Sclerotinia sclerotiorum is a serious pathogen which causes yield loss in many dicotyledonous crops including potato. The objective of this study was to assess the potential of biofumigation using three Brassica crops including Brassica napus, B. juncea and B. campestris against potato stem rot caused by S. sclerotiorum by in vitro tests. Both macerated and irradiated dried tissues were able to reduce radial growth and sclerotia formation of five pathogen isolates on PDA, but macerated live tissues were more effective. Compared with other tested crops, B. juncea showed more inhibitory effect against the pathogen. The volatile compounds produced from macerated tissues were identified using a gas chromatograph-mass spectrometer. The main identified compounds were methyl, allyl and butyl isothiocyanates. Different concentrations of these compounds inhibited mycelial growth of the pathogen in vitro when applied as the vapor of pure chemicals. A negative relationship was observed between chemicals concentrations and growth inhibition percentage. In this study, it became clear that the tissues of local Brassica crops release glucosinolates and have a good potential to be used against the pathogen in field examinations.