A tightening mechanism for adjustably setting a tape holder to an immovable state on a tape dispenser includes a tape holder being provided at an axially outer end with a flared round recess, a bushing ring having a bevel wall and being fitted in the flared round recess, and a locating member slightly smaller than the bushing ring for fitly seating in the bushing ring. A turning knob having a spring put thereon is sequentially extended through the locating member and the bushing ring to screw into a fixing bar, on which the tape holder is mounted. When the knob is turned, a tightness of the bushing ring against the flared round recess is progressively changed to thereby adjust the tape holder to different degrees of immobility, allowing the tape dispenser to apply to tapes of different adhesion ability.