A lactate biosensor based on lactate dehydrogenase/nictotinamide adenine dinucleotide (oxidized form) immobilized on a conducting polymer/multiwall carbon nanotube composite film

An amperometric lactate biosensor was developed based on a conducting polymer, poly-5,2′-5′,2′′-terthiophene-3′-carboxylic acid (pTTCA), and multiwall carbon nanotube (MWNT) composite on a gold electrode. Lactate dehydrogenase (LDH) and the oxidized form of nicotinamide adenine dinucleotide (NAD⁺) were subsequently immobilized onto the pTTCA/MWNT composite film. The modified electrode was characterized by quartz crystal microbalance (QCM), scanning electron microscopy (SEM), and electrochemical experiments. The detection signal was amplified by the pTTCA/MWNT assembly onto which a sufficient amount of enzyme was immobilized and stabilized by the covalent bond formation between the amine groups of enzyme and the carboxylic acid groups of the pTTCA/MWNT film. Experimental parameters affecting the sensor responses, such as applied potential, pH, and temperature, were assessed and optimized. Analytical performances and dynamic ranges of the sensor were determined, and the results showed that the sensitivity, stability, and reproducibility of the sensor improved significantly using pTTCA/MWNT composite film. The calibration plot was linear (r² = 0.9995) over the range of 5 to 90 μM. The sensitivity was approximately 0.0106 μA/μM, with a detection limit of 1 μM, based on a signal/noise ratio of 3. The applicability of the sensor for the analysis of l-lactate concentration in commercial milk and human serum samples was demonstrated successfully.