Signal on-off ratiometric electrochemical sensor coupled with a molecularly imprinted polymer for the detection of carbendazim.

A stable ratiometric electrochemical sensor is introduced for the selective detection of carbendazim (CBD). Specifically, the proposed sensor employs a Co@Mo2C bimetallic nanomaterial as the glassy carbon electrode substrate and a layer of molecularly imprinted polymer (MIP) was in situ fabricated on glassy carbon electrode by electropolymerization, with o-aminophenol as the functional monomer and CBD acting as template. A ratiometric MIP sensor was constructed by adding ferrocene (Fc) internal reference directly to the sample solution. The bimetallic nanomaterials provide a large loading platform for the MIP layer through synergistic effects, amplifying the signal. Excellent CBD binding selectivity is achieved by the templating effect of the three-dimensional (3D) MIP layer. The internal standard is added directly to the electrolyte solution to be tested, allowing the new type of ratiometric electrochemical sensor to avoid the cumbersome steps of other methods and reducing the difficulty and human error of the experimental procedure. Combining a ratiometric strategy with a 3D MIP structure realises the dual-signal detection of CBD. The optimised sensor showed an excellent linear relationship between 0.01 and 1 000 µM, with a correlation coefficient of 0.997 and a detection limit of 3.4 nM (S/N = 3).

A high sensitivity electrochemical sensor based on a dual-template molecularly imprinted polymer for simultaneous determination of clenbuterol hydrochloride and ractopamine.

A nitrogen-doped Fe-MOF shows a high specific surface area and excellent electrical conductivity after high temperature carbonization. A novel electrochemical sensor based on a N@Fe-MOF@C loaded dual-template molecularly imprinted polymer (DTMIP) modified glassy carbon electrode (GCE) was proposed for the rapid and ultra-sensitive simultaneous detection of clenbuterol hydrochloride (CLB) and ractopamine (RAC). N@Fe-MOF@C combined with a MIP significantly enhanced the electrical signal. Cyclic voltammetry (CV) was used to detect CLB and RAC. The electrochemical polymerization was conducted with O-phenylenediamine as the functional monomer and CLB and RAC as template molecules. The factors affecting the sensor response were optimized. Under the optimal experimental conditions, the CV current response showed a linear range of 10-12-8 × 10-9 M for both CLB and RAC, and the detection limit (LOD) for both CLB and RAC was 3.03 × 10-13 M (S/N = 3). This electrochemical sensing system has high sensitivity, selectivity, excellent reproducibility, repeatability and stability. The recoveries of the actual samples (97.4%-101.2%) and reasonable relative standard deviations (RSDs) (1.06%-3.17%) indicate the practicability of the sensor system. The system has high application value in the rapid detection of CLB and RAC in clenbuterol hydrochloride tablets, human urine and raw pork.

GlnR and PhoP regulate ß-glucosidases involved in cellulose digestion in response to nitrogen and phosphate availability.

The limited catalytic efficiency of cellulose-degrading enzymes restricts cellulose digestion. We investigated the transcriptional regulation of genes encoding key cellulose degrading enzymes, namely ß-glucosidases, in the industrial actinobacterium Saccharopolyspora erythraea. We observed that the expression of most ß-glucosidase-encoding genes was controlled by the availability of nitrogen and phosphate via their respective global regulators, namely GlnR and PhoP. Electrophoretic mobility shift assay demonstrated that GlnR and PhoP bound directly to the promoters of ß-glucosidase-encoding genes. Deletion of glnR resulted in lower transcript levels and activity of ß-glucosidases, leading to decreased bacterial growth on cellulose. Overexpression of glnR and phoP or nitrogen/phosphate starvation increased the transcript levels and total activity of ß-glucosidases. Moreover, GlnR/PhoP-mediated cellobiose utilization was also observed in Streptomyces coelicolor A3(2). These findings provide insights into the regulatory roles played by GlnR and PhoP in coordinating nitrogen/phosphate metabolism and carbohydrate utilization, and indicate potential strategies for cellulose fermentation in the production of bio-based chemicals by actinobacteria.

