Biosensor with L-phenylalanine Dehydrogenase and Diaphorase Immobilized on Nanocomposite Electrode for the Determination of L-phenylalanine in Urine

Bienzymatic amperometric biosensor consisting of L-phenylalanine dehydrogenase and diaphorase for the determination of L-phenylalanine has been developed. Enzymes were immobilized between chitosan layers onto the surface of planar nanocomposite electrodes based on multi-walled carbon nanotubes. Wide linear concentration range was obtained for 12–1250 μM of L-phenylalanine in phosphate buffer solution of pH 9.0 and in the presence of 2 mM NAD and 2.5 mM ferricyanide mediator. The limit of detection was 6.4 μM, the sensitivity 22.3 nA μM cm and the response time within 60 s. The biosensor exhibited good storage (90% of initial sensitivity after 12 months) and operational stabilities (sensitivity above 90% after 5 days of use). It is proposed for the potential use in self-monitoring of phenylalanine in urine by phenylketonurics. A low signal was measured when tested for urine samples of healthy people, which indicated a low susceptibility for various interferences. Recovery values of about 100% were obtained after spiking urine samples with phenylalanine.