Understanding the variations of silicon isotopes in terrestrial higher plants can be helpful toward elucidating the global biogeochemical silicon cycle. We studied silicon isotope fractionation in rice and cucumber plants over their entire life cycles. These two different silicon‐absorbing plants were grown hydroponically at different external silicon concentrations. The ranges of δ30Si values in rice were −1.89‰ to 1.69‰, −1.81‰ to 1.96‰, and −2.08‰ to 2.02‰ at 0.17 mM, 1.70 mM, and 8.50 mM silicon concentrations, respectively. The ranges of δ30Si values in cucumber were −1.38‰ to 1.21‰, −1.33‰ to 1.26‰, and −1.62‰ to 1.40‰ at 0.085 mM, 0.17 mM, and 1.70 mM external silicon concentrations, respectively. A general increasing trend in δ30Si values from lower to upper plant parts reflected the preferential incorporation of lighter silicon isotopes from transpired water to biogenic opal. Furthermore, the active uptake mechanism regulated by several transporters might have also played an important role in the preferential transport of heavy silicon isotopes into aboveground plant parts. This suggested that silicon isotope fractionation in both rice and cucumber was a Rayleigh‐like process. The data on δ30Si values for the whole plants and nutrient solutions indicated that biologically mediated silicon isotope fractionation occurred during silicon uptake by roots. At lower external silicon concentrations, heavy silicon isotopes entered plants more readily than light silicon isotopes. Conversely, at higher external silicon concentrations, light silicon isotopes entered plants more readily than heavy silicon isotopes.