Abstract: Abstract: Pepper seedlings were grown for 30 d with 58±1 μmol/(m2•s) light intensity under different photoperiod (day/night) of 24 h (P24,14 h/10 h), 12 h(P12,7 h/5 h) and 6 h (P6,3.5 h/2.5 h), and different photon flux ratios of red and blue light of 7(7RB) and 1(RB). The measurements included plant morphology, growth, the chlorophyll content, photosynthesis and chlorophyll fluorescence. Pepper seedlings under P12-7RB showed the highest shoot dry weight and the largest leaf area. The relative growth rates (RGR) and specific leaf area (SLA) ratio of P6-RB were 55.88% and 23.62% higher than P24-RB, respectively. P6-RB showed higher quantum efficiency of Photosystem II (ΦPSII) (0.68 mol/(m2•s)) and lower non-photochemical quenching (NPQ) (0.17 μmol/mol) compared with P24-RB, although P6-RB and P24-RB had no significant difference among photosynthesis (Pn), stomatal conductance (Gs), CO2 absorption (Ci) and transpiration (Tr). There was no significant difference in terms of chlorophyll contents among the P6, P12 and P24 treatments. Seedlings of RB had significantly better photosynthetic performance compared with those of 7RB. Experiments showed that peppers could adapt to unnatural different photoperiods. The light parameter P6-RB was beneficial to the growth of pepper seedlings and could be efficiently used in plant factory. The results provide a theoretical basis for long photoperiod applications in a closed system.