Direct Colorimetric Detection Research Articles

An Approach to Identifying Single-Nucleotide Mutations Using Noncovalent Associates of Gold Nanoparticles with Fluorescently Labeled Oligonucleotides.

Globally, widespread tuberculosis is one of the acute problems of healthcare. Drug-resistant forms of tuberculosis require a personalized approach to treatment. Currently, rapid methods for detecting drug resistance of Mycobacterium tuberculosis (MTB) to some antituberculosis drugs are often used and involve optical, electrochemical, or PCR-based assays. Despite the large number of these assays, it is necessary to develop new tests (for drug-resistant MTB strains) that are structurally simple and do not require specialized equipment. Colorimetric assays involving a colloidal solution of gold nanoparticles (AuNPs) have good potential for the development of the needed diagnostic tools. Here, conditions were found for the formation of tandem duplexes between DNA probes and DNA targets, representing a part of MTB gene gyrA, either wildtype or containing a single-nucleotide polymorphism associated with fluoroquinolone resistance of MTB. Adsorption of the duplexes on AuNPs allowed to distinguish the two targets owing to the formation of nano-constructs of different structures. Interaction of DNA with AuNPs was analyzed by optical spectroscopy, dynamic light scattering, and transmission electron microscopy. A scheme is proposed for direct colorimetric detection of the fluoroquinolone-resistance-associated single-nucleotide polymorphism at a 2 nM concentration in a liquid system based on a shift of AuNPs' optical absorption maximum.

Direct colorimetric detection of triacetone triperoxide explosive by hydroxylamine-mediated fenton chemistry catalyzed by goethite nanoparticles

Due to the absence of nitro groups or aromatic structures in its structure, spectrophotometric methods that can directly determine triacetone triperoxide (TATP) are absent. In the proposed study, a molecular spectroscopic sensor was developed based on the decomposition of TATP by hydrolysis without the addition of external acid, allowing the direct determination of TATP. The detection principle of the developed sensor is based on the decomposition of TATP by the hydrochloric acid formed as a result of the reaction of TATP with hydroxylamine hydrochloride, taking advantage of the solubility of TATP in acetone, and the hydrogen peroxide formed as a degradation product to form hydroxyl radicals catalyzed by goethite nanoparticles. The formed hydroxyl radicals, thanks to their strong oxidation properties, oxidized N,N-dimethyl-p-phenylenediamine (DMPD) to the pink-colored DMPD•+ radical cation, and the direct determination of TATP was performed by measuring the absorbance of the colored product formed. Thus, a chemo-cybernetic sequence of reactions is proposed for intact TATP assay in a circular self-sustained system, where the reactants at a given step of reactions are supplied by the products of a previous step. By applying the developed method to TATP samples prepared in acetone, absorbances due to DMPD•+ were read at 554 nm to evaluate the results. A calibration curve was created between absorbance versus TATP concentration within a final range of 10 – 33 mg L-1. The limit of detection (LOD) and limit of quantification (LOQ) of the developed TATP method were 3.3 mg L-1 and 10.8 mg L-1, respectively. Five replicate measurements were performed to determine the intra-assay and inter-assay precision of the spectrophotometric determination of TATP, and the relative standard deviations (RSD %) for TATP were found as 2.70 % and 3.76 %, respectively. Interference analysis was carried out to see the effects of ions commonly present in soil/groundwater, and of the substances used as camouflage materials due to their color and appearance similarity to TATP. The selectivity of the method toward TATP was investigated by examining the analyte recovery from explosive mixtures. The developed method was validated against a reference GC–MS method using TATP-contaminated soil samples, and the results were compared statistically.

