AbstractA method using reverse phase thin layer and high performance liquid chromatography was developed to determine the level of the bitter principle occasionally found in zucchini (Cucurbita pepo L.). The system gave excellent separation with recovery rates of about 90% from spiked fruit. The bitter principle was confirmed by C13 NMR and mass spectrometry as cucurbitacin E glycoside which was present in concentrations in the order of 600mg kg−1 in bitter fruits. Bitterness was detected by a taste panel at cucurbitacin E glycoside concentrations of 1 mg kg−1 in aqueous solutions and 2 mg kg−1 in zucchini pulp. The method described should assist the screening for bitterness in commercial and breeding populations of zucchini.