Development of a lactate biosensor based on conducting copolymer bound lactate oxidase

Abstract

An amperometric enzyme electrode was developed for determination of lactate in serum. To prepare this electrode, commercial lactate oxidase from Pediococcus species has been immobilized through glutaraldehyde coupling onto polyaniline-co-fluoroaniline film deposited on an Indium tin oxide (ITO) coated glass plate. This plate acted as working electrode when combined with Pt electrode as counter electrode to the electrometer for the development of a biosensor. The method is based on generation of electrons from H 2O 2, which is formed from lactic acid by immobilized lactate oxidase. The concentration of lactic acid is directly proportional to the current measured. The enzyme electrode showed optimum response when operated at 42 °C in 0.05 M, sodium phosphate buffer pH 6.5 for 50 s. The biosensor showed a good performance with a linear response range from 0.1 to 5.5 mM/l. The minimum detection limit of the method is 0.1 mM/l and sensitivity of the sensor is 1.18 μA/mM/l lactate. This electrode was employed for determination of lactate in serum. The serum values in healthy and diseased persons were in the range 0.51–2.9 and 5.0–15.0 mM, respectively. The analytical recovery of added lactic acid was 71%. Within batch and between batch CV were <4 and <5%, respectively. Among the various serum substances tested only 8-hydroxyquinoline, urea, ammonium molybdate and uric acid caused 64, 38, 34 and 31% inhibition, respectively. The electrode was used 150 times over 26 days without any considerable loss of activity, when stored at 4 °C.