensitive and visual detection of early bacterial contamination in foods.

Enhanced peroxidase-like activity of gold nanoparticles loaded on carbon coated magnetic nanoparticles for colorimetric detection of mercury (II)

Mercury is a toxic heavy metal that has debilitating effects on human health. Anthropogenic and natural processes are responsible for the availability of mercury in the environment that necessitates periodic monitoring. Most of the techniques commonly employed for analysis of mercury, however, are expensive and require highly trained personnel. Here we report on a cheap colorimetric method using gold nanoparticles loaded carbon covered magnetic nanoparticles (Fe3O4@C@AuNPs) as nanozymes. The particles were characterized by Fourier translation infrared spectroscopy (FT-IR), X-ray powder diffraction (XRD), scanning electron microscopy (SEM), and transmission electron microscopy (TEM). The AuNPs establish specific interactions with mercury (II) and the peroxidase-like activity of Fe3O4@C@AuNPs using 3, 3, 5, 5′-tetramethylbenzidine (TMB) as a substrate was enhanced on addition of mercury (II). Our results show that the absorption signal of oxidized TMB increased linearly with concentrations of mercury (II) for a concentration range of 0.001–25 µM. The limit of detection (LOD) was found to be 0.0435 µM (43.5 nM). The feasibility of the method in real sample application was tested by analyzing mercury (II) ions in tap water. From our findings, we anticipate that our method can be applied for analysis of mercury (II) in various environmental samples.

On-site colorimetric detection of Salmonella typhimurium

Rapid qualitative and quantitative detection of Salmonella typhimurium (S. typhimurium) takes an important role in ensuring food safety. Herein, a colorimetric assay aptasensor for S. typhimurium utilizing intrinsic peroxidase-like activity of gold nanoparticles embedded spherical covalent organic framework and the affinity and specificity of S. typhimurium-aptamer has been explored. This aptasensor can capture the S. typhimurium via the selective binding effect of aptamer, and the catalytically active sites were shielded. As a result, the colorimetric signals of the 3,3′,5,5′-tetramethylbenzidine-H2O2 system were turned off. Under optimum conditions, the aptasensor gave a linear response over the range of 10 to 107 CFU/mL for S. typhimurium. The detection limit of 7 CFU/mL was obtained within 45 min and was effectively applied to detect S. typhimurium in milk and lake water samples with recoveries in the range from 96.4 to 101.0%. More importantly, combined with a self-developed smartphone-based image analysis system, the proposed aptasensor can be used for point-of-care testing applications.

A novel gold nanoparticle-based colorimetric assay for highly sensitive detection of ascorbic acid

In this study, a novel approach for colorimetric detection of ascorbic acid (AA) was proposed based on the peroxidase-like activity of gold nanoparticles, which were synthesized using Oldenlandia corymbosa leaves extract (OC-AuNPs). The OC-AuNPs synthesis was optimized at 0.5 mM Au3+, a reaction time of 20 min, and a temperature of 60 °C. The results showed that the biogenic OC-AuNPs presented as highly-efficient peroxidase mimics for catalyzing H2O2 to oxidize 3,3′,5,5′-tetramethylbenzidine forming a typical blue product with the characteristic absorption at 654 nm. The developed colorimetric method exhibited high sensitivity for AA with a detection limit of 0.26 μM in the linear range of 0 – 100 μM. The proposed assay can be serviced as a sensitive biosensor for the determination of AA in aqueous solutions due to its simple detection procedure, fast operation, and high sensitivity.

