Impedance nanobiosensor based on enzyme-conjugated biosynthesized gold nanoparticles for the detection of Gram-positive bacteria.

In this report, gold nanoparticles (GNPS) were synthesized using cell-free extracts of seven different isolates, namely, Pseudomonas aerogenosa CEBP2, Pseudomonas sp. CEBP1, Pseudomonas pseudoalcaligenes CEB1G, Acinetobactor baumani CEBS1, Cuprividus sp. CEB3, Micrococcus luteus CUB12, and Pandoraea sp. CUB2S. The spectroscopic (UV-vis, FTIR, DLS, XRD, EDS) and microscopic (FESEM, TEM) results confirm the reduction of Au3+ to Au0 in the presence of biomolecules having reducing as well as self-stabilizing activity. In this green synthesis approach, the average particle size of biosynthesized GNPS might vary (4-60 nm) depending on the bacterial species, pH of the media, incubation time, and temperature. In this study, GSH-modified BSGNPs (Au-GSH) have shown antimicrobial activity with better stability against Gram-positive bacteria. After conjugation of lysozyme with Au-GSH (lyso@Au-GSH), the zone of inhibition was enhanced from 12 to 23 mm (Au-GSH). The TEM study shows the spherical GNP (16.65 ± 2.84) turns into a flower-shaped GNP (22.22 ± 3.12) after conjugation with lysozyme due to the formation of the protein corona. Furthermore, the nanobioconjugate (lyso@Au-GSH) was immobilized with Nafion on a glassy carbon electrode to fabricate a label-free impedance biosensor that is highly sensitive to monitor changes in the transducer surface due to biomolecular interactions. The uniquely designed biosensor could selectively detect Gram-positive bacteria in the linear range of 3.0 × 101-3 × 1010 cfu mL-1 with RE <5%. The proposed simplest biosensor exhibited good reproducibility (RSD = 3.1%) and excellent correlation (R2 = 0.999) with the standard plate count method, making it suitable for monitoring Gram-positive bacterial contamination in biofluids, food, and environmental samples.

Biosorption of p-chloro meta xylenol (PCMX) by bacterium-encapsulated calcium alginate beads in a novel plug flow process.

P-Chloro-Meta-Xylenol (PCMX) is a widely used disinfectant. In the current pandemic scenario, its consumption has increased largely, and as a result, wastewater is loaded heavily with PCMX as a contaminant. Remediation of this ecologically toxic phenolic compound is therefore a burning issue. This study proposes an eco-friendly biosorption-based remediation technique to remove PCMX. A novel isolated phenol-resistant gram-negative bacterium, Pandoraea sp. strain BT102, is first encapsulated in biopolymeric calcium alginate beads. These beads are packed in a long adsorption tube and the contaminated water was passed through this packed tube resembling a plug flow reactor. This unique plug-flow set-up is capable of reducing PCMX concentration from 100 mg L-1 to 2.85 µg L-1 within 4 h using only 30 g of adsorbent, resulting in 99.99% removal efficiency. Adsorption isotherms and kinetics are studied using batch experimental data. A PCMX loading capacity of the encapsulated calcium alginate beads is found to be 961.7 mg g-1, and the Freundlich isotherm results suggested the phenomenon of cooperative adsorption. A good agreement of the pseudo-second-order kinetic model along with the intra-particle diffusion model suggests a multilayer diffusion-controlled adsorption process. Biosorption of PCMX by the bacterium-modified beads was confirmed by Transmission Electron Microscopy (TEM), Scanning Electron Microscopy (SEM), Energy Dispersive X-ray (EDX), and Fourier-Transform Infrared spectroscopy (FT-IR) analyses. The application of multivariate model-based Response Surface Methodology (RSM) reveals flow rate to be the most important factor controlling the rate of bioremediation.

An interference-free new xanthine biosensor based on immobilized enzyme-nanogold conjugate on carbon nanotube doped poly(3,4-Ethylenedioxythiophene) composite film.

