A dual-mode optical assay for iron(II) and gallic acid based on Fenton reaction

The hydroxyl radicals ( · OH) produced by the Fenton reaction of iron(II) and hydrogen peroxide (H₂O₂) can oxidize the colorless 3,3′,5,5′-tetramethylbenzidine (TMB) to blue oxidized TMB (Ox-TMB), resulting in a decrease in the fluorescence intensity of the reaction system and an increase in ultraviolet absorption. Ox-TMB had a visible absorption peak at 625 nm and a fluorescence peak around 420 nm. When gallic acid (GA) was added to the system, Ox-TMB was reduced to TMB, which made the color of the system disappear and the fluorescence recover. The linear ranges for determination of iron(II) were 0.5–10 μM (fluorometric) and 0.5–20 μM (colorimetric), and the detection limits were 0.25 μM (fluorometric) and 0.28 μM (colorimetric). The linear ranges for determination of GA were 0–80 μM (fluorometric) and 0–60 μM (colorimetric), and the detection limits were 0.31 μM (fluorometric) and 0.8 μM (colorimetric). The results of anti-interference experiments shew that this dual-mode assay had very good selectivity for the determination of iron(II) and GA.     

Ultrasensitive and selective colorimetric detection of uric acid using peroxidase mimetic activity of biogenic palladium nanoparticles

Uric acid (2,6,8-trihydroxypurine) is a metabolic product of purine, which is one of the important markers of human health. The development of a rapid, facile, highly sensitive, and selective method for uric acid detection is critical for the diagnosis of related diseases and is still a strategic challenge. In this study, we developed a highly sensitive and selective colorimetric assay for the detection of uric acid using biogenic palladium nanoparticles (Pd NPs). The synthesized nanoparticles were shown to acquire peroxidase mimetic activity that oxidized 3,3′,5,5′-tetramethylbenzidine and produced a blue colour in an assay. The developed colorimetric assay is instrument-free detection of uric acid with a limit of detection of 0.05 μM and a 1.11 μM limit of quantification (LOQ). This is the first report determining the LOQ for a colorimetric assay that gives the lowest quantity of analyte that can be evaluated with more precision under the specified conditions of the analysis. The developed assay had a linear response at low uric acid concentrations of 0.05 to 1 μM and a 0.99841 linear regression correlation coefficient. This colorimetric detection provides a rapid, cost-effective, and easy-to-use platform for the clinical diagnosis of uric acid biomarkers.     

Colorimetric detection of gallic acid based on the enhanced oxidase‐like activity of floral‐like magnetic Fe3O4@MnO2

In this study, a colorimetric method was developed for rapid and sensitive determination of gallic acid (GA) by using floral‐like magnetic Fe₃O₄@MnO₂ composite material with enhanced oxidase‐like activity. Fe₃O₄@MnO₂ composite material is able to oxidize 3,3′,5,5′‐tetramethylbenzidine (TMB) to a blue product (oxTMB) with apparent color change and absorbance at 652 nm. GA can reduce the oxTMB yielding a fading blue color. Based on these results, a technique is proposed to detect GA quantitatively and qualitatively with UV–vis spectroscopy and bare eyes. A low detection limit of 0.105 μM and a detection range of 0.01 to 15 μM were obtained with UV–vis spectroscopy. This methodology possesses high potential for application in determination of GA.     

A rapid and high‐throughput method for the determination of serum uric acid based on microarray technology and nanomaterial

Serum uric acid (SUA) is a new therapeutic target for non‐alcoholic fatty liver disease (NAFLD). In this study, we introduced a chemiluminescence (CL) method combined with microarray technology and a simple fabrication procedure to obtain a highly sensitive SUA probe based on a mesoporous metal oxide nanomaterial. The high‐throughput method was based on the generation of H₂O₂ from SUA by immobilized uricase and its measurement by a CL reaction catalyzed by mesoporous metal oxide nanomaterials. The CL probe was designed for SUA The linear range of the uric acid concentration was 0.6–9 μM and the detection limit was 0.1 μM. In comparison with the other SUA detection techniques, this method has the advantages of a low detection limit, high sensitivity and simplicity. A new sensitive high‐throughput approach was obtained for the determination of SUA.     