Enhanced oxidoreductase activity of diatom frustule supported Fe3O4 for sensitive colorimetric detection of Cr(VI)

Given Cr(VI) is a ubiquitous water pollutant posing significant public health risks, there is a need for an inexpensive and easy-to-use detection tool. Herein, a diatom frustule-supported Fe3O4 (DF-Fe3O4) with enhanced oxidoreductase mimetic activity is reported for direct colorimetric detection of Cr(VI). Unlike the peroxidase nanozymes which are often reported for colorimetric detection, DF-Fe3O4 triggered a chromogenic redox reaction between TMB and Cr(VI) without requiring H2O2. This enables direct redox reactions while circumventing potential interferences associated with the use of H2O2. DF-Fe3O4 was synthesized with a coprecipitation method and subsequently characterized by SEM, XRD, and XPS techniques, revealing the distribution of Fe3O4 NPs on the frustule matrix. The frustule obtained from Nitzschia sp. of a sediment sample rendered a robust catalytic support enhancing the oxidoreductase activity of pristine Fe3O4 NPs by 20.8 %. Further, the nanozyme maintains 73 % of its activity even at 95 °C while losing only 33 % of its activity after one month of storage. The oxidoreductase mimetic activity was evaluated using the chromogenic redox reaction between TMB and Cr(VI) which rapidly forms a blue color (λmax= 654 nm), where its intensity forms the basis of the colorimetric detection. With a Km value of 0.058 mM, the nanozyme was able to selectively detect Cr(VI) down to 0.21 µM with a linear range between 0.1–500 µM. Recoveries from spiked wastewater samples were between 91.14–102.20 %. The obtained analytical figures of merits demonstrated the applicability of the developed sensor for Cr (VI) analysis devoid of complex instrumentation in the established analytical methods.

Point-of-care detection of hydroxyurea drug in serum using a supramolecular enzyme mimetic

The sole approved therapeutic drug for sickle cell disease management is provided by hydroxyurea, requiring its precise detection and monitoring in biological matrices such as human blood serum for personalized treatment. Herein, we introduce a direct colorimetric detection method for hydroxyurea in serum using a metallo-supramolecular assembly called Vesi-ImCu that mimics the peroxidase enzyme. The Vesi-ImCu enzyme mimetic comprises a catalytically active copper center coordinated to synthetic imidazole containing surfactant and iminodiacetic acid capping moieties. By leveraging the peroxidase-like activity of Vesi-ImCu, we have demonstrated that it catalyzes the efficient production of nitric oxide from hydroxyurea that reacts with the Griess reagent, producing an intense pink color. This approach enables the naked-eye detection of hydroxyurea, with notable efficacy even in complex biological matrices such as human blood serum. The Vesi-ImCu suprazyme-based approach offers several advantages, including high sensitivity (limit of detection of 75 µM), clinically relevant dynamic range of hydroxyurea detection, minimum batch-to-batch variation, and the ability to perform mobile phone-based quantification. To our knowledge, this is the first report on direct colorimetric quantification of HU in serum. With its visual detection ability, high-end equipment-free quantification, user-friendliness and portability, the present enzyme mimetic holds great promise for point-of-care detection applications.

CuCeTA nanoflowers as an efficient peroxidase candidate for direct colorimetric detection of glyphosate.

Conventional nanozyme-based pesticide detection often requires the assistance of acetylcholinesterase. In this work, a CuCeTA nanozyme was successfully designed for the direct colorimetric detection of glyphosate. Direct detection can effectively avoid the problems caused by cascading with natural enzymes such as acetylcholinesterase. By assembling tannic acid, copper sulfate pentahydrate and cerium(III) nitrate hexahydrate, CuCeTA nanoflowers were prepared. The obtained CuCeTA possessed excellent peroxidase-like activity that could catalyze the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) to blue oxidized TMB in the presence of hydrogen peroxide. Glyphosate could effectively inhibit the peroxidase-like activity of CuCeTA while other pesticides (fenthion, chlorpyrifos, profenofos, phosmet, bromoxynil and dichlorophen) did not show significant inhibitory effects on the catalytic activity of CuCeTA. In this way, CuCeTA could be used for the colorimetric detection of glyphosate with a low detection limit of 0.025 ppm. Combined with a smartphone and imageJ software, a glyphosate test paper was designed with a detection limit of 3.09 ppm. Fourier transform infrared spectroscopy demonstrated that glyphosate and CuCeTA might be bound by coordination, which could affect the catalytic activity of CuCeTA. Our CuCeTA-based nanozyme system exhibited unique selectivity and sensitivity for glyphosate detection and this work may provide a new strategy for rapid and convenient detection of pesticides.