Efficient production and characterization of homopolymeric poly(3-hydroxyvalerate) produced by Bacillus strain PJC48.

Aliphatic polyester, poly(3-hydroxyvalerate) (PHV), is commonly produced as a granular component in bacterial cells of various species. Based on 16S rDNA gene sequence analysis, strain PJC48 was identified as a Bacillus species. The current study is aimed to screen for a high-yield strain that can produce PHV efficiently and to increase PHV product yield by optimizing the fermentative process. We identified a high-producer strain based on Nile red staining. Characterization of the PHV produced by PJC48 by nuclear magnetic resonance spectroscopy revealed that it consisted of (R)-3-hydroxyvalerate monomers. The suggested model was validated by response surface methodology. Optimization of the PHV yield resulted in an increase of 32.75% compared to control, with a maximum production of 1.64 g/L after 48 H.

Electrochemical sensor based on molecularly imprinted polymer for sensitive and selective determination of metronidazole via two different approaches.

A molecularly imprinted polymer decorated glassy carbon electrode (MIP/GCE) is facilely developed into an electrochemical sensing platform for detection of metronidazole (MNZ). MIP preparation was carried out via in situ electropolymerization and o-phenylenediamine was selected as the optimal functional monomer. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were employed to characterize and assess the performance of the so-obtained sensor. In particular, two assay methods, which are based on different principles, were involved in the detection procedure. One is based on MIP/catalysis (Method І) and the other is MIP/gate effect (Method II). Comparison of these two methods was made in the aspects including detection range, sensitivity, accuracy, selectivity, repeatability, and long-term stability. It is found that Method І affords a lower detection limit of 3.33 × 10(-10) M (S/N = 3) while the detection limit of Method II is 6.67 × 10(-10) M (S/N = 3). The linear range of Method І and II is 1.0 × 10(-9) to 1.0 × 10(-8) M and 2.0 × 10(-9) to 1.0 × 10(-7) M, respectively. The MIP/GCE exhibits good recognition ability towards the template molecule-MNZ in the presence of the analogues of MNZ and other interferents, which can be ascribed to the successful imprinting effect during MIP membrane preparation. Graphical Abstract Procedure for fabricating MIP/GCE and its application in detecting metronidazole in serum.

Preparation and characterization of erythromycin molecularly imprinted polymers based on distillation-precipitation polymerization.

Erythromycin-imprinted polymers with excellent recognition properties were prepared by an innovative strategy called distillation-precipitation polymerization. The interaction between erythromycin and methacrylic acid was studied by ultraviolet absorption spectroscopy, and the as-prepared materials were characterized by Fourier-transform infrared spectroscopy and scanning electron microscopy. Moreover, their binding performances were evaluated in detail by static, kinetic and selective sorption tests. It was found that the molecularly imprinted polymers afforded good morphology, monodispersity, and high adsorption capacity when the fraction of the monomers was 7 vol% in the whole reaction system, and the adsorption data for imprinted polymers correlated well with the Langmuir model. The maximum capacity of the imprinted and the non-imprinted polymers for adsorbing erythromycin is 44.03 and 19.95 mg/g, respectively. The kinetic studies revealed that the adsorption process fitted a pseudo-second-order kinetic model. Furthermore, the imprinted polymers display higher affinity toward erythromycin, compared with its analogue roxithromycin.

A novel electrochemical sensor based on molecularly imprinted polymer-modified C-ZIF67@Ni for highly sensitive and selective determination of carbendazim.

