A new effective quantitative screening method was developed to detect sulfonamides (SAs) in foodstuffs of animal origin. The method was based on pulsed amperometric detection (PAD) at a glassy carbon electrode (GCE) coupled with a reversed-phase high performance liquid chromatographic separation (RP-HPLC). First, SAs electrochemical behaviour at GCE was investigated by cyclic voltammetry (CV) experiments. In the light of the CV results, a new three-step waveform for pulsed amperometric detection of SAs was designed and optimized by performing flow injection analysis (FIA) of SAs standard solutions. The optimized PAD waveform ensured an excellent response stability and sensitivity with respect to the direct current detection at constant potential. PAD was then applied to detect SAs after RP-HPLC in gradient mode by using a typical organic mobile phase containing acetate buffer, acetonitrile and methanol. The developed method showed, for all the SAs investigated, very low limits of detection and quantification, in the range 2.1-2.7 μg L-1 and 6.9-9.0 μg L-1, respectively, and a wide linear range (up to 0.5 mg L-1). Method selectivity was tested by comparing the chromatograms of SAs mix standard solutions, blank purified milk samples (without SAs) and spiked purified milk samples. Excellent values of precision under repeatability (1.0-5.0 %, n = 6) and within-laboratory reproducibility (11.2-18.6 %, n = 18) conditions, and recoveries (94-105 %) were obtained.