Sensitive Colorimetric Assay Research Articles

Au/Pd Nanocatalysts on Silica Nanoparticle-Coated Indium Tin Oxide for Colorimetric Sensing of Ascorbic Acid

Nanomaterials of gold and palladium have received research attention due to their excellent catalytic properties. Their recovery after the catalytic reaction still remains a challenge rendering the catalytic application of these nanomaterials for single use. Herein, we report a method of fabricating nanomaterials as thin nanoparticle (NP) films on catalytically inert surfaces, while allowing for their recovery and reuse is investigated. Indium tin oxide (ITO) glass slides were first hydroxylated (ITO-OH) and used to attach the silica NPs (SiO2NPs) to form ITO-SiO2NPs. The ITO-SiO2NPs surface was used as an adsorbent for gold and palladium mono- and bimetallic NPs (Au/PdNPs) to form ITO-SiO2-prS-Au/PdNPs, which was evaluated for the enzyme-like catalytic activity. The fabricated film with bimetallic NPs, ITO-SiO2-prS-Au/PdNPs, exhibited excellent peroxidase-like activity for the oxidation of 3,3′,5,5′-tetramethylbenzidine (TMB) with H2O2 when compared with monometallic thin NP films, ITO-SiO2-prS-AuNPs, and ITO-SiO2-prS-PdNPs. A typical Michaelis–Menten enzyme-like behavior of the bimetallic NP film, ITO-SiO2-prS-Au/PdNPs, was observed for H2O2 and TMB with Michaelis–Menten constants (Km) of 0.33 and 3.46 mM, respectively. A simple, sensitive, and selective colorimetric assay for ascorbic acid (AA) detection was fabricated based on the reduction effect of AA on the oxidized TMB. The fabricated AA colorimetric sensor exhibited a good linear range from 0.50 to 30.0 μM, with the limit of detection of 0.50 μM and the limit of quantification of 1.65 μM. Finally, our proposed colorimetric sensor, ITO-SiO2-prS-Au/PdNPs, successfully detected AA in new-born calf serum samples, as a real sample representative.

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.

Isolation of Bacteria Aptamers with Non-SELEX for the Development of a Highly Sensitive Colorimetric Assay Based on Dual Signal Amplification.

In this work, an aptamer against Escherichia coli is isolated via non-SELEX, which executes efficient selection by employing repetitive cycles of centrifugation-based partitioning, and the binding site of the aptamer on E. coli cell surfaces is inferred to be a membrane protein. Moreover, truncated sequence 2-17-2 with a higher affinity (Kd = 101.76 nM) is employed for highly sensitive colorimetric detection of bacteria based on the dual signal amplification strategy. When targets exist, the release of DNA 1 from the polymer activates a hybridization chain reaction (HCR) between DNA 1 and DNA 2, thereby inducing the aggregation of probe 1. Subsequently, DNA 3 dissociated from probe 1 as a linker DNA further assembles probe 2/3. In this system, two types of DNA@gold nanoparticles (AuNPs) coexist and successively aggregate AuNPs based on divergent triggering mechanisms. Under optimal conditions, the dual signal amplification strategy presents excellent sensitivity (10 CFU mL-1) and specificity, as well as the realization of real sample analysis.

A Simple and Highly Selective Colorimetric Sensing of Hg2+ Ions Based on Anti-Aggregation of Gold Nanoparticles

In this work, a highly sensitive and selective colorimetric sensor assay for the detection of Hg2+ ions is presented utilizing an anti-aggregation sensing approach with AuNPs and an amine-based sensor (S). The addition of sensor (S) to gold nanoparticles (AuNPs) leads to their aggregation and, thus, color variation from red to blue. Whereas, in the presence of Hg2+ ions, i.e., target analyte, sensor (S) preferably binds with Hg2+ ions over AuNPs, thus inhibiting their aggregation and subsequent color change. This approach is optimized in terms of various analytical parameters, such as incubation time, pH effect, and S titration, to obtain high sensitivity and selectivity. The UV–Visible studies validate the performance of developed approach for Hg2+ ions detection, and the limit of detection was calculated to be 17 nM, which is lesser than its permissible limit (30 nM) in drinking water. The interference studies also confirm the develop approach selectivity for Hg2+ ions even in the presence of coexisting ions in aqueous media. The experimental results were further correlated with the computational density functional theory (DFT) studies. The presented method is not only cost-effective but also eliminates tedious synthetic schemes and need of laborious manpower.