In this work, we propose a two-step coating method, combining C-ZIF67@Ni with molecular imprinting polymer (MIP), to develop a high-sensitivity and high-selectivity Carbendazim (CBD) electrochemical sensor. ZIF67@Ni was prepared by a simple chemical bath method, and C-ZIF67@Ni was obtained by high-temperature carbonization of ZIF67@Ni. Then, MIP layer was prepared by electrochemical in-situ polymerization, with O-aminophenol as functional monomers, CBD acting as template on the surface of the C-ZIF67@Ni-modified glassy carbon electrode (GCE). During the preparation process, the types of functional monomers, the polymerization solution pH, the ratio of functional monomers to template molecules, and the incubation time are optimized. The morphological characteristics, composition information and electrochemical properties of MIP/C-ZIF67@Ni/GCE were investigated in detail under optimal conditions. Physical characterization and electrochemical tests revealed that ZIF67@Ni significantly improves the electron transmission capacity and surface area of the sensor after high-temperature carbonization. C-ZIF67@ Ni has a good synergistic effect on MIP, allowing rapid and specific identification of the test substance. MIP/C-ZIF67@Ni/GCE showed a good linear relationship with CBD in the concentration range from 4 × 10-13 M to 1 × 10-9 M, the lowest detection limit was 1.35 × 10-13 M (S/N = 3) R2 = 0.9983 and RSD = 2.34. Additionally, the sensor showed good repeatability, stability, and selectivity, and can be used for the detection of carbendazim in soil and water with a recovery of 98% above.

Construction of a p-coumaric and ferulic acid auto-regulatory system in Pseudomonas putida KT2440 for protocatechuate production from lignin-derived aromatics.

Lignin is a robust and underutilized aromatic heteropolymer on earth. The effective valorization of lignin into value-added products is an attractive research topic in lignocellulosic biorefineries. However, a low bioconversion rate, high process cost, low yield, and high toxicity of substrates hinder its further applications. In this study, an auto-regulatory system was developed and identified as an effective solution to diminish these challenging bottlenecks. First, a lignin-derived standard model aromatic p-coumaric acid (p-CA)-responsive biosensor was successfully developed through a series of rational engineering approaches in Pseudomonas putida KT2440. Furthermore, an auto-regulatory system was established, which rapidly coexpressed key rate-limiting enzymes, 4-hydroxybenzoate hydroxylase, vanillate-O-demethylase, and transporter HcnK under the biosensor element, to convert the mixture of p-CA and ferulic acid into a value-added platform chemical protocatechuate with a titer of 12.7 g/L. This study demonstrated that the constructed auto-regulatory platform is effective and economical for lignin valorization.

Multiplexed bead-based mesofluidic system for gene diagnosis and genotyping.

We have developed a novel multiplexed bead-based mesofluidic system (MBMS) based on the specific recognition events on the surface of a series of microbeads (diameter 250 µm) arranged in polydimethylsiloxane (PDMS) microchannels (diameter 300 µm) with the predetermined order and assembled an apparatus implementing automatically the high-throughput bead-based assay and further demonstrated its feasibility and flexibility of gene diagnosis and genotyping, such as ß-thalassemia mutation detection and HLA-DQA genotyping. The apparatus, consisting of bead-based mesofluidic PDMS chip, liquid-processing module, and fluorescence detection module, can integrate the procedure of automated-sampling, hybridization reactions, washing, and in situ fluorescence detection. The results revealed that MBMS is fast, has high sensitivity, and can be automated to carry out parallel and multiplexed genotyping and has the potential to open up new routes to flexible, high-throughput approaches for bioanalysis.

Multicomponent mesofluidic system for the detection of veterinary drug residues based on competitive immunoassay.