An amperometric uric acid biosensor based on multiwalled carbon nanotube-gold nanoparticle composite

An amperometric uric acid biosensor was fabricated by immobilizing uricase (EC 1.7.3.3) onto gold nanoparticle (AuNP)/multiwalled carbon nanotube (MWCNT) layer deposited on Au electrode via carbodiimide linkage. Determination of uric acid was performed by oxidation of enzymically generated H₂O₂ at 0.4 V. The sensor showed optimal response within 7 s at 40 °C in 50 mM Tris–HCl buffer (pH 7.5). The linear working range of the biosensor was 0.01–0.8 mM. The limit of detection (LOD) was 0.01 mM. The sensor measured uric acid levels in serum of healthy individuals and persons suffering from gout. The analytical recoveries of the added uric acid, 10 and 20 mg L^–1, were 98.0% and 96.5%, respectively. Within- and between-batch coefficients of variation were less than 5.6% and less than 4.7%, respectively. A good correlation (r = 0.998) was obtained between serum uric acid values by the standard enzymic colorimetric method and the current method. A number of serum substances had practically no interference. The sensor was used in more than 200 assays and had a storage life of 120 days at 4 °C.     

Electro-optical behaviour of CuFe2O4@rGO nanocomposite for nonenzymatic detection of uric acid via the electrochemical method

Uric acid (UA) is a blood and urine component obtained as a metabolic by-product of purine nucleotides. Abnormalities in UA metabolism cause crystal deposition as monosodium urate and lead to various diseases such as gout, hyperuricemia, Lesch–Nyhan syndrome, etc. Monitoring these diseases requires a rapid, sensitive, selective, and portable detection approach. Therefore, this study demonstrates the hydrothermal synthesis of CuFe₂O₄/reduced graphene oxide (rGO) nanocomposite for selective detection of UA. After the nanocomposite synthesis, characterization was performed by X-ray diffraction spectroscopy, Fourier transform infrared spectroscopy, Raman spectroscopy, X-ray photoelectron spectroscopy, UV–visible spectrometry, atomic force spectroscopy, scanning electron microscopy, and electrochemical analysis. Furthermore, from the electrochemical analysis using cyclic voltammetry (CV), kinetic studies were carried out by varying the scan rate to obtain the diffusion coefficient, surface concentration, and rate of charge transfer to achieve a calibration curve that indicates the quasi reversible nature of the fabricated electrode with a linear regression coefficient of oxidation (R²: 0.9992) and reduction (R²: 0.9971) peaks. Moreover, the fabricated nonenzymatic amperometric sensor to detect UA with a linearity (R²: 0.9989) of 1–400 μM was highly sensitive (2.75 × 10⁻⁴ mAμM⁻¹ cm⁻²) and had a lower limit of detection (0.01231 μM) at pH 7.5 in phosphate-buffered saline solution. Therefore, the CuFe₂O₄/rGO/ITO-based nonenzymatic sensor could detect interfering agents and spiked real bovine serum samples with higher sensitivity and selectivity for UA detection.     

3,5,2',4'-Tetrahydroxychalcone, a new non-purine xanthine oxidase inhibitor

Xanthine oxidase is a key enzyme that catalyses hypoxanthine and xanthine to uric acid and the overproduction of uric acid will lead to hyperuricemia which is an important cause of gout. In the present study, three chalcone derivatives were synthesized and evaluated for inhibitory activity against xanthine oxidase in vitro. Of the compounds, only Compound 1, 3,5,2′,4′-tetrahydroxychalcone, exhibited a significant inhibitory activity on xanthine oxidase with an IC₅₀ value of 22.5 μM. Lineweaver–Burk transformation of the inhibition kinetics data demonstrated that it was a competitive inhibitor of xanthine oxidase and Ki value was 17.4 μM. In vivo, intragastric administration of Compound 1 was able to significantly reduce serum uric acid levels and inhibited hepatic xanthine oxidase activities of hyperuricemic mice in a dose-dependent manner. Acute toxicity study in mice showed that Compound 1 was very safe at a dose of up to 5 g/kg. These results suggest that Compound 1 is a novel competitive xanthine oxidase inhibitor and is worthy of further development.     

