Soybean (Glycine max L.) sprouts are a common vegetable with rich nutrients, such as protein, vitamin C and isoflavones. Soybean is also capable of accumulating selenium (Se). To study the effects of Se biofortification on the nutrient of this crop, soybean sprouts were treated with different concentrations of selenium nanoparticles (SeNPs) or selenite (i.e., 10 μM, 20 μM, 40 μM, 80 μM and 100 μM) in a hydroponic experiment. Results showed that SeNPs and selenite remarkably increased the total Se concentration in soybean sprouts. Five Se speciations, namely, selenocystine, selenomethionine, methyl selenocysteine, selenite and selenate were detected in soybean sprouts, but selenomethionine was found to be the dominant Se speciation. SeNPs and selenite increased the contents of chlorophyll, carotenoid, soluble sugar, soluble protein, vitamin C and isoflavones in soybean sprouts. SeNPs treatments led to less malondialdehyde content compared with selenite. SeNPs and selenite both enhanced the glutathione content. The modest dosage of exogenous Se stimulated the catalase activity, whereas the large amount reduced it. The peroxidase and ascorbate peroxidase activities were stimulated by SeNPs and selenite. SeNPs posed no significant influence on the superoxide dismutase activity. This study suggests that SeNPs are a good exogenous Se source for the production of Se-rich soybean sprouts.