An automated multicomponent mesofluidic system (MCMS) based on biorecognitions carried out on meso-scale glass beads in polydimethylsiloxane (PDMS) channels was developed. The constructed MCMS consisted of five modules: a bead introduction module, a bioreaction module, a solution handling module, a liquid driving module, and a signal collection module. The integration of these modules enables the assay to be automated and reduces it to a one-step protocol. The MCMS has successfully been applied toward the detection of veterinary drug residues in animal-derived foods. The drug antigen-coated beads (varphi250 microm) were arrayed in the PDMS channels (varphi300 microm). The competitive immunoassay was then carried out on the surface of the glass beads. After washing, the Cy3-labeled secondary antibody was introduced to probe the antigen-antibody complex anchored to the beads. The fluorescence intensity of each bead was measured and used to determine the residual drug concentration. The MCMS is highly sensitive, with its detection limits ranging from 0.02 (salbutamol) to 3.5 microg/L (sulfamethazine), and has a short assay time of 45 min or less. The experimental results demonstrate that the MCMS proves to be an economic, efficient, and sensitive platform for multicomponent detection of compound residues for contamination in foods or the environment.

A robust electrochemical sensing of molecularly imprinted polymer prepared by using bifunctional monomer and its application in detection of cypermethrin.

This work describes a hybrid electrochemical sensor for highly sensitive detection of pesticide cypermethrin (CYP). Firstly, Ag and N co-doped zinc oxide (Ag-N@ZnO) was produced by sol-gel method, and then Ag-N@ZnO was ultrasonically supported on activated carbon prepared from coconut husk (Ag-N@ZnO/CHAC). Finally, a layer of molecularly imprinted polymer (MIP) was in situ fabricated on glassy carbon electrode by electro-polymerization, with dopamine and resorcinol as dual functional monomers (DM), CYP acting as template (DM-MIP-Ag-N@ZnO/CHAC). Morphological features, composition information and electrochemical properties of DM-MIP-Ag-N@ZnO/CHAC were investigated in detail. It is worth to mention that for the first time response surface method was used to investigate the effect of double monomers and to optimize the ratio between template and monomers. Compared with typical one-monomer involving MIP, the MIP prepared with dual functional monomers (DMMIP) of monomers showed higher response and better selectivity. Under the optimal conditions, a calibration curve of current shift versus concentration of CYP was obtained in the range of 2 × 10-13~8 × 10-9 M, and the developed sensor gave a remarkably low detection limit (LOD) of 6.7 × 10-14 M (S/N = 3). Determination of CYP in real samples was conducted quickly and accurately with our sensor. The DMMIP-Ag-N@ZnO/CHAC electrochemical sensor proposed in this paper has great potential in food safety, drug residue determination and environmental monitoring.

A robust electrochemical sensing platform using carbon paste electrode modified with molecularly imprinted microsphere and its application on methyl parathion detection.

A highly sensitive electrochemical sensor using a carbon paste electrode (CPE) modified with surface molecularly imprinted polymeric microspheres (SMIPMs) was developed for methyl parathion (MP) detection. Molecular imprinting technique based on distillation precipitation polymerization was applied to prepare SMIPMs and non-surface imprinted microspheres (MIPMs). The polymer properties including morphology, size distribution, BET specific surface area and adsorption performance were investigated and compared carefully. Both MIPMs and SMIPMs were adopted to prepare CPE sensors and their electrochemical behaviors were characterized via cyclic voltammetry and electrochemical impedance spectroscopy. Compared with MIPMs packed sensor, SMIPMs/CPE exhibits a higher sensing response towards MP with linear detection range of 1 × 10-12-8 × 10-9 mol L-1 and detection limit of 3.4 × 10-13 mol L-1 (S/N = 3). Moreover, SMIPMs/CPE exhibits good selectivity and stability in multiple-cycle usage and after long-time storage. Finally, the developed sensor was used to determine MP in real samples including soil and vegetables and only simple pretreatment is needed. The detection results were consistent with those obtained from liquid chromatography. Collectively, this newly developed sensor system shows significant potential for use in a variety of fields like food safety, drug residue determination and environmental monitoring.

Preparation and characterization of monodisperse microcapsules with alginate and bentonite via external gelation technique encapsulating Pseudomonas putida Rs-198.

