Enhancement in ß-galactosidase activity of Streptococcus lactis cells by entrapping in microcapsules comprising of correlated silica nanoparticles.

With the advent of nanotechnology there has been an impetus towards sol gel encapsulation of biocatalyst in mesoporous silica. In this study cells of Streptococcus lactis expressing ß-galactosidase activity were entrapped in silica microcapsules using evaporation induced assembly through spray drying technique. These microcapsules were characterized using complementary techniques of electron microscopy and neutron/X-ray scattering. Under optimized conditions of spray drying process, the cells immobilized in silica microcapsules had higher biocatalytic activity than the free cells. Immobilized cells showed threefold increase in enzyme activity than free cells. Free cells and immobilized Streptococcus lactis cells exhibited pH optimum at 5.0 and 7.0, respectively, whereas, the temperature optimum for both the systems was observed at 45 °C. Michaelis constant for free cells and immobilized cells was 8.33 mM and 4.16 mM, respectively, while Vmax for free cells and immobilized cells was 71.43 µmoles min-1 and 125 µmoles min-1, respectively. The decrease in apparent Km is correlated with the fold increase in apparent Vmax and can be interpreted as favorable for enzyme substrate complex formation when cells were entrapped in microcapsules. Herein, the concept of immobilization using spray dryer at optimal inlet temperature has emerged as an efficient mode for enhancing the catalytic activity of the microencapsulated microbial cells and a plausible mechanism is suggested.

Preparation and application of silica nanoparticles-Ocimum basilicum seeds bio-hybrid for the efficient immobilization of invertase enzyme.

For efficient utilization of immobilized invertase enzyme, a novel bio-hybrid support comprising silica nanoparticles and Ocimum basilicum seed was synthesized. Ocimum basilicum seeds provide a natural fibrouspellicular structure which acts as template for assembly of silica nanoparticles. Bio-hybrids of two different morphologies have been obtained by changing the physico-chemical conditions of the assembly process. Developed bio-hybrids were characterized through small angle X-ray scattering (SAXS), scanning electron microscopy (SEM), Synchrotron radiation based X-ray micro-computed tomography (SRµCT), Brunauer-Emmett-Teller (BET) and Fourier transform infrared spectroscopy (FTIR). Incorporation of the nanoparticles results to a fourfold increase in the available surface area of the seeds which is one of the important criteria for an immobilizing support. Synthesized bio-hybrids were used for the immobilization of commercially applicable invertase enzyme and efficient loading of enzyme was realized. Enzyme immobilized bio-hybrids could be easily separated out and reused up to eight times with 82 % retention of enzyme activity. Present work suggests that the unique features of the bio-hybrid make it suitable candidate for immobilization of enzymes in general.

Gamma radiation-induced in vitro hormetic apogamy in the fern Pityrogramma calomelanos (L.) link.

Pityrogramma calomelanos (L.) Link, popularly known as "Silver fern" has significant importance as a medicinal plant used traditionally for its astringent, analgesic, anti-haemorrhagic, anti-hypertensive, anti-pyretic and anthelminthic properties. This fern demonstrates an increased morphogenetic potential towards sporophyte formation, upon exposure to low doses of gamma radiation. Young sporophytic leaf crosier cultures were established in vitro on agar based Knop's media with and without 20 g/l sucrose. The cultures were subjected to 60Co radiations in the range of 2.5-100 Gy. Apospory (production of gametophytes on sporophytic tissue without spores) was observed on leaf tissue cultured on Knops media with and without sucrose in P. calomelanos, at the end of 60 days. 5 Gy treated explants showed high number of aposporous gametophytes and was comparable to the control. Other tested doses reduced the aposporous gametophyte production significantly. In the second phase of the experimentation, the cultures were retained on the gametophyte induction media for a period of 4 weeks. Aposporous gametophytes were observed to proliferate with occasional development of antheridia. At the end of 4 weeks, morphogenetic development on the gametophytic tissue resulted in a significantly higher number of apogamous sporophytes (production of sporophytes without fusion of gametes) were obtained on 5 Gy treated tissue as compared to control and all the other treated explants. Apogamous sporophytes thus produced were successfully grown in the greenhouse and transferred to the field. Thus the use of gamma radiation in vitro not only reduced the need for sucrose for induction of apospory in P.calomelanos, it also exhibited hormesis at 5 Gy for improved sporophyte production.