A new design of biosensor based on polymeric nano(bio)composite has been proposed for the selective detection of xanthine to be used in the clinical analysis as well as food quality control. The xanthine oxidoreductase (XOR) gene ofPseudomonas aerogenosastrain CEBP1 wascloned to obtainpurifiedenzyme through affinity chromatography. fMWCNTdoped PEDOTwas electrodeposited on the working electrodeto enhance the sensitivity and selectivity of the biosensor. Bio-synthesized gold nanoparticles conjugated XOR (Au-XOR) was covalently immobilized on the polymeric nanocomposite. The enzymatic activity was enhanced 1.12 times with increased substrate affinity. The surface morphology and structural properties of the polymeric layer were investigated using SEM, FESEM, TEM. Electrochemical characteristics were performed by cyclic voltammetry, differential pulse voltammetry (DPV), and electrochemical impedance spectroscopy. Xanthine was oxidized (pH 7.0) on the uniquely designed polymeric nano(bio)composite modified electrode at a lower anodic potential of + 0.446 V vs. Ag/AgCl (3 M NaCl)at optimized DPV conditions. The simple, newly designed Au-XOR/fMWCNT-PEDOT/GCE exhibited interference-free reproducibility and stability (∼4 months) with excellent sensitivity of 16.075 µA.µM-1.cm-2for the quantification of xanthine in biological samples such as blood, tissue, urine. The applicability of thebiosensor was validatedby comparing the sensing results for the real biological fluidic solutions with HPLC data (RE = 0.5-3.1%).

Electrochemical Column Cell for Continuous Oxidative Inactivation of Pathogens and Reductive Removal of Toxic Heavy Metals.

A solar-driven electrochemical column (EC) was developed for cathodic sequestration remediation of heavy metals (HMs) and anodic electroporative inactivation of pathogenic bacteria (PB) with continuous flow capacity for sustainable production of drinking water from wastewater. The method produces "revitalized drinking water" by keeping its natural mineral nutrients boosted with dissolved oxygen. The EC was constructed with graphene oxide (GO) synthesized via photoassisted electrochemical oxidation of CF (PEGO-CF) as the cathode and phytoreduced GO (RPEGO-CF) as the anode. In the EC, effluent is passed upward through the microchannel of CF electrodes to obtain a higher contact time with water molecules, enabling deposition of HMs and oxidative inactivation of PB, collectively termed electroadsorptive dialysis (EAD). PEGO-CF and RPEGO-CF stacked inside the EC resulted in the increased surface area and thereby the removal efficiency. Reactive oxygen species (ROS) produced at the anode damaged the bacterial cell sheath, while the oxygen functional group and the cathodic negative potential had a concurrent effect in "sequestration" of HMs. Density functional calculations showed that PEGO might transfer an electron from the highest occupied molecular orbital (HOMO) to the lowest unoccupied molecular orbital (LUMO) under applied negative potential leading to internal system crossing to the vacant d-orbitals of HMs, allowing for simultaneous coordination and deposition. The EC produced 313 L of revitalized water from wastewater augmented with 500 µg L-1 HMs and 107 CFU mL-1 pathogenic bacteria (Escherichia coli and Staphylococcus aureus). Only a 3.6 J energy investment produced 1 L of revitalized water, which is ∼2000 times less than the usual energy consumption by electroporation and the lowest value obtained to date for bacterial inactivation with heavy metal removal. Laboratory-to-industrial scale-up calculations were performed for this water-purifying technology involving a water-energy nexus, promising high-efficiency bacterial inactivation, and HM remediation to obtain energy-efficient clean and revitalized water.

