Jagriti Narang

Immobilization of rat brain acetylcholinesterase on ZnS and poly(indole-5-carboxylic acid) modified Au electrode for detection of organophosphorus insecticides.

A novel, highly sensitive amperometric biosensor for detection of organophosphorus (OP) compounds has been constructed, based on rat brain acetylcholinesterase (AChE) immobilized onto nanocomposite of ZnS-nanoparticles (ZnSNPs) and poly(indole-5-carboxylic acid) electrodeposited on Au electrode. In the presence of acetylthiocholine chloride (ATCl) as a substrate, ZnSNPs promoted electron transfer reactions at a lower potential and catalyzed electrochemical oxidation of enzymatically formed thiocholine, thus increasing detection sensitivity. Under optimum conditions (phosphate buffer, pH 7.5 and 30°C), the inhibition of AChE by malathion and chlorpyrifos was proportional to their concentrations in the range, 0.1-50nM and 1.5-40nM, respectively. The biosensor determined malathion and chlorpyrifos in spiked tap water samples with a acceptable accuracy (95-100%). The enzyme electrode had long-storage stability (50% retention of initial activity within 2 months, when stored at 4°C).

A non-enzymatic sensor for hydrogen peroxide based on polyaniline, multiwalled carbon nanotubes and gold nanoparticles modified Au electrode

We describe the construction of a polyaniline (PANI), multiwalled carbon nanotubes (MWCNTs) and gold nanoparticles (AuNPs) modified Au electrode for determination of hydrogen peroxide without using peroxidase (HRP). The AuNPs/MWCNT/PANI composite film deposited on Au electrode was characterized by Scanning Electron Microscopy (SEM) and electrochemical methods. Cyclic voltammetric (CV) studies of the electrode at different stages of construction demonstrated that the modified electrode had enhanced electrochemical oxidation of H(2)O(2), which offers a number of attractive features to develop amperometric sensors based on split of H(2)O(2). The amperometric response to H(2)O(2) showed a linear relationship in the range from 3.0 μM to 600.0 μM with a detection limit of 0.3 μM (S/N = 3) and with high sensitivity of 3.3 mA μM(-1). The sensor gave accurate and satisfactory results, when employed for determination of H(2)O(2) in milk and urine.

Silver nanoparticles/multiwalled carbon nanotube/polyaniline film for amperometric glutathione biosensor

A new silver nanoparticles (AgNPs)/carboxylated multiwalled carbon nanotubes (c-MWCNT)/polyaniline (PANI) film has been synthesized on Au electrode using electrochemical techniques. The enzyme glutathione oxidase (GSHOx) (EC 1.8.3.3) was immobilized covalently on the surface of AgNPs/c-MWCNT/PANI/Au electrode to construct the glutathione biosensor. The modified electrode was characterized by scanning electron microscopy (SEM), cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS) and Fourier transform infrared (FTIR) spectrophotometry. The biosensor showed optimum response within 4s at +0.4V vs. Ag/AgCl, pH 6.0 and 35 °C, with a linear working range of 0.3-3500 μM and a detection limit of 0.3 μM. The glutathione biosensor was employed for measurement of glutathione content in hemolysated erythrocyte (RBC). The sensor was evaluated with 97.77% and 99.16% recovery of added glutathione in hemolysated RBC and 2.4% and 6.3% within and between batch coefficients of variation (CVs) respectively. The enzyme electrode lost 50% of its initial activity after 300 uses over a period of 3 months, when stored at 4 °C. The biosensor has the advantages over earlier biosensors in terms of greater stability, lower response time and no interference by a number of RBC hemolysate substances.

Construction of a triglyceride amperometric biosensor based on chitosan-ZnO nanocomposite film.

A method is described for construction of a novel amperometric triglyceride (TG) biosensor based on covalent co-immobilization of lipase, glycerol kinase (GK) and glycerol-3-phosphate oxidase (GPO) onto chitosan (CHIT) and zinc oxide nanoparticles (ZnONPs) composite film deposited on the surface of Pt electrode. The enzymes-ZnONPs-CHIT composite was characterized by X-ray diffraction (XRD), scanning electron microscope (SEM), cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The sensor showed optimum response within 6s at pH 7.5 and temperature of 35°C. The sensor measures current due to electrons generated at 0.4V against Ag/AgCl from H(2)O(2), which is produced from triolein by co-immobilized enzymes. A linear relationship was obtained between a wide triolein concentration range (50-650 mg/dl) and current (mA) under optimum conditions. The biosensor showed high sensitivity, low detection limit (20 mg/dl) and good storage stability (half-life of 7 months at 4°C). The biosensor was unaffected modified by a number of serum substances at their physiological concentrations. The biosensor was evaluated and employed for determination of TG in sera in apparently healthy subjects and persons suffering from hypertriglyceridemia.