An optical microplate biosensor for the detection of methyl parathion pesticide using a biohybrid of Sphingomonas sp. cells-silica nanoparticles.

The previously developed Sphingomonas sp. based optical microplate biosensor for methyl parathion (MP) was good as it detected multiple samples but had poor stability and low sensitivity. The present study aims to overcome these limitations. Silica nanoparticles (Si NP) were thus functionalized with polyethyleneimine (PEI) and the functionalized silica nanoparticles (fSi NP) were then integrated with Sphingomonas sp. cells. The process was optimized for hydrolysis of MP into p-nitrophenol (PNP). Integration of fSi NP with cells was confirmed by FT-IR analysis. Biohybrid of Sphingomonas sp.-fSi NP was immobilized on the wells of microplate and associated directly with the optical transducer of microplate reader. Immobilized biohybrid of Sphingomonas sp.-fSi NP was characterized using SEM. A detection range of 0.1-1ppm MP was achieved from the linear range of calibration plot. After integration with fSi NP the storage stability of biohybrid was enhanced ten times from 18 to 180 days. This study proves that after interaction of cells with fSi NP, improved the sensitivity and stability of the biosensor. Spiked samples were also analyzed and correlated using this biohybrid based biosensor.

Synthesis of one-dimensional gold nanostructures and the electrochemical application of the nanohybrid containing functionalized graphene oxide for cholesterol biosensing.

This manuscript reports a new approach for the synthesis of one dimensional gold nanostructure (AuNs) and its application in the development of cholesterol biosensor. Au nanostructures have been synthesized by exploiting ß-diphenylalanine (ß-FF) as an sacrificial template, whereas the Au nanoparticles (AuNPs) were synthesized by ultrasound irradiation. X-ray diffractometer (XRD), scanning electron microscope (SEM) and energy dispersive analysis of X-rays (EDAX) have been employed to characterize the morphology and composition of the prepared samples. With the aim to develop a highly sensitive cholesterol biosensor, cholesterol oxidase (ChOx) was immobilized on AuNs which were appended on the graphite (Gr) electrode via chemisorption onto thiol-functionalized graphene oxide (GO-SH). This Gr/GO-SH/AuNs/ChOx biosensor has been characterized using cyclic voltammetry (CV), electrochemical impedance spectroscopy and chronoamperometry. CV results indicated a direct electron transfer between the enzyme and the electrode surface. A new potentiostat intermitant titration technique (PITT) has been studied to determine the diffusion coefficient and maxima potential value. The proposed biosensor showed rapid response, high sensitivity, wide linear range and low detection limit. Furthermore, our AuNs modified electrode showed excellent selectivity, repeatability, reproducibility and long term stability. The proposed electrode has also been used successfully to determine cholesterol in serum samples.

Biosorption of uranium by human black hair.

Naturally available low cost materials have gained importance as effective alternative to conventional sorbents for the removal of metal ions from water. The present study describes the use of black hair waste as a sorbent for the removal of uranium ions from an aqueous medium. Alkali treatment of the biomass resulted in a significant increase in its uptake capacity. The optimum pH and contact time for uranium removal were 4.5 and 2 h respectively. It was observed that the experimental data fits well in Ho's pseudo-second order kinetic model. Binding of uranium to the biomass was confirmed using FT-IR spectroscopy. Thus, the present study could demonstrate the utility of human black hair to remove uranium from aqueous medium.

Microplate based optical biosensor for L-Dopa using tyrosinase from Amorphophallus campanulatus.

Developing a biosensor which is capable of simultaneously monitoring l-Dopa levels in multiple samples besides requiring small reaction volume is of great value. The present study describes the detection of l-Dopa using tyrosinase enzyme extracted from Amorphophallus campanulatus and immobilized on the surface of the microplate wells. Among the different approaches used for immobilizing tyrosinase onto the microplate wells, glutaraldehyde treatment was found to be most effective. Besides enzyme activity, ESEM-EDS (environmental scanning electron microscope-energy dispersive system) and Atomic Force Microscopy (AFM) were also carried out to confirm the immobilization of tyrosinase enzyme onto the microplate well surface. This immobilized biocomponent was then integrated with an optical transducer for l-Dopa detection and it showed good reproducibility. The sensing property of the system was studied by measuring the initial rate of dopachrome formation at 475 nm. The calibration plot gave a linear range of detection from 10-1000 µM and the detection limit was calculated to be 3 µM. The immobilized biocomponent was stable for 41 days and was reused up to nine times. Spiked samples (blood plasma) were also analyzed using this biocomponent. This microplate based biosensor thus provides a convenient system for detection of multiple samples in a single run.