Interference-free electrocatalysis of p-chloro meta xylenol (PCMX) on uniquely designed optimized polymeric nanohybrid of P(EDOT-co-OPD) and fMWCNT modified glassy carbon electrode.

p-Chloro-meta-Xylenol (PCMX) is an environmentally hazardous phenolic compound having biocidal and antiseptic activity. Very few research publications addressed monitoring this contaminant. This paper presents a rapid sensing system to quantify it in waste water samples. The electrochemical activity of PCMX was exploited through a unique polymeric nanocomposite modified transducer for its quantification. Poly[(3,4-Ethylenedioxythiophene)-co-(o-phenylenediamine)] [P(EDOT-co-OPD)] was deposited through one-step electropolymerization technique on the glassy carbon electrode (GCE) modified by functionalized multi-wall carbon nanotubes (fMWCNTs). An optimized combination of these constituents was evaluated using response surface methodology (RSM) based Box-Behnken experimental design. This maximized the response for PCMX using differential pulse voltammetry (DPV). The sensing matrix was characterized by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The structural and morphological study of the modified film was conducted by Fourier transform-infrared spectroscopy (FT-IR), Raman spectroscopy, scanning electron microscopy (SEM), and field emission scanning electron microscope (FESEM). The anodic peak current could be read from a wide range of 0.5-225 µM calibration curve with a detection limit of 0.2545 µmol L-1. Interestingly this work did not use any biomaterial in the modification but achieved interference-free response with excellent selectivity, sensitivity (0.4668 µA µM-1 cm-2), reproducibility (RSD = 2.2%), and repeatability. The sensing platform showed good stability (85.7%) of 3 months even after 150 times repetitive use. Its applicability for real samples was established by good correlation with standard methods.

Artificial Bifunctional Photozyme of Glucose Oxidase-Peroxidase for Solar-Powered Glucose-Peroxide Detection in a Biofluid with Resorcinol-Formaldehyde Polymers.

Photozymes or artificial photosynthesis based on alternative natural enzymes is vital for the sustainable development of next-generation healthcare, energy, and materials science. Herein, we report resorcinol-formaldehyde (RF) resins as a solar-driven metal-free bifunctional glucose oxidase-peroxidase stand-alone photozyme for the colorimetric dual detection of hydrogen peroxide and glucose. The π-bond conjugated benzenoid-ortho/para quinoid RF polymers are efficient for glucose oxidation and hydrogen peroxide reduction with concurrent 3,3',5,5'-tetramethylbenzidine oxidation under natural sunlight. The photoinduced colorimetric process could detect H2O2 up to 3.5 µM at 652 nm with the linear range of 0.1-2 mM. A limit of detection of 9.2 µM was exhibited by the system while measuring glucose with a linearity from 0.2 to 8.5 mM. The formation of hydroxyl radicals (•OH) from glucose oxidation reactions was evidenced by spin trapping electron paramagnetic resonance studies conducted herein. The results indicated that RF resins possessed strong intrinsic glucose oxidase and peroxidase (POx)-like activity under natural sunlight with promising storage and operation. This simple photozyme will definitely have potential uses in biomimetic solar-driven catalysis, bioenergy, and biomedicine.

A simple electrochemical approach to fabricate functionalized MWCNT-nanogold decorated PEDOT nanohybrid for simultaneous quantification of uric acid, xanthine and hypoxanthine.

Medical diagnostics and detection of food spoilage require estimation of hypoxanthine (HX), xanthine (XN), and uric acid (UA). A selective sensing platform has been proposed for simultaneous detection of all these species. Functionalized multi-walled carbon nanotube (fMWCNT) stabilized nanogold decorated PEDOT:TOS polymeric nanocomposite (Au-PEDOT-fMWCNT) was synthesized through rapid one-step electropolymerization to enhance conductivity and active surface area by several folds. Electrochemical activities of the proposed sensing platform were analyzed by cyclic voltammetry (CV) and differential pulse voltammetry (DPV), electrochemical impedance spectroscopy (EIS). Analyses through SEM, FESEM and TEM were performed to explore the surface morphology and elemental analysis of the polymeric nanohybrid was investigated by XPS, Raman, FTIR, XRD spectroscopy. Electro-catalysis of UA, XN and HX occurred at low oxidation potentials i.e. 0.082, 0.463 and 0.808 V, respectively in the optimized conditions. The uniquely designed simple, interference free Au-PEDOT-fMWCNT/GCE sensor exhibited high selectivity, good reproducibility, reusability (∼180 times) and stability (∼3 month) with excellent sensitivity of 1.73, 14.31 and 3.82 µA µM-1 cm-2 for UA, XN and HX, respectively. The sensor exhibited linear ranges of detection as 0.1-800, 0.05-175 and 0.1-150 µM with detection limits of 199.3, 24.1 and 90.5 nM for quantification of UA, XN and HX respectively. The performance of the proposed sensor was validated by addition of UA, XN and HX in human serum, urine and fish samples by comparing to those using HPLC. The results indicated good applicability of the proposed sensor for simultaneous detection of UA, XN, HX in real biological fluids.