A colorimetric aptasensing assay with adjustable color mutation points for threshold-readout detection of carcinoembryonic antigen

Direct readout of the clinically significant threshold for the disease judgment is convenient for point-of-care disease screening. Here, we designed a colorimetric aptasensing platform with adjustable color mutation points for threshold-readout detection of carcinoembryonic antigen (CEA) on the test strip. This platform involves a capture module made of a magnetic bead-supported sandwich aptamer, a signal transduction module based on a hybridization chain reaction (HCR) system with tunable mercury ion (Hg2+) binding capacity, and a colorimetric readout based on the Hg2+-regulated peroxidase-like activity of gold nanoparticles (AuNPs). Notably, the tunable Hg2+ binding capacity of the HCR allows the adjustment of the color mutation point over a range of CEA concentrations from 4 to 25 ng mL−1. This platform also exhibits a sensitive detection performance with detection limit of 0.19 ng mL−1 and a reliable clinical diagnostic capability with sensitivity of 100% and specificity of 96.667%. Since the targeting aptamer can be easily changed, this platform also offers the possibility for threshold-readout detection of other disease markers (e.g., prostate-specific antigen (cutoff value of 4 ng mL−1) and alpha-fetoprotein (15 ng mL−1)). Therefore, this platform provides a convenient and straightforward tool for point-of-care screening of disease markers.

Simultaneous quantitative analysis of K+ and Tl+ in serum and drinking water based on UV-Vis spectra and chemometrics.

The simultaneous detection of K+ and Tl+ can serve as a toxicological diagnostic tool for thallium poisoning. Colorimetric-reaction-based nanoprobes have emerged as promising sensors for the rapid and ultrasensitive detection of molecular species in simple systems. However, the development of viable screening tools for multicomponent analysis in complex systems remains challenging owing to interference from coexisting materials in the media. Herein, a simple chemical sensor array based on the peroxidase-like activity of gold nanoparticles modified with single-stranded DNA (AuNPs-ssDNA) and chemometrics was developed for the simultaneous detection of K+ and Tl+ in aqueous solutions and serum. The use of a K+ adapter conferred high selectivity to the developed method. Optimized AuNPs-ssDNAs were used to construct a sensor array, which together with chemometrics provided fingerprints that can facilitate the simultaneous analysis of multiple components. The developed colorimetric reaction in combination with the chemometrics assay was directly used as a biosensor array, which exhibited detection limits of 107.33nM for K+ and 19.26nM for Tl+. The developed method could potentially serve as a diagnostic technique for investigating thallium poisoning and toxicology.

Colorimetric Sensing of Methyl Parathion Using Nanozymatic Activity of Gold Nanoparticles

In this study, we have used new approach for detection of the methyl parathion residues i.e., by inhibiting half of the peroxidase-like activity of gold nanoparticles. We have investigated the peroxidase like activity of gold nanoparticles by colorimetric assay and optimized the pH, temperature, incubation time and different concentrations of H2O2 by using TMB as substrate as well as organophosphates effect on their enzyme mimetic activity. Kinetic study of gold nanoparticles has shown better catalytic activity than horseradish peroxidase at pH 3.5. The peroxidase substrate TMB (3,3,5,5-tetramethylbenzidine) can be oxidized by H2O2 by the enzymatic action of the gold nanoparticles resulting in a blue-coloured product, oxidized TMB. The principle involved behind the inhibition of enzymatic activity of nanozyme is due to hindrance of electron transfer mechanism TMB-H2O2-Au NPs system by the methyl parathion. The peroxidase activity is selectively reduced with increasing methyl parathion concentration. This sensing method has lowest limit of detection of 78.95 nM. This study can be used for development of sensitive and cost effective technique for sensing of harmful pesticides.

Enhancing the peroxidase-like activity and stability of gold nanoparticles by coating a partial iron phosphate shell.