A Novel Preanalytical Strategy Enabling Application of a Colorimetric Nanoaptasensor for On-Site Detection of AFB1 in Cattle Feed

Aflatoxin contamination of cattle feed is responsible for serious adverse effects on animal and human health. A number of approaches have been reported to determine aflatoxin B1 (AFB1) in a variety of feed samples using aptasensors. However, rapid analysis of AFB1 in these matrices remains to be addressed in light of the complexity of the preanalytical process. Herein we describe an optimization on the preanalytical stage to minimize the sample processing steps required to perform semi-quantitative colorimetric detection of AFB1 in cattle feed using a gold nanoparticle-based aptasensor (nano-aptasensor). The optical behavior of the nano-aptasensor was characterized in different organics solvents, with acetonitrile showing the least interference on the activity of the nan-aptasensor. This solvent was selected as the extractant agent for AFB1-containing feed, allowing for the first time, direct colorimetric detection from the crude extract (detection limit of 5 µg/kg). Overall, these results lend support to the application of this technology for the on-site detection of AFB1 in the dairy sector.

Purple to Yellow Silicone Elastomers: Design of a Versatile Sensor for Screening Antioxidant Activity

AbstractAntioxidants play a key role in counteracting the adverse effects of oxidative stress in human organisms. Thus, there is a huge demand for the development of smart and convenient assays for the evaluation of antioxidant activity of synthetic and natural compounds, food samples, plant extracts, etc. The design of a solid‐state, flexible, and portable format of the traditional in vitro 2,2‐diphenyl‐1‐picrylhydrazyl (DPPH) radical test is reported. For the first time, purple DPPH radicals are physically immobilized into a silicone matrix by means of a facile and rapid process. The working principle of the resulting sensor is based on the colorimetric process that is associated with the redox reaction of DPPH radicals with antioxidants. When an antioxidant reacts with the sensor, a color change from purple to yellow can be perceived by the naked eye. The response of the sensor is investigated qualitatively and quantitatively toward food samples and selected antioxidants of different natures, and solubilized in different media. It is demonstrated that the DPPH silicone sensor is highly versatile and can be used as a ready‐to‐use sensor for direct colorimetric detection of antioxidants.

Engineering DNA-Nanozyme Interfaces for Rapid Detection of Dental Bacteria.

Engineering biological interfaces represents a powerful means to improve the performance of biosensors. Here, we developed a DNA-engineered nanozyme interface for rapid and sensitive detection of dental bacteria. We employed DNA aptamer as both molecular recognition keys and adhesive substrates to functionalize the nanozyme. Utilizing different immobilization strategies and DNA designs, a range of DNA nanoscale biointerfaces were constructed to modulate enzymatic and biological properties of the nanozyme systems. These functional biointerfaces improved the accessibility of bacteria to the nanozyme surface, providing large signal change range at optimal DNA probe density. The DNA-functionalized nanozymes demonstrate a rapid, label-free, and highly sensitive direct colorimetric detection of Streptococcus mutans, with a detection limit of 12 CFU mL-1, as well as excellent discrimination from other dental bacteria. We demonstrate the use of this biological nanointerface for identifying dental bacteria in salivary samples, showing its potential in clinical prevention and diagnosis of dental diseases.

Duplex PCR-ELONA for the detection of pork adulteration in meat products

In this work, a duplex PCR–Enzyme Linked Oligonucleotide Assay (ELONA) is reported for the sensitive and reliable detection of pork adulteration in beef and chicken products, two of the most widely consumed meat types in the world. The strategy relies on the use of species-specific tailed primers for duplex amplification and simple dilution of the PCR reactions for direct colorimetric detection via hybridization, eliminating the need for any other post-amplification steps. A high sensitivity was achieved, with as low as 71–188 pg of genomic DNA able to be detected using mixtures of control DNA from each species. The strategy was validated using DNA add-mixtures as well as DNA extracted from raw meat mixtures and 0.5–1% w/w pork could be easily detected when mixed with beef or chicken. The proposed approach is simple, sensitive and cost-effective compared to equivalent commercial kits suitable for detecting adulterant pork levels in meat products.