Immobilization of rat brain acetylcholinesterase on porous gold-nanoparticle–CaCO3 hybrid material modified Au electrode for detection of organophosphorous insecticides

An acetylcholinesterase (AChE) purified from rat brain was immobilized onto gold nanoparticles (AuNPs) assembled on the surface of porous calcium carbonate (CaCO(3)) microsphere. The resulting AChE-AuNPs-CaCO(3) bioconjugate was mounted on the surface of Au electrode with the help of silica sol-gel matrix to prepare the working electrode. This electrode was connected to Ag/AgCl (3M/saturated KCl) as standard and Pt wire as an auxiliary electrode through a potentiostat to construct an organophosphorus (OP) biosensor. The biosensor was based on inhibition of AChE by OP compounds/insecticides. The biosensor showed optimum response at pH 7.0, 30°C, when polarized at +0.2V. Two OP compounds, malathion and chlorpyrifos could be detected in the range of 0.1-100 nM and 0.1-70 nM, respectively at 2.0-3.0% inhibition level of AChE. The sensor was reactivated by immersing it in 0.1 mM 2-pyridine aldoxime for 10 min. The detection limit of the sensor was 0.1 nM for both malathion and chlorpyrifos. The biosensor exhibited good reusability (50 times without considerable loss) and storage stability (50% within 60 days, when stored at 4°C).

Construction of triglyceride biosensor based on nickel oxide-chitosan/zinc oxide/zinc hexacyanoferrate film.

A method is described for construction of an amperometric triglyceride (TG) biosensor based on co-immobilization of lipase, glycerol kinase (GK) and glycerol-3-phosphate oxidase (GPO) onto nickel oxide nanoparticles (NiONPs)-chitosan (CHIT) nanocomposite adsorbed onto zinc oxide/zinc hexacyanoferrate (ZnO-ZnHCF) hybrid film electrodeposited on the surface of an Au electrode. The NiONPs-CHIT/ZnO-ZnHCF hybrid film was characterized by cyclic voltammetry (CV), atomic force microscopy (AFM) and electrochemical impedance spectroscopy (EIS). The biosensor showed optimum response within 4 s at pH 6.0 and 35 °C, when polarized at +0.4 V against Ag/AgCl. There was a linear relationship between sensor response and triolein concentration in the range 50-700 mg/dl with sensitivity of 0.05 μA/mg/dl. The sensor was employed for determination of TG in serum. The detection limit of the biosensor was 10 mg/dl. The biosensor was evaluated with 95-96% recovery of added triolein in sera and 2% and 3% within and between batch coefficients of variation (CVs) respectively. There was a good correlation (r=0.99) between serum TG values by standard enzymic colorimetric method and the present method. The biosensor lost 50% of its initial activity after its 100 uses over a period of 180 days, when stored at 4 °C.

Development of amperometric lysine biosensors based on Au nanoparticles/multiwalled carbon nanotubes/polymers modified Au electrodes

The construction of two amperometric l-lysine biosensors is described in this study. The construction comprises the covalent immobilization of lysine oxidase (LOx) onto nanocomposite composed of gold nanoparticles (AuNPs) and carboxylated multiwalled carbon nanotubes (c-MWCNT), decorated on (i) polyaniline (PANI) and (ii) poly 1,2 diaminobenzene (DAB), electrodeposited on Au electrodes. The biosensors were characterized by scanning electron microscopy (SEM), Fourier transform infrared (FTIR) and electrochemical impedance spectroscopy (EIS) studies. The optimum response (current) was observed within 2 s at pH 7.0 and 25 °C for LOx/AuNPs/c-MWCNT/PANI/Au, and 4 s at pH 7.0 and 30 °C for LOx/AuNPs/c-MWCNT/DAB/Au electrodes. There was a linear relationship between current and lysine concentration ranging from 5.0 to 600 μM for LOx/AuNPs/c-MWCNT/PANI/Au with a detection limit of 5.0 μM, and 20 to 600 μM for the LOx/AuNPs/c-MWCNT/DAB/Au electrode with a detection limit of 20 μM. The PANI modified electrode was in good agreement with the standard HPLC method, with a better correlation (r = 0.992) compared to the DAB modified electrode (r = 0.986). These observations revealed that the PANI modified Au electrode was better than the DAB modified electrode, and hence it was employed for the determination of lysine in milk, pharmaceutical tablets and sera. The PANI modified electrode showed a half life of 120 days, compared to that of 90 days for the DAB modified electrode, after their 100 uses, when stored at 4 °C.