Catalytic Gold-Iridium Nanoparticles as Labels for Sensitive Colorimetric Lateral Flow Assay.

The colorimetric lateral flow assay (CLFA, also known as test strip) is a widely used point-of-care diagnostic technology. It has been a challenge to significantly improve the detection sensitivity of CLFA without involving additional equipment and/or compromising its simplicity. In this work, we break through the detection limit barrier of CLFA by developing a type of catalytic nanoparticles (NPs) used as labels. Specifically, the NPs were engineered by coating conventional gold NPs (AuNPs) with iridium (Ir) to form an Au-Ir core-shell structure. Such Au-Ir NPs possess ultrahigh peroxidase-like catalytic activities. A single Au-Ir NP can generate up to 107 colored molecules per second by catalyzing peroxidase substrates. The strong color signal from the catalysis ensures a high sensitivity of associated CLFA. The Au-Ir NP-based CLFA was successfully applied to the detection of two different cancer biomarkers that achieved limits of detection at the low picogram per milliliter level, hundreds of times lower than those of conventional AuNP-based CLFA.

Sensitive colorimetric assay of hydrogen peroxide and glucose in humoral samples based on the enhanced peroxidase-mimetic activity of NH2-MIL-88-derived FeS2@CN nanocomposites compared to its precursors.

By employing NH2-MIL-88 as a template, we synthesized the intermediate Fe@CN under high-temperature calcination and further fabricated the FeS2@CN nanocomposites in the presence of sulfur powder. Under varying temperatures (300-600 °C) and Fe@CN-to-S ratios (1:3-6), FeS2@CN500-5 nanocomposites had the highest peroxidase-mimetic activity. Under optimized conditions (incubation temperature 40 °C; solution pH 4.0 and nanocomposite concentration 10 μg/mL; 652-nm absorption), the Michaelis-Menten constant (Km) of FeS2@CN was much lower than that of horseradish peroxidase (HRP), therefore demonstrating that it had a higher affinity for both chromogenic substrates than conventional HRP. The limits of detection for H2O2 and glucose were 0.15 and 0.30 μmol/L, respectively, and the recoveries for glucose were 91.8-103% with RSDs <5.2%. The novelty of this study lies in (1) the FeS2@CN was confirmed to possess stronger enzyme-mimetic activity than its precursors (NH2-MIL-88 and Fe@CN); (2) the enhanced activity resulted from the unsaturated sites of N and S doping and the plentiful defects on the porous carbon surface; and (3) free radical trapping experiments evidenced that •OH played a major role in the catalytic reaction, while h+ and •O2- simultaneously participated in the catalytic process. These convincing performance metrics lead us to postulate that the FeS2@CN-based colorimetric biosensor provides a promising approach for several real-world applications, such as point-of-care diagnosis and workplace health evaluations.

DNA-Protein-Interaction (DPI)-ELISA Assay for PPAR-γ Receptor Binding.

Dysregulation of peroxisome proliferator-activated receptor (PPAR)-γ has been described in a plethora of pathological conditions, such as diabetes, obesity, inflammatory-related diseases, and cancer. Therefore, identifying novel drugs that are able to restore PPAR-γ activity is a current challenge, which is however slowed down by the lack of a rapid and reproducible activity assay. To date, only a few methods are able to characterize PPAR-γ activity and most of them are expensive, time-consuming, and not always quantitative.Herein, we presented a sensitive multi-well colorimetric assay, termed DNA-Protein-Interaction enzyme-linked immunosorbent assay (DPI-ELISA). This method is based on the ELISA principle, except that it allows to detect only activated PPAR-γ because, unlike classical ELISA, PPAR-γ is not captured by an antibody but by a double-stranded oligonucleotide probe containing its peroxisome proliferator response elements (PPRE) consensus sequence. Thus, DPI-ELISA represents a useful assay for PPAR-γ studies, as well as for the identification of novel PPAR-γ ligands for the development of innovative therapeutic approaches to human diseases where PPAR-γ signaling is dysregulated.