This paper evaluated the external gelation technique for preparing microcapsules. The microcapsules were consisted of Pseudomonas putida Rs-198 (Rs-198) core and sodium alginate (NaAlg)-bentonite (Bent) shell. Different emulsification rotation speeds and core/shell ratios were used to prepare the microcapsules of each formulation. The near-spherical microcapsules were monodisperse with a mean diameter of 25-100 µm and wrinkled surfaces. Fourier transform infrared spectrophotometry (FTIR) and thermogravimetric analysis (TGA) revealed the physical mixture of the wall material and the superior thermal stability of the microcapsules. Percentage yield, water content, and encapsulation efficiency were evaluated and correlated with the changes in emulsification rotation speed and core/shell ratio. In vitro release experiments demonstrated that 60% of the bacteria were released from the NaAlg-Bent microcapsules within three days. Considerably better survival was observed for encapsulated cells compared to free cells, especially in pH 4.0 and 10.0. In summary, the desired properties of microcapsules can be obtained by external gelation technique and the microcapsules on the bacteria had a good protective effect.

A novel polydopamine-based chemiluminescence resonance energy transfer method for microRNA detection coupling duplex-specific nuclease-aided target recycling strategy.

MicroRNAs (miRNAs), functioning as oncogenes or tumor suppressors, play significant regulatory roles in regulating gene expression and become as biomarkers for disease diagnostics and therapeutics. In this work, we have coupled a polydopamine (PDA) nanosphere-assisted chemiluminescence resonance energy transfer (CRET) platform and a duplex-specific nuclease (DSN)-assisted signal amplification strategy to develop a novel method for specific miRNA detection. With the assistance of hemin, luminol, and H2O2, the horseradish peroxidase (HRP)-mimicking G-rich sequence in the sensing probe produces chemiluminescence, which is quickly quenched by the CRET effect between PDA as energy acceptor and excited luminol as energy donor. The target miRNA triggers DSN to partially degrade the sensing probe in the DNA-miRNA heteroduplex to repeatedly release G-quadruplex formed by G-rich sequence from PDA for the production of chemiluminescence. The method allows quantitative detection of target miRNA in the range of 80 pM-50 nM with a detection limit of 49.6 pM. The method also shows excellent specificity to discriminate single-base differences, and can accurately quantify miRNA in biological samples, with good agreement with the result from a commercial miRNA detection kit. The procedure requires no organic dyes or labels, and is a simple and cost-effective method for miRNA detection for early clinical diagnosis.

Supportless electrochemical sensor based on molecularly imprinted polymer modified nanoporous microrod for determination of dopamine at trace level.

In this work, we developed a novel freestanding metallic microrod as working electrode for highly sensitive and selective electrochemical detection of trace dopamine (DA). The electrode was facilely fabricated via first dealloying smooth Au-Ag alloy microrod (AMR) into nanoporous Au-Ag alloy microrod (NPAMR) and further modifying with electro-polymerized molecularly imprinted polymer (MIP). Influencing factors during electro-polymerization process including pH value and molar ratio of monomer to template molecule were optimized. Under the optimal conditions, a linear range from 2 × 10(-13) to 2 × 10(-8)M for measuring DA was obtained with an ultralow detection limit of 7.63 × 10(-14)M (S/N=3). In addition, the MIP-modified electrode (MIP/NPAMR) was successfully employed to test DA in serum and brain samples.

Ultrasensitive and selective assay of glutathione species in arsenic trioxide-treated leukemia HL-60 cell line by molecularly imprinted polymer decorated electrochemical sensors.

Herein a pair of molecularly imprinted polymer (MIP) modified electrochemical sensors were reported to detect glutathione (GSH) and glutathione disulfide (GSSG) in arsenic trioxide-treated HL-60 cells. MIP film was in situ synthesized onto electrode surface via electro-polymerization in a facile way. The characteristics of the obtained sensors were investigated by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). Both GSH-MIP and GSSG-MIP sensors exhibit the relatively wide linear detection range and low detection limit of 1.33 × 10(-10) M (S/N=3). It is found that N-acetylcysteine and DL-homocysteine, the precursors of GSH, show little influence on the detection of glutathione species, nor did the reactants of arsenite and GSH. Such strategies were successfully applied to discriminate GSH and GSSG in cell samples with acceptable recoveries of 92.0-109.1%, and the results are comparable with classic o-phthalaldehyde fluorospectrophotometry. Moreover, the presented sensors allow for easy disclosure of the reversion of malignant phenotype in leukemia cells via glutathione species analysis.