Chemically modified carbon nitride-chitin-acetic acid hybrid as a metal-free bifunctional nanozyme cascade of glucose oxidase-peroxidase for "click off" colorimetric detection of peroxide and glucose.

Hybrid nanomaterials-based artificial enzymes with numerous utilities are necessary to develop future bionic devices in mimicking physiological processes. This paper demonstrates bifunctional enzyme mimicking roles of a metal-free nanozyme hybrid of chemically modified graphitic carbon nitride (MGCN), chitin and acetic acid (AcOH). The MGCN exhibited glucose oxidase-mimicking activity and chitin-AcOH mirrored peroxidase. MGCN-chitin-AcOH when in contact with glucose, oxidised glucose to gluconic acid and hydrogen peroxide (H2O2) while the chitin-AcOH decomposed the generated H2O2, as proved separately, by concurrent oxidation of 3,3',5,5'-tetramethylbenzidine (TMB). The super-sensitive colorimetric process produced linear regression equation for H2O2 as A = 0.00105C + 0.0630 (C:µM, R2 = 0.9961) with a detection limit of 0.052 µM, whereas for glucose, the linear relationship was A = 0.00084C + 0.0458 (C:µM, R2 = 0.9952) having a detection limit of 0.055 µM. The developed method was also successfully applied for assessment of H2O2 and glucose in human serum and urine samples. Non-enzymatic glucose test strips from MGCN-chitin-AcOH based hydrogel were reported and verified for semi-quantitative analysis of glucose. These compared well with results from standard enzyme-based colorimetric procedure. The developed hybrid nanozyme provided feasible alternatives to the two natural enzymes (peroxidase and glucose oxidase) realized through real sample analysis. The developed hybrid nanozyme can be successfully used for colorimetric detection of peroxide and glucose in medical diagnostics.

UIISScan 1.1: A Field portable high-throughput platform tool for biomedical and agricultural applications.

The colorimetric sensing technology has evolved into an essential tool for high-throughput analysis including portability and cost-effectiveness among available biomedical and agricultural screening approach. In this endeavor, the objective of work is to focus on the development of a field-portable instrument based on an Uniform Illumination Imaging System (UIIS), which will facilitate the colorimetric biochemical sensing. The developed field-portable, wavelength independent UIIS has been exploited for (a) rotavirus detection using commercial enzymatic immunoassay based microplate kit; (b) pesticide residue detection and quantification; The proposed system exhibited a good correlation in comparison to another two conventional techniques, i.e., multi-plate reader (r = 0.9991938) and LC-MS/MS (r = 0.998877399) with a short analysis time of 5 min for 95 test samples. Moreover, the feasibility of UIIS system has also been explored as field-portable enzyme-linked immunosorbent assay (ELISA) plate reader. By incorporating the Mahalanobis distance calculation, the advanced algorithm has been investigated and developed to analyze the data. The overall dataset was transformed into a matrix format to give a good correlation with a conventional plate reader, i.e., r = 0.915389612. Internet of things (IoT) enabled decision support system can be exploited by using big data analytics. Finally, test results can be shared with concerned stakeholders and the remote users. Thus, the developed UIIS will help to identify potential public health threats expeditiosly compared to conventional time consuming process of sample submission to the laboratory for analysis.

Modelling of growth kinetics of isolated Pseudomonas sp. and optimisation of parameters for enhancement of xanthine oxidoreductase production by statistical design of experiments.

