Abstract: The electrochemical behaviour of trimethoprim was investigated by differential pulse polarography and cyclic voltammetry. The optimum conditions for the determination of TMP were as follows: The electrolyte was 0.02 mol/L HAc-NaAc (pH 4.0) buffer solution, initial potential -1.00 V(vs.Ag-AgCl), final potential -1.50 V (vs. Ag-AgCl). The calibration graph was linear over the range of TMP concentrations 5.5×10^-7～5.5×10^-6 mol/L. The detection limit was 2.0×10^-7 mol/L.This paper also reports the construction and performance characteristics of a PVC membrane trimethoprim ion selective electrode. It is based on the use of tetraphenylborate-TMP ion pair complex as the active material. The PVC membrane electrode showed Nernstian response over the trimethoprim concentration range from 4.4×10~(-5) to 2.2×10~(-3) mol/L, with a slope of 57.3 mV/decade, the limit of detection being 4.6×10^-6 mol/L. The electrode gave fast response, and good reproducibility. The electrode potential drift was less than 4 mV for a duration of twenty days.The trimethoprim ion selective electrode and differential pulse polarographic methods have been applied to the determination of TMP in pharmaceutical preparations. The electrode can also be used to determine TMP in urine and serum with satisfactory results.