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Tubeimoside I(TBMS1)was isolated fromthe tubers of Bolbostemma panieulatum(Maxim.)Franquet.TBMS1shows potent antitumor activity.The present studywas conducted to investigate the antimicrotubule roleof TBMS1 and its binding site oftubulin.Methods：Cell growth inhibition Was measuredby MTT after treatment with TBMS1.Uptake kinetics ofTBMS1 bv humannasopharyngeal carcinoma CNE-2Z cell line(CNE-2Z)Wasassayed by HPLC.Microtubule protein(MTP)Was preparedfrom porcinebrain through two cycles of polymerization-depolymerization in a high molaritybuffer.Inhibition of MTP polymerization induced byTBMS1 Was determined by a turbidity measurement and asedimentation assay；the interactions of TBMS1 withtubulin within CNE-2Zcells were investigated by immunofluoresceneemicroscopy and immunoblotting.TBMS1 Was tested forits ability to inhibit binding ofknown tubulin ligands through competitive bindingassay.Results：TBMS1 displayed growth inhibitoryactivity against CNE-2Z cellswith IC50 value of 16.7 μM for 72 h.HPLC analysis ofTBMS1 uptake by CNE-2Z eeHs displayed the initialslow TBMS1 uptake andthen gradually reaching an maximum uptake neax 18h.CNE-2Z cells treated with TBMS1(25μM，3 h)weresufficient to cause themicrotubular network disruption.Immunoblot analysisshowed that the proportion of cytosolic tubulin ofceils treated with TBMS1increased in a time-and concentration-dependentmanner.TBMS1 did not inhibit the binding ofvinblastine tO tubtdin.Colchicinebinding to tubulin was inhibited in the presence ofTBMS1.Conclusions：TBMS1 is an anti-mierotubuleagent，and its binding site oftubulin is the colchicine binding site of tubulin.