Colorimetric assay based on peroxidase-like activity of dodecyl trimethylammonium bromide-tetramethyl zinc (4-pyridinyl) porphyrin for detection of organophosphorus pesticides.

A simple and sensitive colorimetric assay for detecting organophosphorus pesticides (OPs) was developed based on 3,3',5,5'-tetramethylbenzidine (TMB)/hydrogen peroxide (H2O2)/dodecyl trimethylammonium bromide (DTAB)-tetramethyl zinc (4-pyridinyl) porphyrin (ZnTPyP). In this system, based on the peroxidase-like activity of DTAB-ZnTPyP, H2O2 decomposes to produce hydroxyl radicals, which oxidize TMB, resulting in blue oxidation products. The OPs (trichlorfon, dichlorvos, and thimet) were first combined with DTAB-ZnTPyP through electrostatic interactions. The OPs caused a decrease in the peroxidase-like activity of DTAB-ZnTPyP due to spatial site blocking. At the same time, π-interactions occurred between them, and these interactions also inhibited the oxidation of TMB (652 nm), thus making the detection of OPs possible. The limits of detection for trichlorfon, dichlorvos, and thimet were 0.25, 1.02, and 0.66 μg/L, respectively, and the corresponding linear ranges were 1-35, 5-45, and 1-40 μg/L, respectively. Moreover, the assay was successfully used to determine OPs in cabbage, apple, soil, and traditional Chinese medicine samples (the recovery ratios were 91.8-109.8%), showing a great promising potential for detecting OPs also in other complex samples.

Analysis of Stable Chelate-free Gadolinium Loaded Titanium Dioxide Nanoparticles for MRI-Guided Radionuclide Stimulated Cancer Treatment.

Recent studies demonstrate that titanium dioxide nanoparticles (TiO2 NPs) are an effective source of reactive oxygen species (ROS) for photodynamic therapy and radionuclide stimulated dynamic therapy (RaST). Unfortunately tracking the in vivo distribution of TiO2 NPs noninvasively remains elusive. Given the use of gadolinium (Gd) chelates as effective contrast agents for magnetic resonance imaging (MRI), this study aims to (1) develop hybrid TiO2-Gd NPs that exhibit high relaxivity for tracking the NPs without loss of ROS generating capacity; and (2) establish a simple colorimetric assay for quantifying Gd loading and stability. A chelate-free, heat-induced method was used to load Gd onto TiO2 NPs, which was coated with transferrin (Tf). A sensitive colorimetric assay and inductively coupled plasma mass spectrometry (ICP-MS) were used to determine Gd loading and stability of the TiO2-Gd-Tf NPs. Measurement of the relaxivity was performed on a 1.4 T relaxometer and a 4.7 T small animal magnetic resonance scanner to estimate the effects of magnetic field strength. ROS was quantified by activated dichlorodihydrofluorescein diacetate fluorescence. Cell uptake of the NPs and RaST were monitored by fluorescence microscopy. Both 3 T and 4.7 T scanners were used to image the in vivo distribution of intravenously injected NPs in tumor-bearing mice. A simple colorimetric assay accurately determined both the loading and stability of the NPs compared with the expensive and complex ICP-MS method. Coating of the TiO2-Gd NPs with Tf stabilized the nanoconstruct and minimized aggregation. The TiO2-Gd-Tf maintained ROS-generating capability without inducing cell death at a wide range of concentrations but induced significant cell death under RaST conditions in the presence of F-18 radiolabeled 2-fluorodeoxyglucose. The longitudinal (r1 = 10.43 mM-1s-1) and transverse (r2 = 13.43 mM-1s-1) relaxivity of TiO2-Gd-Tf NPs were about twice and thrice, respectively, those of clinically used Gd contrast agent (Gd-DTPA; r1 = 3.77 mM-1s-1 and r2 = 5.51 mM-1s-1) at 1.4 T. While the r1 (8.13 mM-1s-1) reduced to about twice that of Gd-DTPA (4.89 mM-1s-1) at 4.7 T, the corresponding r2 (87.15 mM-1s-1) increased by a factor 22.6 compared to Gd-DTPA (r2 = 3.85). MRI of tumor-bearing mice injected with TiO2-Gd-Tf NPs tracked the NPs distribution and accumulation in tumors. This work demonstrates that Arsenazo III colorimetric assay can substitute ICP-MS for determining the loading and stability of Gd-doped TiO2 NPs. The new nanoconstruct enabled RaST effect in cells, exhibited high relaxivity, and enhanced MRI contrast in tumors in vivo, paving the way for in vivo MRI-guided RaST.