Using citrate-capped gold nanoparticles (AuNPs) as peroxidase-mimicking enzymes to design biosensors is hindered by their low catalytic activity and poor colloidal stability, resulting in limited sensitivity and large variations. Herein, the growth of a partial iron phosphate (FeP) shell with Fe2+ ions on citrate-capped AuNPs boosted the activity of the AuNPs by up to 20-fold. The FeP-enhanced activity was demonstrated on AuNPs of different sizes, and gold nanostars. When the FeP layer is thick enough to block the access to the Au/FeP interface, the activity was inhibited. Capping the remaining Au surface by thiol also inhibited the activity, suggesting that faster reactions occurred at the interfaces of Au/FeP. Moreover, a FeP shell can stabilize AuNPs against freezing and a high NaCl concentration of 1 M. Sensitive detection of Fe2+ was achieved with a detection limit of 0.41 μM, while no other tested transition metal phosphates enhanced the peroxidase-like activity of AuNPs.

Colorimetric Sensing of Dopamine Based on Peroxidase-Like Activity of Gold Nanoparticles

Development of a simple and label-free colorimetric sensing assay for dopamine is urgent, because of the important roles dopamine plays in many physiological processes. Herein, a label-free colorimetric method for determination of dopamine has been proposed based on the enhancement of the peroxidase-like activity of bare gold nanoparticles (AuNPs). The assay is based on specific interactions between protonated dopamine and gold atoms through the catechol group, which induces aggregation of AuNPs and significantly enhances their peroxidase-like activity. The catalytic activity is evaluated by chromogenic reaction of 3,3',5,5'-tetramethylbenzidine with H2O2. The mechanism for enhancement of catalytic activity is discussed. The detection limit of dopamine was 25 nM by virtue of UV-Vis spectroscopy, and 0.21 μM by naked eye. The whole detection procedure can be completed in 15 min. The proposed sensor is simple, fast, cost-effective and sensitive. These advantages make this sensor a powerful tool for the determination of dopamine in water samples.

Detection of Malachite Green using a colorimetric aptasensor based on the inhibition of the peroxidase-like activity of gold nanoparticles by cetyltrimethylammonium ions.

A specific and sensitive colorimetric aptasensor is described for the determination of Malachite Green (MG). It is exploiting the inhibition of the peroxidase-like activity of gold nanoparticles (AuNPs). The AuNPs act as enzyme mimics that catalyze the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) by H2O2 to yield a dark blue solution. The catalytic activity is inhibited by hexadecyl trimethyl ammonium ion, specifically by cetyltrimethylammonium bromide (CTAB), which causes the aggregation of AuNPs. If a (negatively charged) RNA-aptamer against MG is added, it binds to the positively charged CTAB and prevents aggregation. This enhances the enzyme mimicking activity of the AuNPs and leads to the formation of a dark blue solution. However, in the presence of MG, the aptamer binds to MG, and leads to the aggregation of AuNPs again. The aggregated AuNPs possess a light blue color. A colorimetric method (best performed at 650nm) was work out that can detect MG in a concentration range from 10 to 500nmolL-1. The detection limit based on 3σ/k criterion is 1.8nmolL-1. The assay is highly specific and accurate. Recoveries from spiked real samples (aquaculture water) ranged from 80% to 120%. Graphical abstract Based on the inhibition of cetyltrimethyal ammonium ion and the enhancement of RNA-aptamer, the differences of the peroxidase-like activities of AuNPs can be greatly enlarged with and without MG, by which a colorimetric aptasensor can be constructed for the detection of Malachite Green (MG).

Peroxidase-Like Activity of Gold Nanoparticles and Their Gold Staining Enhanced ELISA Application

We found that the peroxidase-like activity of gold nanoparticles (GNPs) followed the Michaelis-Menten kinetic model and was dependent on environmental pH and temperature, which was very similar to natural Horseradish Peroxidase (HRP). However, unlike HRP, which needs a lower H2O2 concentration with a very narrow range to reach a maximum reaction rate and avoid enzyme poisoning, GNPs have very high activity, even at an H2O2 concentration two orders of magnitude higher than HRP. It was demonstrated that H2O2 treatment could enhance the peroxidase-like activity of GNPs, resulting thus in the activity increase in a circular catalytic reaction by the reduplicative use of GNPs. It was also found that the peroxidase-like activity of GNPs responded sensitively to nanoparticle size and surface modifications. When used in an immunoassay, GNPs were generally conjugated with antibody and blocked with hydrophilic macromolecules to construct a nanoprobe. This strongly reduced the peroxidase-like activity and detection sensitivity of GNPs, therefore, restricting their use as peroxidase mimetics. We presented a novel strategy that combined the nanoprobes with gold staining to expose fresh catalytic gold surfaces and obtained a great increase in detection sensitivity.

