Graphdiyne　(GDY)　and　graphene　are　regarded　as　two　promising　two-dimensional　carbon-based　materials,　which　have　unique　planar　structure　and　novel　electronic　properties.　Differences　between　the　two　carbon　allotropes　in　their　physicochemistry　biology　and　cytotoxicity　have　never　been　explored.　Here,　we　chemically　functionalized　the　surface　of　the　two　carbon　allotropes　using　similar　oxidation　processes　and　compared　their　physicochemistry,　biology,　and　mutagenesis.　Graphene　oxide　(GO)　and　GDY　oxide　(GDYO)　showed　similarities　in　their　size,　morphology,　and　physical　spectral　characteristics,　excepting　the　differences　in　sp-　and　sp(2)-hybridizations　and　Fourier　transform　infrared　spectroscopy.　GDYO　was　well　soluble　in　various　media.　In　contrast,　GO　was　only　soluble　in　H2O,　but　kinetically　aggregated　in　0.9%　NaCl,　phosphate　buffered　saline,　and　cell　media　within　24　h　incubation　when　its　concentrations　increased.　GO　nanoparticles　adhered　and　aggregated　to　the　surface　of　a　human　hepatocyte　membrane,　resulting　in　cell　membrane　ruffle,　methuosis,　and　apoptosis.　Adhesion　of　GO　to　cells　caused　cell　stress　and　induced　reactive　oxygen　species.　In　contrast,　GDYO　did　not　adhere　to　the　cell　membrane　to　produce　the　related　consequences.　Both　GDYO　and　GO　showed　in　vivo　mutagenesis　potential　but　no　erythrocyte-killing　effect,　and　both　were　antioxidant　and　bioequivalent　at　binding　to　single-stranded　DNA　and　doxorubicin,　thus　causing　fluorescence　quenching.　The　present　study　significantly　enriches　our　existing　knowledge　of　GO/alkene　and　GDYO/alkyne　chemistry.