In order to explore the allelopathy effect of forest gap litter on the natural regeneration of Castanopsis kawakamii seedlings, we illustrated forest gap, litter allelopathy effect, and their interactions on the seedling height, chlorophyll fluorescence parameters, and osmotic adjustment substance of C. kawakamii. In this experiment, we simulated the light intensity of different gaps and non-gaps, and used aqueous extracts of litter as the main allelochemicals with different concentrations' treatments (1:10, 1:30, 1:50 and 1:100). The results showed that:(1) the relative chlorophyll content, PSⅡ potential activity (F_v/F_o), PSⅡ maximal photochemical efficiency (F_v/F_m), and the soluble sugar content of seedlings in non-gaps were higher than those of other gaps, whereas the seedlings height on the late stage and initial fluorescence (F_o) were lower than those of other gaps. (2) Comparing with the control treatment, the litter extracts with different concentrations decreased the F_v/F_o value, while the concentrations of 1:50 and 1:100 increased seedling height in the late stage, the soluble protein content and relative chlorophyll content. (3) The formation of forest gap improved seedling height under the different concentrations of litter extract in general. In addition, the relative chlorophyll content, F_v/F_o, and F_v/F_m under different concentrations in small gaps and non-gaps were generally higher than those in other gaps. (4) The results of entropy method indicated that the comprehensive evaluation value of C. kawakamii seedling in non-gaps was higher than that of other gaps. The influences of litter extract concentration on seedlings growth expressed an obvious trend with "low promotion and high inhibition", and the comprehensive evaluation value of 1:50 concentration was higher than that of other concentrations. Under the higher concentrations (1:10 and 1:30), the comprehensive evaluation value in medium gaps was higher than that of other gaps. It concluded that forest gap and litter allelopathic effect affected the seedling growth of C. kawakamii mutually, and the medium gaps (50-100 m²) weakened the toxic effect with higher extract concentration of litter on the seedling growth. In the future, we could regulate gap size, and clear the forest litter by human interference to promote the natural regeneration of C. kawakamii seedlings.