Non-protein coding RNA sequences mediate specific colorimetric detection of Staphylococcus aureus on unmodified gold nanoparticles

Nonprotein coding RNA (npcRNA) is a transcribed gene sequence that is not able to translate into protein, yet it executes a specific function in modulation and regulation mechanisms. As npcRNA is highly resistant to the mutation, the Sau-02 npcRNA gene and its probe oligonucleotide, which are specifically present in Staphylococcus aureus and in methicillin-resistant S. aureus only, used to develop a highly specific and sensitive colorimetric assay on unmodified gold nanoparticles (AuNPs). Hybridization between the npcRNA Sau-02 gene sequences was detected through noncrosslinking AuNP aggregation in salt solution in the presence of probe-target gene sequences. AuNPs of 10 and 15 nm in sizes with monovalent ion salt (NaCl) solution were optimized as the ideal tool for investigating the stability of AuNPs upon the addition of gene sequences. The state dispersed and aggregated forms of 10 nm AuNPs with the presented colorimetric assay were justified through field emission scanning electron microscopy and atomic force microscopy. The particle distribution of two different AuNP states was evaluated through particle distribution analysis. The lowest detection amount of S. aureus npcRNA from the colorimetric assay performed was 6 pg/µL, as the color of AuNPs turned blue with the presence of probe oligonucleotides and target gene sequences.

Target-induced recycling assembly of split aptamer fragments by DNA toehold-mediated displacement for the amplified colorimetric detection of estradiol

Estradiol is an endogenous hormone that affects many physiological functions in humans; thus, the demand for its detection has been increasing. Gold nanoparticle (AuNP)-based colorimetric sensors with split aptamers offer an easy-to-operate detection method for estradiol. However, the assay performance is still unmet. Here, we report a simple and sensitive colorimetric assay for the detection of estradiol, based on the target-induced recycling assembly of split aptamer fragments. This method contains three DNA fragments that remain in a metastable state without a target. After adding estradiol, the split aptamers assemble with the target, which can be regenerated by helper DNA, resulting in the formation of a three-way junction (3WJ)-like structure and the recycling of recognition. The generation of the 3WJ-like DNA structure causes the formation of unstable AuNPs, enhancing the salt-induced aggregation of AuNPs. The assay shows a detection limit of 0.7 nM, which is better than those of other aptamer-based detection methods. Moreover, the assay enables the detection of estradiol spiked in urine by avoiding any interference from contaminants. The colorimetric assay includes anti-estradiol DNA split aptamers, accounting for no false responses to non-target molecules. Therefore, the results demonstrated that it has application potential for the detection of estradiol in a complex matrix.

Highly sensitive visual colorimetric sensor for xanthine oxidase detection by using MnO2-nanosheet-modified gold nanoparticles

In this study, a highly sensitive colorimetric assay has been constructed for the determination of xanthine oxidase (XOD) activity by the GNP@MnO2 core-shell nanoparticles as probe. In the presence of XOD, xanthine can be oxidized to produce H2O2, which makes the MnO2 shell fallen off. With the single particle detection (SPD) based dark field microscopy (DFM), the scattering color of GNP@MnO2 NP probe shows obvious change before and after etching process. At the single particle level, noticeable color change of the single probe can be easily detected in the existence of trace XOD. This SPD-based colorimetric strategy displays broad linear dynamic range (0.02–4 mU/mL) and low detection limit of 7.82 μU/mL, which is more sensitive than the results from ensemble sample measurement. In addition, we tested the inhibitory effect of quercetin on the activity of XOD and obtained good inhibition effect. As a consequence, this SPD-based colorimetric strategy provides new perception for the ultrasensitive detection of molecules in complex system.