Unveiling the role of ATP in amplification of intrinsic peroxidase-like activity of gold nanoparticles.

Peroxidase enzyme-like activity of gold nanoparticles (AuNPs) is currently being investigated for the potential application in the several realms of biomedicines. However, little is explored about the peroxidase activity of AuNPs decorated with different surface charges. It is well-documented that the catalytic activity and the interaction with mammalian cells are significantly different among AuNPs carrying different surface charges. We have recently reported that ATP enhances the peroxidase-like activity of AuNPs and iron oxide nanoparticles. However, a comprehensive and systematic study to reveal the role of surface charge on nanoparticles peroxidase-like activity has not been studied. In this work, we have shown that AuNPs coated with PEG (PEG AuNPs), citrate (citrate AuNPs) or CTAB (CTAB AuNPs) exhibit varying peroxidase-like activity and the boosting effect imparted by ATP was also different. We found that the peroxidase-like activity of PEG AuNPs and citrate AuNPs is dependent on hydroxyl radical formation, whereas CTAB AuNPs did not show any significant activity under the same experimental conditions. We also studied the boosting effect of ATP on the peroxidase-like activity of PEG and citrate AuNPs. Although the use of ATP resulted in enhanced peroxidase-like activity; however, contrary to the expectation, it did not facilitate the enhanced production of hydroxyl radical. In further studies, we found that the likely mechanism of boosting effect by ATP is the stabilization of oxidized TMB after peroxidase reaction. ATP imparts stabilization to the oxidized TMB produced due to PEG AuNPs, citrate AuNPs as well as HRP.

Lab-on-nanoparticle as a multidimensional device for colorimetric discrimination of proteins

We report on a method for sensitive detection and discrimination of multiple proteins. The peroxidase-like activity of gold nanoparticles (AuNPs) is employed to catalyze the oxidation of 3′,3′,5′,5′-tetramethylbenzidine (TMB) in the presence of H2O2. Depending on the protein, this produces colorations with different absorbance. The catalytic activity of the reaction system is affected because different proteins are absorbed onto the surface of the AuNPs. The absorbance values at 370, 450, and 650 nm are different in the presence of different proteins. The array can discriminate 8 proteins at 50 nM concentration levels with 100% accuracy. It also can discriminate proteins being present in different concentrations and mixtures with different molar ratios of proteins. Human urine spikes with 8 proteins (Concanavalin A), Cytochrome C, Horseradish peroxidase, Human serum albumin, Myoglobin, Pepsin, Trypsin, Papain) can be differentiated by applying hierarchical cluster analysis. The system also can discriminate proteins at different concentrations and the mixtures with different molar ratios in bovine serum. This indicates that the array has a great potential in terms of analyzing biological fluids.

Ultrasensitive and selective colorimetric detection of acetamiprid pesticide based on the enhanced peroxidase-like activity of gold nanoparticles

A colorimetric aptasensor based on the peroxidase activity of AuNPs has been proposed for the rapid and ultrasensitive detection of acetamiprid.