This report presents the substrate inhibitory effect of xanthine (XN) on microbial growth and optimisation of effective parameters to achieve high enzyme activity of xanthine oxidoreductase (XOR) through statistical design. Three efficient isolated strains (Pseudomonas aeruginosa CEBP1 and CEBP2, Pseudomonas sp. CEB1G) were screened for growth kinetic studies. Substrate inhibitory models (eg. Aiba, Edward) could explain the growth kinetics of CEBP1, CEBP2 and CEB1G very well with various initial [XN] (S0), e.g., 0.1-35 g L-1. Highest XOR activity was obtained at stationary phase when biomass yield was high. Highest catalytic efficiency (kcat/KM) of XOR was obtained by CEBP1 at optimum specific growth rate of 0.082 h-1 and biomass yield of 0.196 g g-1 at S0 = 5 g L-1. The effects of S0, pH and temperature were studied by Box-Behnken experimental design to evaluate the interactive effects of the significant variables influencing XOR production by CEBP1. ANOVA with high correlation coefficient (R2 > 0.99) and lower 'Prob > F'value (< 0.05) validated the second order polynomial model for the enzyme production. The highest XOR activity of 31.2 KU min-1 mg-1 was achieved by CEBP1 under optimised conditions (35 °C; S0=5 g L-1; pH = 7.0) as compared to any report in literature. A sevenfold substrate affinity of the enzyme was observed after purification.

3­Mercapto­propanoic acid modified cellulose filter paper for quick removal of arsenate from drinking water.

This paper reports a simple, facile and rapid preparation of 3­mercapto­propanoic acid (MPA) modified cellulose filter paper (MPA-Cell paper) for arsenate removal from drinking water. The MPA was covalently grafted to the cellulose filter paper (Cell) by esterification process through the formation of O­acylisourea intermediate and characterized by the FTIR, SEM, EDS and XPS analyses. The arsenate adsorption efficiency was studied for batch and semi-continuous systems while exploring the adsorption kinetics, isotherm and the effect of pH for the former. The experimental data fitted well with Langmuir, Dubinin-Radushkevich (DR) and pseudo second order kinetic models. The mechanism of adsorption was studied by FTIR spectroscopy utilizing the adsorption isotherm, kinetic model and XPS results. The modified filter paper performed well at nearly neutral pH in arsenate removal through adsorption and demonstrated a significant arsenate uptake capacity of 92.59 mg/g. The DR and FTIR results indicated that the adsorption of arsenate ion occurred through ion exchange process. The MPA-Cell paper could have a potential use as low-cost but efficient commercial adsorbent for arsenate abatement from contaminated drinking water by both batch as well as semi-continuous operating systems. The MPA-Cell paper could purify ground water containing high level of arsenate.

Silver nanoparticles decorated eggshell membrane as an effective platform for interference free sensing of dopamine.

In this paper a simple electrochemical sensing of dopamine by a new effective immobilization of tyrosinase (Tyr) enzyme on eggshell membrane (ESM) along with silver nanoparticles (AgNPs) is reported. The modified membrane was characterized by scanning electron microscope (SEM), energy dispersive spectroscopy (EDAX), X-Ray diffraction (XRD). A simple solution based approach was used to prepare AgNPs on biomembrane followed by glutaraldehyde activation to immobilize Tyr on the nanoparticles decorated ESM. The direct electrochemistry of DA oxidation was performed through cyclic voltammetry (CV) and differential pulse voltammetry (DPV). Characterization of membrane was accomplished by electrochemical impedance spectroscopy (EIS). Prepared electrode showed very good stability, reproducibility, high selectivity, easy preparation and regeneration of electrode. The proposed sensor exhibited low detection limits 1.7ngL-1 with wide linear range 10-1000 ngL-1, excellent sensitivity (14.28µA µgL-1cm-2) with good storage and operational stabilities. The accurate measurement of dopamine in blood serum and good recoveries in spiked serum samples ensured great potential for medical diagnostics.

