A bismuth oxide-carboxylated multi-walled carbon nanotube/glassy carbon electrode (Bi2O3-CMWCNTs/GCE) was prepared for the detection of the flavonoid drug baicalein. The Bi2O3 was grown on the surface of CMWCNTs by in-situ synthesis, and a simple, and efficient Bi2O3-CMWCNTs composite was prepared. The materials were characterized by XRD, SEM and EDS. CV and EIS techniques were used to study the electrochemical sensing performance of the composite electrode surface. The electrochemical performance and optimal detection conditions of baicalein on Bi2O3-CMWCNTs/GCE were investigated by LSV. The oxidation peak current of baicalein has a linear relationship with its concentration in the range of 0.01–15 μmol·L−1, with a detection limit of 2.0 nmol·L−1. The method is both stable and effective, and was successfully used for determination of baicalein in human urine and the Chinese herb Oroxylum indicum. It shows that the in situ synthesized materials have more stable characteristics, and have promising applications in the detection of flavonoid drugs.