Aptamer based photometric assay for the antibiotic sulfadimethoxine based on the inhibition and reactivation of the peroxidase-like activity of gold nanoparticles

It is known that gold nanoparticles (AuNPs) possess peroxidase-like activity. They can catalyze the oxidation of 3,3,5,5-tetramethylbenzidine by H2O2 which leads to a color change from red to blue. It is shown here that the peroxidase-like activity of AuNPs can be inhibited by passivating its surface passivation with a ssDNA aptamer against sulfadimethoxine. If, however, the target molecule (sulfadimethoxine) is present, the aptamer is desorbed from the AuNPs surface, and this results in the reactivation of the catalytic property of the AuNPs. The color change of the solution (from purple to blue) is related to the analyte concentration, and this can be judged visually or by UV-visible absorptiometry at 650 nm. The assay, under optimized conditions, has a detection limit of 10 ng·mL−1 of sulfadimethoxine, and the calibration plot is linear over a rather wide concentration range (0.01–1000 μg·mL−1). The assay can be performed within <15 min, is sensitive, and therefore is well suited for fast screening in food analysis. Conceivably, it can be extended to many other small analytes for which aptamers are available.

Pitfalls and capabilities of various hydrogen donors in evaluation of peroxidase-like activity of gold nanoparticles.

Catalytic nanomaterials, widely used as substitutes of peroxidase, exhibit unique properties, which are unattainable for native enzymes. However, their activity is usually examined by means of substrates developed and methods standardized for horseradish peroxidase (HRP). The aim of the presented work was to determine the scope of usefulness of chromogenic substrates for gold nanoparticle (AuNP) activity studies under conditions which significantly extend beyond the activity range of a native HRP. The applicability of chromogens such as 3,3′5,5′-tetramethylbenzidine (TMB), o-phenylenediamine (OPD), 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) beyond the typical range of pH, and for the samples of high concentration of hydrogen peroxide was examined. The conducted research confirmed the usefulness of ABTS and TMB in acidic media (pH 2.5–3.5). At the same time, potential interferences from chloride anion, unobservable for HRP-based assays, were indicated. Moreover, a number of potentially useful hints concerning relations of concentration of substrates and catalyst for aromatic amine oxidation (TMB and OPD) were proposed. By increasing the concentration of chromogens and thanks to assuring the relatively low conversion of the reaction, the stability of TMB and OPD oxidation product was improved even in acidic media. The comparative studies of H2O2 affinity to the surface of AuNPs in the presence of various hydrogen donors underlined the superiority of phenolic compounds over aromatic amines and ABTS in the case of the samples of relatively low H2O2 concentration. This work highlights some improvements in the methods of HRP-like activity characterization of NPs. It provides a critical analysis of the major challenges, which may emerge in a case of bioanalytical assays employing the catalytic nanoparticles as labels.

Direct electrochemical detection of kanamycin based on peroxidase-like activity of gold nanoparticles

An enzyme-free, ultrasensitive electrochemical detection of kanamycin residue was achieved based on mimetic peroxidase activity of gold nanoparticles (AuNPs) and target-induced replacement of the aptamer. AuNPs which were synthesized using tyrosine as a reducing and capping agent, exhibited mimetic peroxidase activity. In the presence of kanamycin-specific aptamer, however, the single-stranded DNA (ssDNA) adsorbed on the surface of AuNPs via the interaction between the bases of ssDNA and AuNPs, and therefore blocked the catalytic site of AuNPs, and inhibited their peroxidase activity. While in the presence of target kanamycin, it bound with the adsorbed aptamer on AuNPs with high affinity, exposed the surface of AuNPs and recovered the peroxidase activity. Then AuNPs catalyzed the reaction between H2O2 and reduced thionine to produce oxidized thionine. The latter exhibited a distinct reduction peak on gold electrode in differential pulse voltammetry (DPV), and could be utilized to quantify the concentration of kanamycin. Under the optimized conditions, the proposed electrochemical assay showed an extremely high sensitivity towards kanamycin, with a linear relationship between the peak current and the concentration of kanamycin in the range of 0.1–60 nM, and a detection limit of 0.06 nM. Moreover, the established approach was successfully applied in the detection of kanamycin in honey samples. Therefore, the proposed electrochemical assay has great potential in the fields of food quality control and environmental monitoring.

