Defender Alessandro Nesta has been ruled out of Ukraine's clash after failing to recover from a groin injury.
"I visited a clinic here in Duisburg and there's still something not right. It has improved a great deal, but I can't play in this match," Nesta was quoted as saying by the official FIFA website on Thursday.
"However I'm confident of making the next one if we get there. The doctor has given me some hope," he added.
The news is a blow to Lippi as Nesta's usual replacement Marco Materazzi will play no part in the quarter-finals due to suspension after he was sent-off in Italy's 1-0 victory over Australia in the second round.
Daniele De Rossi will be out as he is currently serving a four-match suspension for elbowing United States striker Brian McBride.
Udinese striker Vincenzo Iaquinta, who came off the bench to score Italy's second goal against Ghana in their World Cup opener, will also miss the game as he picked up a slight muscle problem.
Meanwhile, Lippi said his side's match against Ukraine is not an easy one, noting that the opponents have very good individuals at their disposal.
“Ukraine don’t have only [Andriy] Shevchenko, this is a good team with excellent elements who had a fantastic qualification campaign to win a place at the Finals,” Lippi was quoted as saying by Football Italia magazine.
“It’s true that they haven’t played difficult opponents after their Spanish loss in their World Cup opener, but there must be a reason if they are in the last eight," the former Juventus boss added.