Royal　jelly　(RJ)　is　a　biologically　active　substance　secreted　by　the　hypopharyngeal　and　mandibular　glands　of　worker　honeybees.　It　is　widely　claimed　that　RJ　reduces　oxidative　stress.　However,　the　antioxidant　activity　of　RJ　has　mostly　been　determined　by　in　vitro　chemical　detection　methods　or　by　external　administration　drugs　that　cause　oxidative　stress.　Whether　RJ　can　clear　the　endogenous　production　of　reactive　oxygen　species　(ROS)　in　cells　remains　largely　unknown.　Here,　we　systematically　investigated　the　antioxidant　properties　of　RJ　using　several　endogenous　oxidative　stress　models　of　Drosophila.　We　found　that　RJ　enhanced　sleep　quality　of　aging　Drosophila,　which　is　decreased　due　to　an　increase　of　oxidative　damage　with　age.　RJ　supplementation　improved　survival　and　suppressed　ROS　levels　in　gut　cells　of　flies　upon　exposure　to　hydrogen　peroxide　or　to　the　neurotoxic　agent　paraquat.　Moreover,　RJ　supplementation　moderated　levels　of　ROS　in　endogenous　gut　cells　and　extended　lifespan　after　exposure　of　flies　to　heat　stress.　Sleep　deprivation　leads　to　accumulation　of　ROS　in　the　gut　cells,　and　RJ　attenuated　the　consequences　of　oxidative　stress　caused　by　sleep　loss　and　prolonged　lifespan.　Mechanistically,　RJ　prevented　cell　oxidative　damage　caused　by　heat　stress　or　sleep　deprivation,　with　the　antioxidant　activity　in　vivo　independent　of　Keap1/Nrf2　signaling.　RJ　supplementation　activated　oxidoreductase　activity　in　the　guts　of　flies,　suggesting　its　ability　to　inhibit　endogenous　oxidative　stress　and　maintain　health,　possibly　in　humans.　©　2023　Institute　of　Zoology,　Chinese　Academy　of　Sciences.