Victor Barrio, 29, a professional bullfighter, was killed when the bull's horn pierced his chest.
The fight, in the eastern town of Teruel, was being broadcast live on TV.
Also on Saturday, a 28-year-old man died after being gored by a bull during a bull run through the village of Pedreguer, near Valencia.
The last matador to die in a bullfight in Spain was Jose Cubero, or Yiyo, in 1985.
Television footage shows the bull throwing Barrio into the air before goring him on the right-hand side of his chest and violently throwing him.