Development of a simple bioelectrode for the electrochemical detection of hydrogen peroxide using Pichia pastoris catalase immobilized on gold nanoparticle nanotubes and polythiophene hybrid.

In this paper, a simple and innovative electrochemical hydrogen peroxide biosensor has been proposed using catalase (CATpp) derived from Pichia pastoris as bioelectrocatalyst. The model biocomponent was immobilized on gold nanoparticle nanotubes (AuNPNTs) and polythiophene composite using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide and N-hydroxysuccinimide (EDC-NHS) coupling reagent. In this present work, we have successfully synthesized gold nanoparticles (AuNPs) by ultrasonic irradiation. The tubular gold nanostructures containing coalesced AuNPs were obtained by sacrificial template synthesis. The assembly of AuNPNTs onto the graphite (Gr) electrode was achieved via S-Au chemisorption. The latter was pre-coated with electropolymerized thiophene (PTh) to enable S groups to bind AuNPNTs. The combination of AuNPNTs-PTh, i.e., an inorganic-organic hybrid, provides a stable enzyme immobilization platform. The physical morphology of the fabricated biosensor Gr/PTh/AuNPNTs/EDC-NHS/CATpp was investigated using scanning electron microscopy and energy-dispersive microscopy. The analytical performance of the bioelectrode was examined using cyclic voltammetry, differential pulse voltammetry and chronoamperometry. Operational parameters such as working potential, pH, and thermal stability of the modified electrode were examined. The beneficial analytical characteristics of the proposed electrode were demonstrated. Our results indicate that the Gr/PTh/AuNPNTs/EDC-NHS/CATpp bioelectrode exhibits a wide linear range from 0.05 mM to 18.5 mM of H2O2, fast response time of 7 s, excellent sensitivity of 26.2 mA mM(-1) cm(-2), good detection limit of 0.12 µM and good Michaelis-Menten constant of 1.4 mM. In addition, the bioelectrode displayed good repeatability, high stability and acceptable reproducibility, which can be attributed to the AuNPNTs-PTh composite that provides a biocompatible micro-environment.

Evaporation induced self assembled microstructures of silica nanoparticles and Streptococcus lactis cells as sorbent for uranium (VI).

An assembled microstructure of silica nanoparticles and Streptococcus lactis (S. lactis) cells has been synthesized by evaporation induced self assembly, with the objective of its application in bioremediation. Different morphologies have been realized by tuning the physico-chemical conditions of the assembly process. The potential of these microstructures in removal of uranium (VI) has been evaluated. Morphology dependent uptake has been demonstrated and maximum uptake was seen for the spray dried doughnut shaped microstructure (SDSM). For a fixed morphology, the variation in uptake varies with solution pH, contact time, temperature and initial uranium (VI) concentration. The U (VI) removal was significantly rapid, with more than 85 ± 2% of total uptake in 10 min. The maximum sorption capacity (qmax) of U (VI) at pH 5.0 and temperature 298 K was 169.5 mg/g using SDSM as sorbent. The kinetic data of adsorption of U (VI) are best described by a pseudo-second-order kinetic model. Calculated thermodynamic parameters reveal an endothermic and a spontaneous adsorption process. The present work opens up the possibility of a means for the functionalization of silica microstructures through the incorporation of micro-organism and the potential for the use of these functionalized materials for bioremediation.

Biosorption of uranium by melanin: kinetic, equilibrium and thermodynamic studies.

Limitation of conventional techniques for the removal of heavy metals present at low concentrations, has led to the need for developing alternate technologies like biosorption. In the present study we describe the use of melanin pigment synthesized through green technology, for sorption of uranium from aqueous system. Biosynthesized melanin showed good uptake over a broad pH range. Removal of uranium was rapid and equilibrium was reached within 2h of contact. It was observed that the kinetic data fits well into Lagergren's pseudo-second order equation. A maximum loading capacity of 588.24 mg g(-1) was calculated from Langmuir plot. Thermodynamic studies performed revealed that sorption process was favorable. Binding of uranium on the surface of melanin was confirmed by FT-IR and energy dispersive spectroscopy (EDS). Thus, biosynthesized melanin can be efficiently used as a sorbent for removal of uranium from aqueous solution.