A colorimetric detection of dopamine in urine and serum based on the CeO2@ZIF-8/Cu-CDs laccase-like nanozyme activity

This study reports a sensitive and selective colorimetric approach for the analysis of dopamine (DA) based on CeO₂@ZIF-8/Cu-CDs laccase-like nanozymes activity. The CeO₂@ZIF-8/Cu-CDs was synthesized using cerium oxide (CeO₂) and copper-doped carbon dots (Cu-CDs) with 2-methylimidazole by a facilely hydrothermal approach. The CeO₂@ZIF-8/Cu-CDs exhibited excellent laccase-like nanozymes activity and can oxidize the colorless substrate (DA) to red product with 4-aminoantipyrine as the chromogenic agent. The Michaelis–Menten constant (K_m) and the maximal velocity (V_max) of CeO₂@ZIF-8/Cu-CDs are 0.20 mM and 1.48 μM/min, respectively. The detection method has a linear range of 0.05–7.5 μg/mL and a detection limit as low as 8.5 ng/mL with good reproducibility. The developed colorimetric sensor was applied to rapid and precise quantitative evaluation of DA levels in serum and urine samples. This study presents a new approach for detecting biological molecules by utilizing the controlled regulation of nanozymes' laccase-like activity.     

Improved peroxyl radical scavenging TOSC assay to quantify antioxidant capacity using SIFT-MS

Abstract

We report a new, fast, sensitive variation of the total oxyradical scavenging capacity (TOSC) assay for measuring the antioxidant capacity of pure compounds, plant extracts and biological fluids using selected ion flow tube mass spectrometry (SIFT-MS). The TOSC assay examines the partial inhibition of ethene formation in the presence of antioxidants that compete with α-keto-γ-methiolbutyric acid (KMBA) for reactive oxygen species. The SIFT-MS-TOSC assay takes 15 s for each ethene analysis and the time interval between consecutive analyses is 20 s. We demonstrate the method by monitoring the antioxidant capacity of several standard radical scavengers of peroxyl radicals. For peroxyl radicals the measured SIFT-MS-TOSC concentrations necessary to produce 50% inhibition of radical reaction with KMBA are 6.1 ± 0.3 μM for Trolox, 5.7 ± 0.3 μM for ascorbic acid, 8.4 ± 0.4 μM for uric acid and 38 ± 2 μM for reduced glutathione.     

Polyethylene terephthalate membrane as a support for covalent immobilization of uricase and its application in serum urate determination

A method is described for covalent immobilization of uricase onto polyethylene terephthalate (PET) membrane with a conjugation yield of 4.44 μg/cm² and 66.6% retention of initial activity of free enzyme. The enzyme exhibited an increase in optimum pH from pH 7.0 to 8.5 and K_m for uric acid from 0.075 mM to 0.13 mM but slight decrease in temp. for maximum activity from 37 °C to 35 °C after immobilization. A colorimetric method for determination of serum uric acid was developed using immobilized uricase, which is based on measurement of H₂O₂ by a color reaction consisting of 3,5-dichlorobenzene sulphonic acid (DHBS), 4-aminoantipyrine and peroxidase as chromogenic system. Minimum detection limit of the method was 0.05 mM. Analytical recovery of added uric acid (5 mg/dl and 10 mg/dl) was 94.3% and 89.8%, respectively. Within and between batch coefficient of variation (CV) were <3.2% and <4.3%, respectively. A good correlation (r = 0.98) was found between uric acid values by standard enzymic colorimetric method and the present method. The immobilized uricase was reused 100 times during the span of 60 days without any considerable loss of activity, when stored in reaction buffer at 4 °C. The support chosen for the present study was biocompatible, antimicrobial, inert, impact resistant, light weight and had good shelf life.     