Acoustofluidics-manipulated triple-emission fluorescent nanoprobe aggregates with multicolor-variation for colorimetric quantitative assay

Acoustofluidics-assisted colorimetric method, a promising point-of-care test (POCT) strategy, has been applied in convenient detection of target molecules using the sensing nanomaterials, but remains a formidable challenge to design fluorescent probe with profuse color evolution, realizing sensitive, simple and accurate determination of analyte. Here, a novel triple-emission fluorescent probe is constructed through a simple blend of two ratiometric fluorescent probes (blue-organosilane-polymerized carbon dots@SiO2 nanoparticles@red-CdTe/CdS quantum dots (b@SiO2@r) and blue-organosilane-polymerized carbon dots@SiO2 nanoparticles@green-CdTe/CdS quantum dots (b@SiO2@g)) at the optimized volume ratio of 11:8. As signal reporters of the triple-emission probe, outer modified “r” and “g” will be quenched in sequence by target via two-step response strategy while the inner “b” as internal standard remains constant. When acoustic field is applied, the effect of acoustofluidics-based nanoparticles concentration makes triple-emission probe aggregate completely, accompanying fluorescence color signal being enhanced, sensitivity of colorimetric assay being improved and multicolor-variation (from orange to dark-goldenrod to dark-olive-green to sea-green to dark-cyan to final steel-blue) with the increase of analyte amount. For POCT demonstration, hemoglobin (Hb) is used as a model target and the captured fluorescence photos of triple-emission probe aggregates are analyzed by a chromaticity analysis application on the smartphone to calculate the red, green and blue values, which are used for accurate Hb quantification with the limit of detection of 1.99 mg/L. Whole test is finished within ∼12 min. The acoustofluidics-manipulated triple-emission probe biosensing strategy has also been applied to the rapid and visual quantitative detection of Hb in human blood.

Arsenic speciation analysis in porewater by a novel colorimetric assay

Arsenic is common toxic contaminant, but tracking its mobility through submerged soils is difficult because microscale processes dictate its speciation and affinity to minerals. Analyses on environmental dissolved arsenic (As) species such as arsenate and arsenite currently require highly specialized equipment and large sample volumes. In an effort to unravel arsenic dynamics in sedimentary porewater, a novel, highly sensitive, and field-usable colorimetric assay requiring 100 μL of sample was developed. Two complementary protocols are presented, suitable for sub-micromolar and micromolar ranges. Phosphate is a main interfering substance, but can be separated by measuring phosphate and arsenate under two different acidities. Arsenite is assessed by oxidation of arsenite to arsenate in the low-acidity reagent. Optimization of the protocol and spectral analyses resulted in elimination of various interferences (silicate, iron, sulfide, sulfate), and the assay is applicable across a wide range of salinities and porewater compositions. The new assay was used to study As mobilization processes through the soil of a contaminated brook. Water column sources of arsenic were limited to a modest input by a groundwater source along the flow path. In one of the sites, the arsenite and arsenate porewater profiles showed active iron-driven As redox cycling in the soil, which may play a role in arsenic mobilization and releases arsenite and arsenate into the brook water column. Low arsenic concentrations downstream from the source sites indicated arsenic retention by soil and dilution with additional sources of water. Arsenic is thus retained by the Bossegraben before it merges with larger rivers.

Colorimetric detection of hydrogen peroxide and cholesterol using Fe3O4-brominated graphene nanocomposite.

Fe3O4-brominated graphene (Fe3O4-GBR) nanocomposites were synthesized via an in situ method using the precursors FeSO4.7H2O and GBR in different (1:1, 1:2, 2:1, 1:5, 1:10, 1:20, and 5:1) weight ratios at pH11.5. The Fe3O4-GBR (1:5) nanocomposite in combination with H2O2 and 3,3',5,5'-tetramethylbenzidine (TMB) showed swift and superior intrinsic peroxidase mimetic enzyme activity compared with the other Fe3O4-GBR composites, GBR and Fe3O4, as observed by colorimetry. It was characterized using high-resolution scanning electron microscopy (HRSEM), energy dispersive X-ray spectroscopy (EDX), Fourier transform infrared (FTIR) spectroscopy, powder X-ray diffraction (PXRD), and thermogravimetric analysis (TGA). Its catalytic activity was optimized by varying different parameters, and the optimum conditions for peroxidase mimetic activity were observed using 100μL Fe3O4-GBR (1mg/mL), 50μL TMB (1mg/mL), and 200μL H2O2(1mM) in 400μL of acetate buffer of pH2.3 at 30°C temperature. Kinetic analysis has revealed the Michaelis-Menten kinetic behavior of peroxidase activity with Michaelis-Menten constants (Km) and maximum initial velocities (Vmax) of 0.082mM and 14.1 nMs-1 respectively, for H2O2 and 0.086mM and 5.1 nMs-1, respectively for TMB. The limit of detection and linear range were found to be 49.6μM and 100-880μM, respectively, for H2O2 and 41.9μM and 47.6-952.3μM, respectively, for cholesterol. On this basis, a simple, swift, sensitive, selective, and reproducible colorimetric assay to detect cholesterol levels in blood serum samples using Fe3O4-GBR nanocomposite has been developed. Thus, Fe3O4-GBR composite as compared to Fe3O4 and GBR has shown better peroxidase mimicking activity for biosensing.

