Tyrosinase is inextricably related to the development of Alzheimer’s disease. The effects of natural tyrosinase inhibitors on human health have attracted widespread attention. This study aimed to isolate and analyze the tyrosinase (TYR) inhibitory peptides in the enzymatic digestion products of royal jelly. We first analyzed optimal process conditions for the enzymatic digestion of royal jelly by single-factor and orthogonal experiments and then used gel filtration chromatography to obtain five fractions (D1~D5) with molecular weights ranging from 600 to 1100 Da. LC-MS/MS was applied to identify the fractions with the highest activity, and the obtained peptides were screened and molecularly docked using AutoDock Vina. The results showed that the optimal enzymatic conditions for tyrosinase inhibition rate were acid protease, enzyme addition 10,000 U/g, initial pH 4, feed-to-liquid ratio 1:4, enzymatic temperature 55 °C, and enzymatic time 4 h. The D4 fraction had the most significant TYR inhibitory activity. The IC50values of the three new peptides with the strongest TYR inhibitory activity, TIPPPT, IIPFIF, and ILFTLL, were obtained as 7.59 mg/mL, 6.16 mg/mL, and 9.25 mg/mL, respectively. The molecular docking results showed that aromatic and hydrophobic amino acids were more favorable to occupy the catalytic center of TYR. In conclusion, the new peptide extracted from royal jelly has the potential to be used as a natural TYR inhibitory peptide in food products with health-promoting properties.