Rapid and sensitive colorimetric detection of ascorbic acid in food based on the intrinsic oxidase‐like activity of MnO2 nanosheets

In this paper, we report a colorimetric sensor for the rapid, selective detection of ascorbic acid (AA) in aqueous solutions. Single‐layered MnO₂ nanosheets were established as an artificial oxidase; consequently colorless 3,3´,5,5´‐tetramethylbenzidine (TMB) was oxidized to a blue product (oxTMB), with increase in absorbance at 650 nm. The absorbance of the reaction system decreased after introduction AA, which reduced MnO₂ into Mn²⁺. Under optimum conditions, a detection limit of 62.81 nM for AA in aqueous solutions could be achieved. The linear response range for AA was 0.25–30 μM with a correlation coefficient of 0.996. Importantly, the MnO₂ nanosheet–TMB chromogenic reaction exhibited great selectivity as there was no interference from other metal ions, amino acids and small biological molecules. The proposed colorimetric sensing of AA could be applied for fruit, juice and pharmaceutical samples. Moreover, the proposed sensor showed satisfying performance, including low cost, easy preparation, rapid detection, and good biocompatibility.     

Nano RNA aptamer wire for analysis of vitamin B₁₂

A simple and stable RNA aptamer-based colorimetric sensor for the detection of vitamin B₁₂ using gold nanoparticles (AuNPs) has been proposed. Vitamin B₁₂ belongs to the B vitamin group and prevents pernicious anemia, which is caused by vitamin B₁₂ deficiency. A highly stable RNA aptamer that binds to vitamin B₁₂ was employed by structural modification of 2′-hydroxyl group of ribose to 2′-flouro in all pyrimidines indicated in lowercase in 35-mer aptamer (5′ GGA Acc GGu GcG cAu AAc cAc cuc AGu GcG AGc AA 3′). Aggregation of AuNPs was specifically induced by desorption of the vitamin B₁₂ binding RNA aptamer from the surface of AuNPs as a result of the aptamer–target interaction, leading to the color change from red to purple. The level of detection of vitamin B₁₂ was 0.1 μg/ml by successful optimization of the amount of the aptamer, AuNPs, salts, and stability of the aptamer. Analysis of vitamin B₁₂ was carried out, and the observed recovery was 92 to 95.3% with a relative standard deviation in the range of 2.08 to 8.27%. The results obtained were compared with those of the ultraviolet–visible (UV–vis) spectrometry method. This colorimetric aptasensor is advantageous for on-site detection with the naked eye.     

Concerning the presence and formation of ascorbic acid in yeasts

The selective, sensitive method of analysis of ascorbic acid by high performance liquid chromatography with electrochemical detection (HPLC/EC) has been used to determine the ascorbic acid content of cell extracts from yeasts grown in glucose-free medium, 0.3 M D-glucose, and 0.112 M L-galactono-1,4-lactone. Saccharomyces cerevisiae (strain G-25 and its tetraploid) and a commercial baker's yeast contained less than 2 μg ascorbic acid g^?1 wet wt. of cells when grown for 22 h in glucose-free medium. In 0.3 M D-glucose, only the commercial baker's yeast gave a slight increase (2–50 μg g^?1 wet wt. in 22 h). In 0.112 M L-galactono-1,4-lactone, all three strains produced ascorbic acid (372–587 μg g^?1 wet wt. in 22 h). Lypomyces starkeyi, a species previously reported to contain a significant amount of ascorbic acid (Heick et al., Can. J. Biochem., 47 (1972) 752), was essentially devoid of ascorbic acid under all three conditions of incubation although it did contain an HPLC/EC reactive peak (R_T = 0.87 relative to ascorbic acid) that was readily oxidized by charcoal in the presence of oxygen. The identity of this new compound remains to be determined.     