Direct colorimetric detection of aspartic acid in rat brain based on oriented aggregation of Janus gold nanoparticle

We report a colorimetric assay based on oriented Janus gold nanoparticles (AuNPs) aggregation for highly selective and sensitive detection of aspartic acid (Asp) in rat brain. Initially, the Janus AuNPs with relatively large size was prepared, during which the stabilizing agent polyethylene glycol (PEG) and highly selective recognition agent cysteine (Cys) were attached to the selected area at the surface. The addition of Asp causes the oriented and controllable aggregation due to the specific recognition between Cys and Asp. Such a way of aggregation leads to significantly improved long-term stability and an at least 2 orders of magnitude wider dynamic range (1.8 nM–180 μM) compared with conventional colorimetric sensors. In addition, the strategy to increase the size of AuNPs (13–45 nm) enables to efficiently increase the limit of detection (18 μM–1.8 nM). Surprisingly, such a sensor can be applied to direct sensing of cerebral Asp in rat brain, which is determined round 33.9 ± 1.3 μM with satisfactory spiked recovery (86.5 ± 7.5%) due to the excellent anti-interference ability. This work provides a direct and reliable method to evaluate the Asp involved physiological and pathological process and opens a new avenue for real sample detection.

Photoelectrocatalytic Synthesis of Hydrogen Peroxide by Molecular Copper-Porphyrin Supported on Titanium Dioxide Nanotubes.

We report on a self‐assembled system comprising a molecular copper‐porphyrin photoelectrocatalyst, 5‐(4‐carboxy‐phenyl)‐10,15,20‐triphenylporphyrinatocopper(II) (CuTPP‐COOH), covalently bound to self‐organized, anodic titania nanotube arrays (TiO2 NTs) for photoelectrochemical reduction of oxygen. Visible light irradiation of the porphyrin‐covered TiO2 NTs under cathodic polarization up to −0.3 V vs. Normal hydrogen electrode (NHE) photocatalytically produces H2O2 in pH neutral electrolyte, at room temperature and without need of sacrificial electron donors. The formation of H2O2 upon irradiation is proven and quantified by direct colorimetric detection using 4‐nitrophenyl boronic acid (p‐NPBA) as a reactant. This simple approach for the attachment of a small molecular catalyst to TiO2 NTs may ultimately allow for the preparation of a low‐cost H2O2 evolving cathode for efficient photoelectrochemical energy storage under ambient conditions.

A simple assay for direct colorimetric detection of prostatic acid phosphatase (PAP) at fg levels using biphosphonated loaded PEGylated gold nanoparticles

A fast and sensitive colorimetric assay for the determination of prostatic acid phosphatase (PAP) levels using biphosphonates-PEG-coated gold nanoparticles (BP-PEG-AuNPs) has been developed. Addition of different amounts of PAP to the AuNPs results in a significant change of color, from a red-wine color to purple, and finally to blue. The PAP should bind onto the AuNPs surface and crosslink the AuNPs inducing their aggregation and as a consequence a color change of the solution from wine red to purple/blue. The color change was monitored by UV–vis spectrophotometry or with the naked eye. The ensuing concentration-dependent red-shift of surface plasmon resonance absorption allows the determination concentration within the fg range in 10 min. Importantly, the visual detection limit (1 ng/mL) allows the rapid differential diagnosis with naked eye or image analyzer. This demonstrates that the color modification observed with BP-PEG-AuNP is effectively due to an interaction between PAP and BP.

