In this article, a novel and efficient analysis method is described to simultaneously and quantitatively determine glucoraphanin and its major degradation product sulforaphane by reversed-phase high-performance liquid chromatography (HPLC) coupled with an evaporative light-scattering detector (ELSD). This method was validated according to the regulatory guidelines with respect to precision, accuracy and linearity. The HPLC-ELSD method was successfully used to assess the contents of glucoraphanin and sulforaphane in varieties of Brassica oleracea seeds. The developed analytical method not only avoids the underestimation of total glucoraphanin content, but also provides a reduction in analysis time and good efficiency and sensitivity compared to with conventional HPLC methods.