Matrix-assisted　laser　desorption/ionization　time-of-flight　mass　spectrometry　(MALDI-TOF　MS)　has　become　an　indispensable　tool　for　high-throughput　analysis　of　macromolecules,　but　many　challenges　still　remain　in　detection　of　small　molecules　due　to　the　severe　matrix-related　background　interference　in　the　low-molecular-weight　ranges　(MW　＜　700　Da).　Herein,　a　gallic　acid　(GA)-functionalized　zirconium　1,4-dicarboxybenzene　metal-organic　framework　(MOF)　(denoted　as　UiO-66-GA)　was　designed　to　serve　as　a　new　substrate,　and　a　novel　strategy　on　the　basis　of　the　synergistic　effect　of　MOF　and　GA　was　developed　to　enhance　the　LDI　process.　In　comparison　with　conventional　organic　matrices,　the　UiO-66-GA　substrate　showed　superior　LDI　performance　in　the　analysis　of　a　wide　variety　of　molecules　including　amino　acids,　unsaturated　fatty　acids,　bisphenols　(BPs),　oligosaccharides,　peptides,　protein,　and　polyethylene　glycol　(PEG)　of　various　average　molecular　weights　from　200　to　10000.　Perfluorooctanoic　sulfonate　(PFOS)　was　used　to　evaluate　the　ability　of　quantitative　analysis,　and　its　corresponding　limit　of　detection　as　low　as　1　fmol　was　achieved.　High　sensitivity　and　good　salt　tolerance　of　the　UiO-66-GA-assisted　LDI-MS　were　allowed　to　determine　ultratrace　PFOS　in　the　spiked　human　urine　and　serum　samples.　In　addition,　the　synergistic　mechanism　of　MOF　and　GA　in　the　enhanced　LDI　process　was　investigated　by　comprehensively　comparing　GA-and　its　analogue-functionalized　UiO-66,　and　the　results　revealed　that　two　aspects　contributed　to　the　enhanced　LDI　process:　(1)　an　enhancement　in　the　metal-phenolic　coordination　system　of　UiO-66-GA　promoted　laser　absorption　and　energy　transfer;　(2)　introduction　of　carboxyl　and　hydroxyl　groups　of　GA　onto　UiO-66　facilitated　the　LDI　process　in　both　positive　and　negative　ion　modes.　This　work　expands　a　new　domain　for　the　MOF　applications　and　provides　a　promising　alternative　for　various　molecule　analyses.　Copyright　©　2019　American　Chemical　Society.