Arabic gum was purified and used as modifier of carbon paste electrode (PAG-CPE) for the simultaneous determination of quercetin (QT) and rutin (RT) in 0.1 M phosphate buffer solution, pH 6.0. Fourier transform infrared spectroscopy was used for the identification of different chemical functional groups present at the surface of the raw and the purified arabic gum. The electrochemical behaviour of QT and RT on the modified electrode was studied using cyclic voltammetry (CV) and square wave voltammetry. The sensitivities of PAG-CPE for both QT and RT were respectively 2.8 and 3.3-fold greater than signals obtained on the unmodified CPE in CV. The linear dynamic range for both flavonoids was found within the concentration range of 0.099–1.090 µmol/L with a detection limit of 0.029 µM and 0.058 µM respectively for QT and RT (3S/m). The proposed approach was successfully applied, first for the direct quantification of QT and RT in crude natural fruits and secondly, for the simultaneous detection of both flavonoids in human urine.