Matrix-assisted　laser　desorption/ionization　(MALDI)　mass　spectrometry　(MS)　holds　great　promise　for　the　rapid　and　sensitive　detection　of　biomolecules,　but　its　precise　detection　of　small　molecule　metabolites　is　hindered　by　severe　background　interference　from　the　organic　matrix　in　the　low　molecular　weight　range.　To　address　this　issue,　nanomaterials　have　commonly　been　utilized　as　substrates　in　LDI-MS.　Among　them,　covalent　organic　frameworks　(COFs),　known　for　their　unique　optical　absorption　and　structural　properties,　have　garnered　significant　attention.　Despite　these　advantages,　their　low　ionization　efficiency　remains　a　challenge.　Herein,　a　composite　material　of　COF-S@Au　nanoparticles　(NPs),　by　incorporating　Au　NPs　into　a　sulfur-functionalized　COF　(COF-S)　through　postsynthetic　modification,　was　designed　and　adopted　as　substrates.　This　hybrid　material　leverages　the　synergistic　effects　of　COF-S　and　Au　NPs　to　improve　the　desorption/ionization　efficiency　and　minimize　background　interference.　The　COF-S@Au　NPs　demonstrated　a　5-16-fold　improvement　in　MS　signals　of　small　biomolecules　along　with　a　clean　background　and　excellent　resistance　to　salt　and　protein　interference.　Their　corresponding　limits　of　detection　(LODs)　were　achieved　at　∼pmol.　Furthermore,　the　COF-S@Au　NPs　were　applied　to　analyze　metabolites　in　a　triclosan　(TCS)-exposed　mouse　model,　successfully　identifying　10　differential　metabolites　associated　with　TCS　toxicity.　This　work　provides　a　foundation　for　developing　advanced　LDI-MS　materials　for　high-performance　metabolic　analysis　and　offers　valuable　insights　into　TCS　metabolic　toxicity　with　potential　applications　in　environmental　toxicology.　©　2025　American　Chemical　Society.