Direct colorimetric detection of unamplified pathogen DNA by dextrin-capped gold nanoparticles

The interaction between gold nanoparticles (AuNPs) and nucleic acids has facilitated a variety of diagnostic applications, with further diversification of synthesis match bio-applications while reducing biotoxicity. However, DNA interactions with unique surface capping agents have not been fully defined. Using dextrin-capped AuNPs (d-AuNPs), we have developed a novel unamplified genomic DNA (gDNA) nanosensor, exploiting dispersion and aggregation characteristics of d-AuNPs, in the presence of gDNA, for sequence-specific detection. We demonstrate that d-AuNPs are stable in a five-fold greater salt concentration than citrate-capped AuNPs and the d-AuNPs were stabilized by single stranded DNA probe (ssDNAp). However, in the elevated salt concentrations of the DNA detection assay, the target reactions were surprisingly further stabilized by the formation of a ssDNAp-target gDNA complex. The results presented herein lead us to propose a mechanism whereby genomic ssDNA secondary structure formation during ssDNAp-to-target gDNA binding enables d-AuNP stabilization in elevated ionic environments. Using the assay described herein, we were successful in detecting as little as 2.94 fM of pathogen DNA, and using crude extractions of a pathogen matrix, as few as 18 spores/µL.

Method for colorimetric detection of double-stranded nucleic acid using leuco triphenylmethane dyes

Because loop-mediated isothermal amplification (LAMP) can amplify substantial amounts of DNA under isothermal conditions, its applications for simple genetic testing have attracted considerable attention. A positive LAMP reaction is indicated by the turbidity caused by by-products or by the color change after adding a metallochromic indicator to the reaction solution, but these methods have certain limitations. Leuco crystal violet (LCV), a colorless dye obtained after sodium sulfite treatment of crystal violet (CV), was used as a new colorimetric method for detecting LAMP. LCV is reconverted into CV through contact with double-stranded DNA (dsDNA). Therefore, the positive reaction of LAMP is indicated by color change from colorless to violet. The assay is sensitive enough to detect LAMP products, with a detection limit of 7.1ng/μl for dsDNA. It is also highly selective to dsDNA, and interference with single-stranded DNA and deoxynucleotide triphosphates (dNTPs) is not observed. LCV facilitates direct colorimetric detection of the main product rather than a by-product of the LAMP reaction; therefore, this method can be used under various reaction conditions such as those with added pyrophosphatase in solution. This colorimetric LAMP detection method using LCV is useful for point-of-care genetic testing given its simplicity.

Functionalized Gold Nanoparticles as an Approach to the Direct Colorimetric Detection of DCNP Nerve Agent Simulant

AbstractNew functionalized gold nanoparticles have been synthesized and their ability to act as colorimetric molecular probes for the naked‐eye detection of nerve agent simulant DCNP has been studied. The detection process is based on the compensation of charges at the surfaces of the functionalized AuNPs, which triggers their aggregation with the resulting change in the color of the solution.

Ultrasensitive optical detection of trinitrotoluene by ethylenediamine-capped gold nanoparticles

This study found that 1,2-ethylenediamine (EDA) as a primary amine could be modified onto the surface of citrate-stabilized gold nanoparticles (Au NPs), and the EDA-capped Au NPs were successfully used as an ultrasensitive optical probe for TNT detection. The strong donor–acceptor (D–A) interactions between EDA and trinitrotoluene (TNT) at the Au NP/solution interface induced significant aggregation of the EDA-capped Au NPs, and enabled to easily realize the direct colorimetric detection of ultratrace TNT. The results showed that such a color change was readily seen by the naked eye, and the colorimetric detection could be down to 400pM level of TNT with excellent discrimination against other nitro compounds. UV–vis absorption spectroscopy was used to examine the TNT-induced changes in local surface plasmon resonance (LSPR) of EDA-capped Au NPs, and a new LSPR band at ca. 630nm arose along with the addition of TNT, which produced a detection limit of TNT down to ca. 40pM. Furthermore, dynamic light scattering measurements evidenced the ultratrace TNT-induced small changes in the size of the EDA-capped Au NPs, and realized the quick and accurate detection of TNT in 0.4pM level. These results demonstrated the ultrahigh sensitivity of this optical probe for TNT detection. Moreover, this optical probe is sample, stable, low-cost, and these excellent properties make it quite promising for infield and rapid detection of TNT.

