In the present study, a peptide showing anti-inflammatory activity was isolated from Arctoscopus (A.) japonicus sandfish protein hydrolysate by hydrolysis and analyzed using ultrafiltration (UF), prep-high-performance liquid chromatography (HPLC), and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Effects of heat, pH, salt, and intestinal proteases on stability of the isolated anti-inflammatory peptide were determined. The nitric oxide (NO) scavenging activities of the peptide from meat and roe hydrolysates were 18.43 ± 3.21% and 52.35 ± 8.85%, respectively, at a concentration of 0.1 mg/mL. The anti-inflammatory peptide maintained a similar level of NO scavenging activity to the control in the presence of salt, whereas it showed no resistance to heat, pH, and intestinal proteases. These results suggest that the anti-inflammatory peptide derived from A. japonicus could be used for producing functional foods, protein supplements, and pharmaceutical agents. However, it is necessary to find methods for protection against heat, pH, and proteolysis.