Performance of nanopolyaniline-fungal enzyme based biosensor for water pollution

B.S. Kushwah1*, S.C. Upadhyaya2, Shipra Shukla3, Apurva Singh Sikarwar3, R.M.S. Sengar4, Seema Bhadauria5

1Department of Mechanical Engineering, Faculty of Engineering and Technology, Bichpuri, R.B.S. College, Agra, India

2Department of Optical Instrumentation, Dau Dayal Institute of Vocational Education, Dr. B. R. Ambedkar University, Agra, India

3Department of Biotechnology, R.B.S. College, Agra, India

4Department of Botany, Agra College, Agra, India

5Department of Botany, R.B.S. College, Agra, India

Adv. Mater. Lett., 2012, 2 (1), pp 43-51

DOI: 10.5185/amlett.2010.8149

Publication Date (Web): Apr 08, 2012

E-mail: b_kushwah@rediffmail.com

Abstract


The Laccases are oxidoreductases belonging to the multinuclear copper-containing oxidases; they catalyse the monoelectronic oxidation of substrates at the expense of molecular oxygen. These essentially ecofriendly enzymes work with air and produce water as the only by-product. Their uses span from the textile to the pulp and paper industries, and from food applications to bioremediation processes. Laccases also have uses in organic synthesis, where their typical substrates are phenols and amines, and the reaction products are dimers and oligomers derived from the coupling of reactive radical intermediates. Laccase from Pleurotus ostreatus was extracted from the Shaken flask cultures of Pleurotus ostreatus and grown at 25°C with continuous agitation (110 rpm) in baffled Erlenmeyer flasks (1000 mL) containing 200 mL medium. The basal glucose yeast extract peptone agar medium (GYP medium) used for cultures unless otherwise stated contained 20 g glucose L-1, 5 g yeast extract L-1, 5 g peptone L−1 and 1 g MgSO4.7H2O L−1. The pH was adjusted to 5±0 with H3PO4 prior to sterilization. The kinetics of oxidation reactions catalyzed by laccase was studied using 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid (ABTS). The laccase showed lower specific activity. Enzyme modified electrodes were fabricated with polyaniline. Electrochemical polymerization of aniline was performed to get the film of polymer on the surface of glass electrode. First, ITO/PANI electrode was reduced by a 15 min cathodic polarization of the sensor at -500 mV in 0.1 M acetate buffer, pH 5.5. After cathodic polarization, the film was immersed in 0.1 M of acetate buffer, pH 5.5 containing enzyme solution for the deposition of enzyme in polymer layer at +650 mV for 20 min. During this oxidation process laccase become electrostatically attached to polymer film. The ITO/PANI/LAC electrode was rinsed with deionised water to remove any loosely bounded enzyme, and stored in buffer solution at 4oC, when not in use. Conducting polymer/enzyme modified electrodes prepared by immobilization of enzyme were tested for electrocatalytic activities towards amperometric sensing of phenol in industrial effluent.

Keywords

Polyaniline, laccase, nanotechnology, amperometric analysis, phenol biosensor.

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