The biocompatiable and low-toxic poly(thiophene-3-acetic acid)(PTAA) matrix was successfully electrosynthesized in ionic liquid 1-butyl-3-methylimidazolium hexafluorophosphate(BmimPF_6) in comparison with the electrosynthesis of PTAA matrix in acetonitrile(ACN).Ascorbate oxidase(AO) was used as a model for the development and application of biosensor.Vitamin C(VC) biosensors were facilely fabricated by the covalent immobilization of AO molecules on PTAA matrices electrosynthesized in ACN containing tetrabutylammonium tetrafluoroborate and BmimPF_6, respectively.Electrochemical impedance spectroscopy,scanning electron microscopy and FTIR spectroscopy indicated that AO molecules were covalently immobilized on PTAA matrices.Parameters of the as-obtained biosensors such as working potential,pH and temperature have been optimized.The amperometric biosensor based on PTAA matrix electrosynthesized in BmimPF_6 exhibited wider linear range,lower detection limit,higher sensitivity and bioaffinity,and better operational and storage stability than that electrosynthesized in ACN under optimal conditions.The as-obtained biosensor based on PTAA matrix electrosynthesized in BmimPF_6 was employed for the detection of VC content in commercial juices,and the result was close to the data given by manufacturers.Excellent results indicate that the PTAA matrix electrosynthesized in ionic liquid is a promising platform for the covalent immobilization of biologically-active species and the development of biosensors. The biocompatiable and low-toxic poly (thiophene-3-acetic acid) (PTAA) matrix was successfully electrosynthesized in ionic liquid 1-butyl-3-methylimidazolium hexafluorophosphate (BmimPF_6) in comparison with the electrosynthesis of PTAA matrix in acetonitrile (ACN). Ascorbate oxidase (AO) was used as a model for the development and application of biosensor. Vitamin C (VC) biosensors were facilely fabricated by the covalent immobilization of AO molecules on PTAA matrices electrosynthesized in ACN containing tetrabutylammonium tetrafluoroborate and BmimPF_6, respectively. Electrochemical impedance spectroscopy, scanning electron microscopy and FTIR spectroscopy indicated that AO molecules were covalently immobilized on PTAA matrices. Parameters of the as-obtained biosensors such as working potential, pH and temperature have been optimized. amperometric biosensor based on PTAA matrix electrosynthesized in BmimPF_ linear range, lower detection limit, higher sensitivity and bioaffinit y, and better operational and storage stability than that electrosynthesized in ACN under optimal conditions. the as-obtained biosensor based on PTAA matrix electrosynthesized in BmimPF_6 was employed for the detection of VC content in commercial juices, and the result was close to the data given by manufacturers. Excellent results that that the PTAA matrix electrosynthesized in ionic liquid is a promising platform for the covalent immobilization of biologically-active species and the development of biosensors.