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Glucosinolates are an important class of secondary plant metabolites, possessing health‐promoting properties. Young broccoli plants are a very good source of glucosinolates with concentrations several times greater than in mature plants. The aim of our study was to develop a liquid chromatography–mass spectrometry and liquid chromatography/tandem mass spectrometry qualitative and quantitative method for the measure of glucosinolates in broccoli sprouts. The described method provides high sensitivity and specificity, allowing a rapid and simultaneous determination of 14 glucosinolates. The proposed method has been validated for eight glucosinolates: glucobrassicin, glucoraphanin, glucoiberin, glucoerucin, progoitrin, gluconapin, sinigrin and glucocheirolin. The linear range was 1–150 µg ml⁻¹, the intra‐day and inter‐day precision values are within 6% and 8% at the lower limit of quantification, while the …