In this study, novel dendritic TiO2 nanofibers (D-Ti NFs) with ultrahigh surface area were for the first time synthesized through in situ hydrothermal treatment of the electrospun TiO2 nanofibers (E-Ti NFs) and their microstructure, formation mechanism, in vitro biocompatibility and drug delivery property were investigated. SEM observations showed that D-Ti NFs had dendritic and fibrous morphology with the controllable diameter of 899±26nm to 1955±64nm. TEM observations showed the individual D-Ti NF was assembled by numerous primary nanowires with the diameter of 7±1nm. XRD pattern showed that the D-Ti NFs displayed the characteristic peaks at 24.1°, 28.1°, and 48.3°, all of which were assigned to hydrogen titanium oxide hydrate. BET measurement indicated that D-Ti NFs had the ultrahigh specific surface area of 213.41m2/g. D-Ti NFs showed good in vitro biocompatibility when exposed to Hela cells. After soaked in solution of tetracycline hydrochloride (TH, one of the representative antibiotics, model drug), D-Ti NFs supported the loading of TH with the loading efficiency of 63.73±4.61%. TH-loaded D-Ti NFs supported a sustained release model for TH when immersed in the phosphate buffer saline. After separately incubated with two representative types of bacteria: Escherichia coli and Staphylococcus aureus, TH-loaded D-Ti NFs showed excellent antibacterial property to inhibit the growth of the bacteria, indicating that the released TH was biologically active.