A novel colorimetric fluorescent probe for detecting hydrazine in living cells and zebrafish

Hydrazine (NH₂NH₂) is a highly toxic organic substance that poses a threat to human health. Monitoring hydrazine with high sensitivity and selectivity is very important. Here, a simple colorimetric fluorescent probe for hydrazine detection, which is a seminaphthorhodafluor derivative containing thiophene-2-carboxylic acid ester reaction site, was rationally constructed. The probe itself exhibits weak fluorescence. The fluorescence is significantly enhanced when hydrazine is added. The probe exhibited a broad linear range (0–1 mM) with satisfactory selectivity and sensitivity (limit of detection 36.4 μM), which turned out to be an excellent fluorescent probe for monitoring hydrazine. Additionally, the probe was used to track hydrazine in living cells and zebrafish with great success, and the detection performance was satisfying. These results proved that this type of fluorescent probe with the thiophene-2-carboxylic acid ester structure can detect hydrazine with higher selectivity and sensitivity.     

Sensitive colorimetric visualization of dihydronicotinamide adenine dinucleotide based on anti-aggregation of gold nanoparticles via boronic acid-diol binding

A facile, highly sensitive colorimetric strategy for dihydronicotinamide adenine dinucleotide (NADH) detection is proposed based on anti-aggregation of gold nanoparticles (AuNPs) via boronic acid-diol binding chemistry. The aggregation agent, 4-mercaptophenylboronic acid (MPBA), has specific affinity for AuNPs through Au-S interaction, leading to the aggregation of AuNPs by self-dehydration condensation at a certain concentration, which is responsible for a visible color change of AuNPs from wine red to blue. With the addition of NADH, MPBA would prefer reacting with NADH to form stable borate ester via boronic acid-diol binding dependent on the pH and solvent, revealing an obvious color change from blue to red with increasing the concentration of NADH. The anti-aggregation effect of NADH on AuNPs was seen by the naked eye and monitored by UV-vis extinction spectra. The linear range of the colorimetric sensor for NADH is from 8.0 × 10(-9)M to 8.0 × 10(-6)M, with a low detection limit of 2.0 nM. The as-established colorimetric strategy opened a new avenue for NADH determination.     

Uric acid mediates photodynamic inactivation of caprine alpha-2-macroglobulin

Uric acid (2,6,8 trioxopurine), the end product of purine metabolism in mammalian systems, has shown a wide range of antioxidant properties including scavenging of hydroxyl radical and singlet oxygen. In this study we show that in the presence of visible light, uric acid disrupted caprine alpha-2-macroglobulin (α₂M) structure and antiproteolytic function in vitro. Proteinase cleaves the bait region of caprine inhibitor inducing major conformational changes and entrapping the enzyme within its molecular cage. In contrast to native α₂M, modified antiproteinase lost half of its antiproteolytic potential within 4 hours of uric acid exposure. The changes in uv-absorption spectra of the treated protein suggested possible spatial rearrangement of subunits or conformational change. Analysis of the mechanism by which α₂M was inactivated revealed that the process was dependent on generation of superoxide anion and hydrogen peroxide. Our findings suggest that antiproteolytic activity of caprine α₂M could be compromised via oxidative modification mediated by uric acid. Moreover, low concentrations of α₂M were found to stimulate superoxide production by some unknown mechanism.     

