Cyclocarya paliurus has been used for drug formulations and ingredients in functional foods in China. Field studies were conducted to examine the relationships between environmental factors and flavonoid accumulation. A split-plot randomized design was used to establish three shading treatments and three fertilization levels, and growth parameters and flavonoid contents were detected. The greatest biomass production was achieved in intermediate shade and fertilization treatment, and leaf production per seedling increased by 139.5% compared to the treatment without shade and fertilization. Overall, shade and fertilization had a significantly negative effect on contents of total flavonoid, kaempferol, quercetin, and isoquercitrin in leaves of C. paliurus. However, the greatest accumulation of total flavonoid in the leaves was observed in intermediate shade and fertilization treatment, achieving 364.4 g/plant. The results suggest that manipulating the field growing conditions and optimizing the silvicultural system would be important for obtaining the greatest yield of targeted health-promoting substances.