Remediation of arsenic in mung bean (Vigna radiata) with growth enhancement by unique arsenic-resistant bacterium Acinetobacter lwoffii.

Arsenic, a carcinogenic and toxic contaminant of soil and water, affects human health adversely. During last few decades, it has been an important global environmental issue. Among several arsenic detoxification methods remediation using arsenic resistant microbes is proved to be environment-friendly and cost-effective. This study aimed to test the effects of arsenic utilizing bacterial strain Acinetobacter lwoffii (RJB-2) on arsenic uptake and growth of mung bean plants (Vigna radiata). RJB-2 exhibited tolerance up to 125mM of arsenic (V) and 50mM of arsenic (III). RJB-2 produced plant growth promoting substances e.g. indole acetic acid (IAA), siderophores, exopolysaccharide (EPS) and phosphate solubilization in the absence and in presence of arsenic. Pot experiments were used to scrutinize the role of RJB-2 on arsenic uptake and growth of mung bean plants grown in soil amended with 22.5mgkg-1 of sodium arsenate (Na2HAsO4·7H2O). RJB-2 could arrest arsenic uptake in just 7days and increase plant growth, number of plants per pot, chlorophyll and carotenoid content of the mung bean plants. RJB-2 formed biofilm and its root-association helped to abate arsenic uptake in mung bean. Confocal and light microscopic studies also revealed the abatement of arsenic uptake and increase in chlorophyll content in mung bean plants in presence of RJB-2. RJB-2 was also responsible for less production of reactive oxygen species (ROS) in mung bean plants reducing the oxidative damage caused by arsenic. The lower percentage of electrolytic leakage (EL) in RJB-2 inoculated mung bean plants proved arsenic abatement. The study also reported the distribution of arsenic in various parts of mung bean plant. RJB-2 owing to its intrinsic abilities of plant growth promotion even in presence of high concentrations of arsenic could inhibit arsenic uptake completely and therefore it could be used in large-scale cultivation for phytostabilization of plants.

Statistical optimization of arsenic biosorption by microbial enzyme via Ca-alginate beads.

Bioremediation of arsenic using green technology via microbial enzymes has attracted scientists due to its simplicity and cost effectiveness. Statistical optimization of arsenate bioremediation was conducted by the enzyme arsenate reductase extracted from arsenic tolerant bacterium Pseudomonas alcaligenes. Response surface methodology based on Box-Behnken design matrix was performed to determine the optimal operational conditions of a multivariable system and their interactive effects on the bioremediation process. The highest biosorptive activity of 96.2 µg gm-1 of beads was achieved under optimized conditions (pH = 7.0; As (V) concentration = 1000 ppb; time = 2 h). SEM analysis showed the morphological changes on the surface of enzyme immobilized gluteraldehyde crosslinked Ca-alginate beads. The immobilized enzyme retained its activity for 8 cycles. ANOVA with a high correlation coefficient (R2 > 0.99) and lower "Prob > F"value (<0.0001) corroborated the second-order polynomial model for the biosorption process. This study on the adsorptive removal of As (V) by enzyme-loaded biosorbent revealed a possible way of its application in large scale treatment of As (V)-contaminated water bodies.

Differential pulse anodic stripping voltammetry for detection of As (III) by Chitosan-Fe(OH)3 modified glassy carbon electrode: A new approach towards speciation of arsenic.

An efficient electrochemical sensor for As(III) was developed based on adsorption of arsenic on a specially modified electrodes at some applied potential and subsequent i) stripping at a fixed potential by anodic stripping voltammetry ii) analysis by generating surface plasmon resonance (SPR). The working glassy carbon electrode was modified by Chitosan-Fe(OH)3 composite and a reducing agent L-cysteine. The composite enhanced adsorption of As(III) and subsequent reduction to As(O) moieties and measurement by anodic stripping. The surface property of modified electrode was characterized by SEM, AFM, FTIR, XPS and electrochemistry was analyzed by impedance spectroscopy (EIS). Surface Plasmon resonance (SPR) was also employed to investigate the As(III) binding capability of polymer matrix. Several optimum voltammetric parameters e.g supporting electrolyte; 0.1M acetate buffer (pH 5.2) deposition potential, -0.9V; deposition time, 100s were established for anodic stripping voltammetry (ASV). A linear correlation was obtained in the range of 2-100ppb for ASV (R(2) 0.974) with limit of detection 0.072ppb. A variety of common coexistent ions such as Mn, Zn, Pb, Cu, Cd in water samples showed no interferences on the As (III) determination. The method was applied successfully to real samples collected from arsenic affected areas of West Bengal, India.