Green spectrophotometric and spectrofluorimetric determination of biperiden hydrochloride using erythrosine B sensing probe

Erythrosine B (EB) is a food colorant antiviral xanthene dye that has many applications as a color additive in pharmaceuticals and cosmetics. Its use as a sensor for spectrofluorimetric and spectrophotometric analysis of amine-based pharmaceuticals renders many advantages because of its availability, low cost, rapid labeling, and high sensitivity. Herein, two fast and sensitive spectrofluorimetric and spectrophotometric methods were established for the estimation of the anti-Parkinson drug, biperiden (BIP) hydrochloride (HCl), in its raw material and tablet forms. The proposed methods depended on the interaction between the phenolic group of EB and the tertiary amino group of the studied analyte to form an ion-pair complex at pH 4 using the Britton Robinson buffer. The spectrofluorimetric method is based on the measurement of the quenching power of BIP HCl on the fluorescence intensity of EB at λ_ex/em = 527.0/550.9 nm. This method was rectilinear over the concentration range of 0.1–1.0 μg/mL with a limit of detection (LOD) = 0.017 μg/mL and a limit of quantification (LOQ) = 0.05 μg/mL. Meanwhile, the colorimetric method involved monitoring the absorbance of the formed ion-pair complex at 555 nm, showing a linearity range of 0.4–5.0 μg/mL with LOD = 0.106 μg/mL and LOQ = 0.322 μg/mL. The proposed methods were assessed for the greenness, indicating the greenness of the developed methods.     

A new colorimetric and far‐red fluorescent probe for hydrazine with a large red‐shifted absorption spectrum

Recently, growing attention has been paid to the detection of hydrazine (NH₂NH₂) because of its important roles in industrial chemical and high toxicity to human beings. Herein, we have constructed a new colorimetric and far‐red fluorescent probe containing a receptor of 4‐bromobutanoate to selectively detect hydrazine. The probe could detect hydrazine quantitatively in the range of 40–500 μM with the detection limit of 2.9 μM. In addition, the probe could monitor hydrazine by the ratiometric method with a large (185 nm) red‐shifted absorption spectrum, and the color changes from yellow to blue make it as a ‘naked‐eye’ indicator for hydrazine. Consequently, our proposed probe would be of great benefit for monitoring hydrazine in aqueous solution.     

Sensitive and Robust Colorimetric Sensing of Sulfide Anion by Plasmonic Nanosensors Based on Quick Crystal Growth

A highly sensitive and selective method for colorimetric sensing of sulfide anions in aqueous solutions is illustrated. The sensing mechanism is based on quick crystallization from Ag to Ag₂S in the presence of sulfide anions which alter the dielectric properties of the Au/Ag core/shell nanorods. The longitudinal surface plasmon resonance peak of the Au/Ag nanorods at about 686 nm undergoes a redshift and the color of the nanorod solution also changes from light green to purple. Sulfide ions at a concentration of 4.0 μM (1.3 ppb) can be detected visually and a sensitivity of 0.5 μM (167 ppt) is achieved by Vis–near-infrared spectrophotometry. Compared to other plasmonic sensors, our Au/Ag nanorod probe does not require surface modification while exhibiting high stability and robustness under different pH conditions. This simple and cost-effective sensing platform provides a rapid and convenient detection for sulfide anions at concentrations far below the hazardous limit in aqueous media.     

Hemin-incorporated nanoflowers as enzyme mimics for colorimetric detection of foodborne pathogenic bacteria

Rapid, sensitive and point-of-care detection of foodborne pathogenic bacteria is essential for food safety. In this study, we found that hemin-concanavalin A hybrid nanoflowers (HCH nanoflowers), as solid mimic peroxidase, could catalyze oxidation of 2,2′-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) diammonium salt (ABTS) in the presence of H₂O₂ to a green-colored product. HCH nanoflowers, integrating the essential functions of both biological recognition and signal amplification, meet the requirements of signal labels for colorimetric immunoassay of bacteria. In view of the excellent peroxidase mimetic catalytic activity of HCH nanoflowers, a colorimetric biosensing platform was newly constructed and applied for sensitive detection of foodborne Escherichia coli O157:H7 (E. coli O157:H7). The corresponding detection limits was as low as 4.1?CFU/mL with wide linear ranges (10¹–10⁶?CFU/mL).