PURPOSE: To investigate the contributions of ocular crystalline lens transmittance spectra and pupil size on age-related differences in the magnitude of light-induced melatonin suppression at night. The first aim was to demonstrate that spectral lens transmittance in children can be measured in vivo with a Purkinje image-based system. The second aim was to test the hypothesis that the magnitude of melatonin suppression in children is enhanced by larger pupils and higher lens transmittance of short wavelengths. METHODS: Fourteen healthy children and 14 healthy adults participated in this study. The experiment was conducted for two nights in our laboratory. On the first night, the participants spent time under dim light conditions (<10 lux) until one hour after their habitual bedtime (BT+1.0). On the second night, the participants spent time under dim light conditions until 30 min before their habitual bedtime (BT-0.5). They were then exposed to LED light for 90 min up to BT+1.0. Individual pupil sizes were measured between BT and BT+1.0 for both conditions. Lens transmittance spectra were measured in vivo using the Purkinje image-based system during the daytime. Non-visual photoreception was calculated from lens transmittance and pupil size. This was taken as an index of the influence of age-related ocular changes on the non-visual photopigment melanopsin. RESULTS: Measured lens transmittance in children was found to be higher than for adults, especially in the short wavelength region (p < 0.001). Pupil size in children was significantly larger than that of adults under both dim (p = 0.003) and light (p < 0.001) conditions. Children's non-visual photoreception was 1.48 times greater than that of adults, which was very similar to the finding that melatonin suppression was 1.52 times greater in children (n = 9) than adults (n = 9). CONCLUSIONS: Our Purkinje image-based system can measure children's lens transmittance spectra in vivo. Lens transmittance and pupil size may contribute to differences in melatonin suppression between primary school children and middle-aged adults.