An amperometric glutathione biosensor based on chitosan-iron coated gold nanoparticles modified Pt electrode.

A method is described for development of an amperometric biosensor for determination of glutathione (GSH), by immobilizing covalently a glutathione oxidase (GSHOx) onto the surface of gold coated magnetic nanoparticles (Fe@AuNPs) modified Pt electrode. Chitosan was used to introduce amino groups onto the surface of Fe@AuNPs. The morphology and covalent linkage of GSHOx led to high enzyme loading and better shelf life. The enzyme electrode was characterized by scanning electron microscopy (SEM), cyclic voltammetry (CV), Fourier transform infrared (FTIR) and electrochemical impedance spectroscopy (EIS). The electrode showed maximum response within 4s, when polarized at +0.4V, pH 7.0 and 25°C. There was a linear relationship between electrode response and glutathione concentrations in the range 5.0-4000 μmol L(-1) with a detection limit of 0.1 μmol L(-1). An amperometric method of GSH determination was developed using this biosensor. The evaluation studies showed that the method was reliable as mean analytic recoveries of 50 μM and 100 μM of GSH were 97.5±1.7 and 96.1±1.3 respectively and within and between CVs for glutathione determination in blood RBCs were <2.14% and <2.39% respectively. The biosensor showed 50% loss in its initial activity after its 150 uses over a period 4 months, when stored at 4°C. GSH concentration in hemolysated erythrocytes as measured by the present biosensor was 2.8 mmol L(-1) in apparently healthy persons.

Determination of serum triglyceride by enzyme electrode using covalently immobilized enzyme on egg shell membrane.

A mixture of commercial lipase, glycerol kinase and glycerol-3-phosphate oxidase was co-immobilized onto egg shell membrane through covalent coupling. A method is described for fabrication of a triglyceride (TG) biosensor using egg shell membrane bound enzymes. The biosensor measured current, i.e. flow of electrons generated from H(2)O(2), maximally when polarized at 0.4V. The biosensor showed optimum response within 10 sec at pH 7.0 and 35°C. The current was in proportion to concentration of TG in the range 0.56-2.25 mM. An amperometric method was developed for determination of TG employing this enzyme electrode. The minimum detection limit of the method was 0.28 mM. The analytic recovery of added TG was 95.00% and 96.50%. Within batch and between batch coefficients of variations (CV) were <2.14% and <3.48% respectively. A good correlation (r=0.985) was obtained between serum TG level by standard enzymic colorimetric method and the present method. Serum substances such as urea, uric acid, glucose, cholesterol, ascorbic acid and pyruvic acid had no interference. The enzyme electrode was used 200 times over a period of 70 days without any considerable loss of activity, when stored at 4°C.

Electrochemical impediometric detection of anti-HIV drug taking gold nanorods as a sensing interface.

In present work, gold nanorods were used for amplification of electrochemical sensing of anti-HIV replication drug i.e. deferiprone. Gold nanorods (nano Au) deposited onto pencil graphite electrode (PGE) has been utilized for covalent immobilization of horse radish peroxidase (HRP), via glutaraldehyde (Glu), for deferiprone detection using impedimetric technique. Gold nanorods (nano Au) prepared were characterized by TEM and XRD. The resulting nano Au sensor exhibited a good response to deferiprone with a wide linear range (0.005-1000µM) and a low detection limit 0.005µM. The biosensor also showed a short response time (within 15s). In addition, the biosensor exhibited high reproducibility, good storage stability and anti-interference ability. The applicability of the nano Au sensor is to determine deferiprone level in spiked urine and serum samples.