Novel electrochemical sensing platform based on a molecularly imprinted polymer decorated 3D nanoporous nickel skeleton for ultrasensitive and selective determination of metronidazole.

A novel electrochemical sensor has been developed by using a composite element of three-dimensional (3D) nanoporous nickel (NPNi) and molecularly imprinted polymer (MIP). NPNi is introduced in order to enhance the electron-transport ability and surface area of the sensor, while the electrosynthesized MIP layer affords simultaneous identification and quantification of the target molecule by employing Fe(CN)6(3-/4-) as the probe to indicate the current intensity. The morphology of the hybrid film was observed by scanning electron microscopy, and the properties of the sensor were examined by cyclic voltammetry and electrochemical impedance spectroscopy. By using metronidazole (MNZ) as a model analyte, the sensor based on the MIP/NPNi hybrid exhibits great features such as a remarkably low detection limit of 2 × 10(-14) M (S/N = 3), superb selectivity in discriminating MNZ from its structural analogues, and good antiinterference ability toward several coexisting substances. Moreover, the proposed method also demonstrates excellent repeatability and stability, with relative standard deviations of less than 1.12% and 1.4%, respectively. Analysis of MNZ in pharmaceutical dosage form and fish tissue is successfully carried out without assistance of complicated pretreatment. The MIP/NPNi composite presented here with admirable merits makes it a promising candidate for developing electrochemical sensor devices and plays a role in widespread fields.

A PDMS/paper/glass hybrid microfluidic biochip integrated with aptamer-functionalized graphene oxide nano-biosensors for one-step multiplexed pathogen detection.

Infectious pathogens often cause serious public health concerns throughout the world. There is an increasing demand for simple, rapid and sensitive approaches for multiplexed pathogen detection. In this paper we have developed a polydimethylsiloxane (PDMS)/paper/glass hybrid microfluidic system integrated with aptamer-functionalized graphene oxide (GO) nano-biosensors for simple, one-step, multiplexed pathogen detection. The paper substrate used in this hybrid microfluidic system facilitated the integration of aptamer biosensors on the microfluidic biochip, and avoided complicated surface treatment and aptamer probe immobilization in a PDMS or glass-only microfluidic system. Lactobacillus acidophilus was used as a bacterium model to develop the microfluidic platform with a detection limit of 11.0 cfu mL(-1). We have also successfully extended this method to the simultaneous detection of two infectious pathogens - Staphylococcus aureus and Salmonella enterica. This method is simple and fast. The one-step 'turn on' pathogen assay in a ready-to-use microfluidic device only takes ~10 min to complete on the biochip. Furthermore, this microfluidic device has great potential in rapid detection of a wide variety of different other bacterial and viral pathogens.

Overoxidized polypyrrole/multi-walled carbon nanotubes composite modified electrode for in vivo liquid chromatography-electrochemical detection of dopamine.

Overoxidized polypyrrole/multi-walled carbon nanotubes (OPPy/MWNTs) modified electrode has been developed for sensitively detecting dopamine (DA). OPPy films developed outside MWNTs might have a porous morphology. Thus, OPPy/MWNTs films developed by this method do not reject ascorbic acid (AA). However, OPPy/MWNTs modified electrode shows largely enhancing oxidative current responses of DA. When combined with liquid chromatography, it not only obtains a low detection limit of 7.5 x 10(-10) mol L(-1) for DA, but also improves the selectivity of DA detection. Mechanisms for the enhancement are also well discussed in this paper. With this approach, microdialysis has been employed for successful assessment of DA in rat striatum.