Giourkas Seitaridis was out with a thigh injury, Fanis Gekas fractured a cheekbone in the match against Russia and will be out for six weeks, while Vassilis Torosidis was also injured, Greek FA spokesman Michalis Tsapidis said.
"Giorgios Karagounis is also doubtful," he said. Karagounis has been nursing a leg injury since the start of the tournament.
None of the injuries, apart from that of Karagounis, had been announced before.
Tziolis said anyone coming into the starting XI was ready for the match.
"In football, major success and major failure should be left behind after 24 hours," he said. "All the players want to enjoy themselves against Spain and want to show off their abilities and skills."