A novel third generation uric acid biosensor using uricase electro-activated with ferrocene on a Nafion coated glassy carbon electrode.

A new uric acid biosensor was constructed using ferrocene (Fc) induced electro-activated uricase (UOx) deposited within Nafion (Naf) on glassy carbon electrode (GCE). Electro-activation of UOx was successfully achieved by cyclic voltammetry through the electrostatic interaction of Fc with Trp residues within the hydrophobic pockets in UOx. The Naf/UOx/Fc composite was characterised by AFM, FTIR and EDX to ensure proper immobilisation. The interaction of Fc with the enzyme was analysed by Trp fluorescence spectroscopy and the α-helicity of the protein was measured by CD spectropolarimetry. The charge transfer resistance (Rct), calculated from electrochemical impedance spectroscopy, for the modified sensor was lowered (1.39 kΩ) and the enzyme efficiency was enhanced by more than two fold as a result of Fc incorporation. Cyclic voltammetry, differential pulse voltammetry and amperometry all demonstrated the excellent response of the Naf/UOx/Fc/GCE biosensor to uric acid. The sensor system generated a linear response over a range of 500 nM to 600 µM UA, with a sensitivity and limit of detection of 1.78 µA µM(-1) and 230 nM, respectively. The heterogeneous rate constant (ks) for UA oxidation was much higher for Naf/UOx/Fc/GCE (1.0 × 10(-4) cm s(-1)) than for Naf/UOx/GCE (8.2 × 10(-5) cm s(-1)). Real samples, i.e. human blood, were tested for serum UA and the sensor yielded accurate results at a 95% confidence limit.

Isolation of a novel uric-acid-degrading microbe Comamonas sp. BT UA and rapid biosensing of uric acid from extracted uricase enzyme.

Uric-acid-utilizing soil bacteria were isolated, and 16s rRNA sequence was studied for strain identification. The most prominent uricase-producing bacterium was identified as Comamonas sp BT UA. Crude enzyme was extracted, freeze-dried and its Km and Vmax were determined as 40 meu M and 0.047 meu M min-1ml-1 using Line-weaver Burke plot. An activity of 80 U/mg of total protein was observed when cultured at 37 degree C for 84 h at pH 7. The purified enzyme was used to measure uric acid by spectrophotometric method and electrochemical biosensor. In the biosensing system the enzyme was immobilized on the platinum electrode with a biodegradable glutaraldehyde-crosslinked gelatin film having a swelling percentage of 109+/- 3.08, and response was observed by amperometry applying fixed potential. The electrochemical process as obtained by the anodic peak current and scan rate relationship was further configured by electrochemical impedance spectroscopy (EIS). The polymer matrix on the working electrode gave capacitive response for the electrode-electrolyte interaction. The sensitivity of the biosensor was measured as 6.93 meu A meu M -1 with a sensor affinity [Km(app)] of 50 mu M and 95 percent reproducibility after 50 measurements. The spectrophotometric method could be used in the range of 6-1000 mu M, whereas the biosensor generated linear response in the 1.5- 1000 mu M range with a response time of 24 s and limit of detection of 0.56 meu M. Uric acid was estimated in human blood samples by the biosensor and satisfactory results were obtained.

Biosensing and bioremediation of Cr(VI) by cell free extract of Enterobacter aerogenes T2.