A naked-eye colorimetric assay for detection of Hg2+ ions in real water samples based on gold nanoparticles-catalyzed clock reaction

Designing a simple, inexpensive, and sensitive colorimetric sensor for the detection of toxic species in drinking water is of great importance. In this paper, we have demonstrated a simple and sensitive colorimetric assay for naked-eye detection of mercury ions (Hg2+) in water based on gold nanoparticles-catalyzed clock reaction. Citrate-capped Au nanoparticles (AuNPs) were used to catalyze the reduction of methylene blue (MB) to leucomethylene blue (LMB) by hydrazine. However, Hg2+ ions can reduce the catalytic activity of AuNPs in this clock reaction, resulting in the longer reaction time for color transformation of MB from blue to colorless LMB solution. Based on this principle, we have developed an ultrasensitive colorimetric assay that can selectively detect Hg2+ in the concentration range of 5 to 1000 nM with a limit of detection (LOD) of 4.3 nM, which is well below the maximum allowable concentration for Hg2+ in drinking water (10 nM) as defined by the United States Environmental Protection Agency. Owing to the rapidity and simplicity of the naked eye detection process, we expect that this AuNPs-catalyzed clock reaction could put forward an insight for the detection of toxic Hg2+ ions in real water samples, opening up new avenues for developing cost-effective and portable nanosensors for environmental applications.

Using activated charcoal to remove substances interfering with the colorimetric assay of inorganic phosphate in plant extracts

AimOrganic substances in leaves of several southwest Australian native species interfere with sensitive colorimetric assays and prevent quantification of inorganic phosphate concentration ([Pi]). We aimed to develop a reproducible routine procedure for treating leaf extracts with activated charcoal (AC) to remove interfering substances, allowing the determination of [Pi] by the malachite green spectrophotometric assay.MethodsLeaf extracts of native plants from southwest Australia in 1% (v/v) acetic acid were treated with 10 mg mL−1 acid-washed AC for removal of interfering substances. Standard solutions (0 to 18 μM Pi) with and without AC treatment were compared to quantify Pi loss. A spiking and recovery test was performed to validate the AC treatment.ResultsLeaf extracts treated with AC exhibited distinguishable absorbance peaks for the malachite green-orthophosphate complex between 630 and 650 nm, as opposed to untreated samples. The Pi-adsorption by AC represented a relatively larger fraction of [Pi] in solutions at 0–4 μM Pi range and stabilised at higher [Pi] when maximum adsorption capacity of AC reached at 11.7 μg Pi g−1AC. The Pi recovery after AC treatment in spiked samples ranged between 100 and 111%.ConclusionThe AC treatment successfully removed interfering substances from samples but caused Pi loss. Thus, quantification of [Pi] in AC-treated extracts requires sample [Pi] ≥ 6 μM Pi and the use of AC-treated standards. The error of the AC treatment was minor compared with environmental variability of leaf [Pi]. The AC treatment was a reproducible time- and cost-effective method to remove interfering substances from leaf extracts.