Uranium (VI) recovery from aqueous medium using novel floating macroporous alginate-agarose-magnetite cryobeads.

This study presents a novel development of a floating polymeric-magnetite cryobead for the recovery of hexavalent uranium from the aqueous sub-surfaces. The alginate-agarose-magnetite cryobeads were synthesized by the process of cryotropic-gelation at subzero-temperature. The physico-chemical properties of cryobeads showed high surface area and high interconnected porosity (≈ 90%). Low density of these cryobeads explains their floating property in the aqueous medium. The rheological analysis of cryobeads showed its stability and increased stiffness after uranium adsorption. The presence of magnetite nanoparticles in the porous cryobeads facilitates the recovery of these beads by applying an external magnetic field. Maximum uranium adsorption (97 ± 2%) was observed in the pH range of 4.5-5.5. The thermodynamic parameters suggest passive endothermic adsorption behaviour. HCl was found to be an efficient eluent for the uranium desorption. Five repeated cycles for the desorption of uranium from biosorbent showed 69 ± 3% of uranium recovery. These results suggest stability of these novel floating magnetite-cryobeads under environmental conditions with potential for the recovery of uranium from contaminated aqueous subsurfaces.

An amperometric bienzymatic cholesterol biosensor based on functionalized graphene modified electrode and its electrocatalytic activity towards total cholesterol determination.

Cholesterol oxidase (ChOx) and cholesterol esterase (ChEt) have been covalently immobilized onto functionalized graphene (FG) modified graphite electrode. Enzymes modified electrodes were characterized using cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). FG accelerates the electron transfer from electrode surface to the immobilized ChOx, achieving the direct electrochemistry of ChOx. A well defined redox peak was observed, corresponding to the direct electron transfer of the FAD/FADH(2) of ChOx. The electron transfer coefficient (α) and electron transfer rate constant (K(s)) were calculated and their values are found to be 0.31 and 0.78 s(-1), respectively. For the free cholesterol determination, ChOx-FG/Gr electrode exhibits a sensitive response from 50 to 350 µM (R=-0.9972) with a detection limit of 5 µM. For total cholesterol determination, co-immobilization of ChEt and ChOx on modified electrode, i.e. (ChEt/ChOx)-FG/Gr electrode showed linear range from 50 to 300 µM (R=-0.9982) with a detection limit of 15 µM. Some common interferents like glucose, ascorbic acid and uric acid did not cause any interference, due to the use of a low operating potential. The FG/Gr electrode exhibits good electrocatalytic activity towards hydrogen peroxide (H(2)O(2)). A wide linear response to H(2)O(2) ranging from 0.5 to 7 mM (R=-0.9967) with a sensitivity of 443.25 µA mM(-1) cm(-2) has been obtained.

Functionalized-graphene modified graphite electrode for the selective determination of dopamine in presence of uric acid and ascorbic acid.

Graphene is chemically synthesized by solvothermal reduction of colloidal dispersions of graphite oxide. Graphite electrode is modified with functionalized-graphene for electrochemical applications. Electrochemical characterization of functionalized-graphene modified graphite electrode (FGGE) is carried out by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The behavior of FGGE towards ascorbic acid (AA), dopamine (DA) and uric acid (UA) has been investigated by CV, differential pulse voltammetry (DPV) and chronoamperommetry (CA). The FGGE showed excellent catalytic activity towards electrochemical oxidation of AA, DA and UA compared to that of the bare graphite electrode. The electrochemical oxidation signals of AA, DA and UA are well separated into three distinct peaks with peak potential separation of 193mv, 172mv and 264mV between AA-DA, DA-UA and AA-UA respectively in CV studies and the corresponding peak potential separations in DPV mode are 204mv, 141mv and 345mv. The FGGE is successfully used for the simultaneous detection of AA, DA and UA in their ternary mixture and DA in serum and pharmaceutical samples. The excellent electrocatalytic behavior of FGGE may lead to new applications in electrochemical analysis.