Hexavalent chromium or Cr(VI) enters the environment through several anthropogenic activities and it is highly toxic and carcinogenic. Hence it is required to be detected and remediated from the environment. In this study, low-cost and environment-friendly methods of biosensing and bioremediation of Cr(VI) have been proposed. Crude cell free extract (CFE) of previously isolated Enterobacter aerogenes T2 (GU265554; NII 1111) was prepared and exploited to develop a stable biosensor for direct estimation of Cr(VI) in waste water, by using three electrodes via cyclic voltammetry. For bioremediation studies, a homogeneous solution of commercially available sodium alginate and CFE was added dropwise in a continuously stirred calcium chloride solution. Biologically modified calcium alginate beads were produced and these were further utilized for bioremediation studies. The proposed sensor showed linear response in the range of 10-40 µg L(-1) Cr(VI) and the limit of detection was found to be 6.6 µg L(-1) Cr(VI). No interference was observed in presence of metal ions, e.g., lead, cadmium, arsenic, tin etc., except for insignificant interference with molybdenum and manganese. In bioremediation studies, modified calcium alginate beads showed encouraging removal rate 900 mg Cr(VI)/m(3) water per day with a removal efficiency of 90%, much above than reported in literature. The proposed sensing system could be a viable alternative to costly measurement procedures. Calcium alginate beads, modified with CFE of E. aerogenes, could be used in bioremediation of Cr(VI) since it could work in real conditions with extraordinarily high capacity.

Isolation and characterization of phenol utilizing bacteria from industrial effluent-contaminated soil and kinetic evaluation of their biodegradation potential.

Microbial degradation of phenol by pure bacterial species is a well-known approach towards alleviation of environmental pollution. In this study, five phenol-degrading bacterial species designated as CUPS-1 to CUPS-5 were isolated from the oil-effluent dumped sites of Haldia Industrial area of West Bengal, India. Detailed morphological, biochemical and molecular characterization identified CUPS-3 as a novel strain- Stenotrophomonas maltophilia (GU358076), while the others could be identified as Pseudomonas (CUPS-2, 5), Delftia (CUPS-1) and Micrococcus (CUPS-4) genera, respectively. Although all of these strains utilized phenol as their sole carbon source supporting growth, three among them, CUPS-2, CUPS-3 and CUPS-5 proved potential phenol degraders and hence used for further biodegradation studies. Degradation experiments were carried out for several initial phenol concentrations of 500 mg/L, 750 mg/L, 1000 mg/L, 1250 mg/L and 1500 mg/L. The novel strain, CUPS-3 could completely degrade 500 mg/L phenol within 48 h, with 0.0937/h substrate degradation rate and 16.34 mg/L/h substrate consumption rate. The strains degraded phenol via meta-cleavage pathway. Prediction of kinetic parameters of the biodegradation was accomplished Haldane model using the experimental data of degradation rate and phenol concentration as function of time.

Low-cost field test kits for arsenic detection in water.

Arsenic, a common contaminant of groundwater, affects human health adversely. According to the World Health Organization (WHO), the maximum recommended contamination level of arsenic in drinking water is 10 µg/L. The purpose of this research was to develop user-friendly kits for detection of arsenic to measure at least up to 10 µg/L in drinking water, so that a preventive measure could be taken. Two different kits for detection of total arsenic in water are reported here. First, the arsenic in drinking water was converted to arsine gas by a strong reducing agent. The arsine produced was then detected by paper strips via generation of color due to reaction with either mercuric bromide (KIT-1) or silver nitrate (KIT-2). These were previously immobilized on the detector strip. The first one gave a yellow color and the second one grey. Both of these kits could detect arsenic contamination within a range of 10 µg/L-250 µg/L. The detection time for both the kits was only 7 min. The kits exhibited excellent performance compared to other kits available in the market with respect to detection time, ease of operation, cost and could be easily handled by a layman. The field trials with these kits gave very satisfactory results. A study on interference revealed that these kits could be used in the presence of 24 common ions present in the arsenic contaminated water. Though the kits were meant for qualitative assay, the results with unknown concentrations of real samples, when compared with atomic absorption spectrophotometer (AAS) were in good agreement as revealed by the t-test.