Three Brassicaceae species, Brassica napus (low glucosinolate content), Brassica nigra (including sinigrin), and Sinapis alba (including sinalbin) were used as host plants for two aphid species: the generalist Myzus persicae and the specialist Brevicoryne brassicae. Each combination of aphid species and prey host plant was used to feed the polyphagous ladybird beetle, Adalia bipunctata. Experiments with Brassicaceae species including different amounts and kinds of glucosinolates (GLS) showed increased ladybird larval mortality at higher GLS concentrations. When reared on plants with higher GLS concentrations, the specialist aphid, B. brassicae, was found to be more toxic than M. persicae. Identification of GLS and related degradation products, mainly isothiocyanates (ITC), was investigated in the first two trophic levels, plant and aphid species, by high-performance liquid chromatography and gas chromatography-mass spectrometry, respectively. While only GLS were detected in M. persicae on each Brassicaceae species, high amounts of ITC were identified in B. brassicae samples (allyl-ITC and benzyl-ITC from B. nigra and S. alba, respectively) from all host plants. Biological effects of allelochemicals from plants on predators through aphid prey are discussed in relation to aphid species to emphasize the role of the crop plant in integrated pest management in terms of biological control efficacy.