The shell nacre protein was extracted from the nacre layer in the Pinctada martensti shell by a combined technology of acidification-high pressure, with the extraction process optimized through single-factor and response surface experiments using the protein extraction rate as index. The protease hydrolysis products of the shell nacre protein were then prepared, and evaluated for its amino acid composition, anti-inflammatory activity as well as skin repair activity. The results showed that under the optimal extraction conditions of hydrochloric acid concentration 0.18 mol/L, alkali material-liquid ratio (g∶mL) 1∶4.9, mass fraction of NaOH solution 1.00%, extraction temperature 65 ℃, extraction time 3 h, acid precipitation pH 3.5 and resting time 32 h, the extraction rate of shell nacre protein was 17.31%±0.84%. The enzymatic hydrolysate of the shell nacre protein produced via bromelain hydrolysis for 3 h showed no cytotoxicity and was beneficial to the macrophage proliferation. The inhibitory rate of the hydrolysate with a concentration of 800 μg/mL on NO release reached up to 51.85%±1.39%. Meanwhile, the hydrolysate exhibited activities of inhibition against the pro-inflammatory factors IL-6 and TNF-α, and up-regulation of the anti-inflammatory factor IL-10 release, thus demonstrating good anti-inflammatory performance. The enzymatic hydrolysate of the shell nacre contained 17 kinds of amino acids. Among them, the total relative content of 8 kinds of essential amino acids for the human body was 32.626%, the total relative content of 7 kinds of hydrophobic amino acids was 34.513%, while that of 3 kinds of positively charged amino acids was 16.173%. The shell nacre active peptides could significantly improve cell proliferation and cell migration of fibroblasts L929, and improve cell viability induced by H₂O₂, inhibit ROS production, and exhibit good skin repair activity.