Colorimetric detection of urea, urease, and urease inhibitor based on the peroxidase-like activity of gold nanoparticles

Herein, we reported for the first time that gold nanoparticles-catalyzed 3,3′,5,5′-tetramethylbenzidine-H2O2 system can serve as an ultrasensitive colorimetric pH indicator. Gold nanoparticles acted as a catalyst and imitated the function of horseradish peroxidase. The absorbance at 450 nm of the yellow-color product in the catalytic reaction exhibited a linear fashion over the pH range of 6.40–6.60. On the basis of this property, we constructed a novel sensing platform for the determination of urea, urease, and urease inhibitor. The limit of detection for urea and urease was 5 μM and 1.8 U/L, respectively. The half-maximal inhibition value IC50 of acetohydroxamic acid was found to be 0.05 mM. Urea in human urine and urease in soil were detected with satisfied results.

Naked-eye sensitive ELISA-like assay based on gold-enhanced peroxidase-like immunogold activity.

A naked-eye sensitive ELISA-like assay was developed based on gold-enhanced peroxidase-like activity of gold nanoparticles (AuNPs). Using human IgG (H-IgG) as an analytical model, goat anti-human IgG antibody (anti-IgG) adsorbed on microtiter plate and AuNPs-labeled anti-IgG acted as capture antibody and detection antibody, respectively. Because the surfaces of AuNPs were blocked by protein molecules, the peroxidase-like activity of AuNPs was almost inhibited, evaluated by the catalytic oxidation of peroxidase enzyme substrate 3,3',5,5'-tetramethylbenzidine (TMB), which could produce a bright blue color in the presence of H2O2. Fortunately, the catalytic ability of AuNPs was dramatically increased by the deposition of gold due to the formation of a new gold shell on immunogold. Under optimal reaction conditions, the colorimetric immunoassay presented a good linear relationship in the range of 0.7-100 ng/mL and the limit of detection (LOD) of 0.3 ng/mL calculated by 3σ/S for UV-vis detection, and obtained LOD of 5 ng/mL for naked-eye detection. The obtained results were competitive with conventional sandwich ELISA with the LOD of 1.6 ng/mL. Furthermore, this developed colorimetric immunoassay was successfully applied to diluted human serum and fetal bovine serum samples, and predicted a broad prospect for the use of peroxidase-like activity involving nanomaterials in bioassay and diagnostics.

Peroxidase-like activity of gold nanoparticles stabilized by hyperbranched polyglycidol derivatives over a wide pH range

The aim of this work was to carry out comparative studies on the peroxidase-like activity of gold nanoparticles (AuNPs) stabilized with low molecular weight hyperbranched polyglycidol (HBPG-OH) and its derivative modified with maleic acid residues (HBPG-COOH). The influence of the stabilizer to gold precursor ratio on the size and morphology of nanoparticles obtained was checked, and prepared nanoparticles were characterized by means of transmission electron microscopy and UV-Vis spectroscopy. The results indicated the divergent effect of increasing the concentration of stabilizers (HBPG-OH or HBPG-COOH) on the size of the nanostructures obtained. The gold nanoparticles obtained were characterized as having intrinsic peroxidase-like activity and the mechanism of catalysis in acidic and alkaline mediums was consistent with the standard Michaelis–Menten kinetics, revealing a strong affinity of AuNPs with 2, 2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 3, 3′, 5, 5′-tetramethylbenzidine (TMB), and significantly lower affinity towards phenol. By comparing the kinetic parameters, a negligible effect of polymeric ligand charge on activity against various types of substrates (anionic or cationic) was indicated. The superiority of steric stabilization via the application of tested low-weight hyperbranched polymers over typical stabilizers in preventing salt-induced aggregation and maintaining high catalytic activity in time was proved. The applied hyperbranched stabilizers provide a good tool for manufacturing gold-based nanozymes, which are highly stable and active over a wide pH range.