Direct Colorimetric Detection of Hydrogen Peroxide Using 4-Nitrophenyl Boronic Acid or Its Pinacol Ester

A colorimetric method for the direct determination of hydrogen peroxide in aqueous solution is described. H2O2 stoichiometrically converts 4-nitrophenyl boronic acid or 4-nitrophenyl boronic acid pinacol ester into 4-nitrophenol, which can be quantified by measuring the absorption at 400 nm in neutral or basic media. The reactions proceed fast under basic conditions and complete in 2 minutes to at pH 11 and 80?C. The linear range for the colorimetric method extends beyond 1.0 to 40 µM H2O2, and the limit of detection is ~1.0 µM H2O2. This method offers a convenient and practical process for rapid determination of hydrogen peroxide in aqueous media. Compared to many other techniques in H2O2 detection, this process is a direct measurement of H2O2, and is relatively unaffected by the presence of various salts, metal ions and the chelator EDTA.

Polydiacetylenes carrying amino groups for colorimetric detection and identification of anionic surfactants

A series of diacetylene lipids carrying primary amine, secondary amine or ammonium head groups are prepared and converted to blue polydiacetylene (PDA) sols by UV irradiation. The blue to pink colorimetric transition of polymerized diacetylenes in the presence of anionic surfactants such as sodium dodecanoate (SDC), sodium docecyl sulphate (SDS), and sodium dodecyl benzene sulphate (SDBS) are observed by the naked eye at the micromolar concentration level while there is no change with cationic surfactants and little response with nonionic surfactants. An identification of common anionic surfactants can be accomplished based on a combination of the colorimetric pattern of structurally diverse PDAs using the principal component analysis technique. Moreover, PDA was successfully fabricated on filter paper and the colorimetric response of PDA embedded on the paper was investigated which allowed the direct colorimetric detection of anionic surfactants.

A Water-Soluble Organometallic Conjugated Polyelectrolyte for the Direct Colorimetric Detection of Silver Ion in Aqueous Media with High Selectivity and Sensitivity

A water-soluble organometallic conjugated polyelectrolyte P1 and its corresponding model complex M1 based on an aspartic acid-substituted fluorene spacer are reported, which possess good water solubility as well as intriguing fluorescent and phosphorescent dual-emissive properties in a completely organic-free aqueous medium at room temperature. A new colorimetric silver ion sensor based on P1 is developed, which shows high selectivity and sensitivity for Ag+ ions in buffered water solution because of the Ag+-induced intersystem crossing from the singlet to triplet states. The obvious color change from colorless to yellow upon exposure to Ag+ ion is visible to the naked eyes and can be quantified colorimetrically by the visible absorption spectroscopic method. On the basis of the fluorescence intensity of P1 obtained in the fluorescence titration curves, a linear relationship is observed in the Stern−Volmer plot at low concentrations (1−5 μM), and the corresponding Stern−Volmer quenching constant (KSV) of ...

Direct colorimetric detection of copper(II) ions in sampling using diffusive gradients in thin-films

A novel binding phase was developed for use in diffusive gradients in thin-film (DGT) sampling for Cu(II) by employing methylthymol blue as a chelating and chromogenic agent. Methylthymol blue was adsorbed onto beads of Dowex 1 × 8 resin (200–400 mesh) and the resin beads were then immobilised onto an adhesive disc. Analysis of exposed binding discs by either UV–vis spectrophotometry or computer imaging densitometry provided robust quantification of adsorbed Cu(II) in the 0.2–1 μg cm −2 range, allowing detection at μg L −1 concentrations in the test solution (ca. 17 μg L −1 for a 24 h deployment), and in good agreement with established DGT theory. The method was shown to be a potential replacement for binding phases based on Chelex 100 where a colorimetric response to a specific metal is desired.