Varnish polish functionalized paper as a new solid platform for colorimetric determination of copper (II) in water

ABSTRACT In the current strategy, a highly sensitive and fast one-step colorimetric assay for trace determination of Cu2+ in water is presented. The planned assay was based upon the use of varnish polish-modified filter paper printed with 2-[5-(2-hydroxy-5-sulfophenyl)-3-phenyl-1-formazyl] benzoic acid monosodium salt, commercially named zincon. Under optimal conditions, the designed assay readily recognised the formation of blue–purple-coloured Cu–ZI chelate in less than 10 min. The proposed probe was able to detect Cu2+ in the concentration range from 0.04 to 10 μg/mL (6.34 × 10−1 to 1.59 × 103µM), with low limit of detection (LOD) of 0.01 μg/mL (1.59 × 10−1 µM). The LOD was 10-fold lower than the allowable level for Cu2+ in drinking water set by WHO legislation. The assay was applied for Cu2+determination in water with acceptable recoveries (89.6–102.1%) and precision (<3%). Therefore, the proposed sensing assay might be considered as a potential rapid and low-cost candidate for total determination and speciation of Cu2+in water samples.

BSA-stabilized manganese phosphate nanoflower with enhanced nanozyme activity for highly sensitive and rapid detection of glutathione

The development of an efficient protein-inorganic nanohybrid with superior nanozyme activity for highly sensitive detection of glutathione (GSH) is essential for early diagnosis of human diseases. Herein, a rapid and highly sensitive colorimetric assay using self-assembled bovine serum albumin-hydrated manganese phosphate nanoflowers (MnPNF) as a biomimic oxidase is developed for GSH detection in human serum. The BSA can complex with Mn2+ to serve the nucleation center to produce MnPNF in the presence of phosphate-buffered saline (PBS). The morphology and surface characterization results show that the MnPNF is assembled with hierarchical nanoplates to form 500 nm nanoflowers. The oxidase-like activity of MnPNF is based on the redox reaction with 3,3′,5,5′-tetramethylbenzidine. However, the addition of GSH can reduce MnPNF to Mn2+, and subsequently supresses the oxidase-like activity and a yellow color at 450 nm is observed in the presence of H2SO4. The MnPNF-based nanozyme exhibits excellent sensing ability toward GSH detection, and a good linear relationship between the change in absorbance at 450 nm and the added amounts of GSH at 50 nM–10 μM with low limits of detection of 20 and 26.6 nM in the PBS and diluted human serum, respectively, is observed. Moreover, the sensing probe shows a superior selectivity over the other 16 interferences, which drive the determination of GSH feasible in real human serum. Since the MnPNF can be simply prepared at room temperature and no functionalization is required, this assay can be used to design the highly efficient biomimic oxidase for effective sensing of GSH and other disease-related biomolecules in biological fluid samples.

Antibody conjugated magnetic nanoparticle based colorimetric assay for the detection and quantification of aflatoxin B1 in wheat grains

Aflatoxin B1 (AFB1) is a most potent carcinogenic secondary metabolite produced byAspergillus flavus. As a food safety concern, development of a rapid, cost effective, sensitive and easy to use method for the detection of aflatoxin is of prime requirement. In this study, AFB1 was conjugated with bovine serum albumin (BSA), and AFB1-BSA conjugate was purified by HPLC. Purification was confirmed by UV-Vis spectroscopy, FTIR and MALDI-TOF mass spectrometry. The polyclonal antibody was raised against AFB1-BSA conjugate in rabbit and purified by protein A sepharose and BSA sepharose affinity columns. Iron oxide nanoparticles (MNPs) were synthesised by co-precipitation method and their surface was functionalised with (3-aminopropyl) triethoxysilane (APTES). Size of APTES conjugated MNPs was determined by electron microscopy, and characterised by several biophysical techniques. The purified anti-AFB1 antibody was conjugated with surface functionalised MNPs and the conjugation was confirmed by determining the sizes of free and antibody conjugated MNPs by field emission scanning electron microscope where increase of particle sizes from 10-20 to 40-50 nm was observed due to antibody conjugation. Anti-AFB1 antibody conjugated MNPs were used for capturing AFB1 from the aflatoxin spiked wheat grains with a recovery percentage of more than 80% and used effectively five times. The captured AFB1 was then quantified by a sensitive colorimetric assay where colourless AFB1 was first converted into coumaric acid by NaOH. Subsequently, coumaric acid reacted with 2,6-dibromoquinone-4-chloroimide (DBQC) to a green-coloured indophenol product which was quantified spectrophotometrically. AFB1 contamination as low as 2 μg/kg in wheat grains was detected by the developed technique suggesting its potential application for both qualitative and quantitative analysis of aflatoxins present in feed and food materials.