The Argentine midfielder did not fly out to Florence for the Group E match after pulling a hamstring in training, which means Steven Gerrard may be used in a less advanced position to cover.
"It's not a problem. We can manage with Stevie or another player," Benitez told a news conference, adding that Mascherano should only be out for a few days.
"It's not good to lose a player like Javier but we'll try to find a solution. There are three or four others who can play there."
Fellow midfielder Yossi Benayoun should be fit, Benitez said, after reports that the Israeli had suffered a leg injury during Liverpool's 6-1 demolition of Hull City on Saturday.
The five-times European champions have roared back to form after a shaky start by winning their last four league games.
But Benitez admitted his free-scoring side still have work to do to regain the defensive solidity they displayed in previous seasons.
"It's a question of little things that we can fix, if we keep working together," he said.
"It's a question of time to find the right balance. Sometimes when you're attacking a lot you have more problems in defence."
TAKE HEART
Liverpool beat Hungarian newcomers Debrecen 1-0 in their first group match, with Dirk Kuyt scoring the winner and the Dutchman said Liverpool would take heart from their last trip to Italy when they beat Inter Milan 1-0.
"That was a great night in the San Siro, I enjoyed every minute of it and hopefully we can do the same against Fiorentina," he told Liverpool's website.
"It will be great if we can win in Florence and get six points from the first two games. It would give us more confidence ahead of the other group games."
Fiorentina coach Cesare Prandelli said he would need good luck to shut out Fernando Torres after the Spain striker continued his hot form with a hat-trick against Hull.
"You stop him with good organisation, tactical order and lots of fortune," said Prandelli. "He's an extraordinary player. You have to hope you don't meet him on a good night."
Fiorentina lost their opening group game 1-0 at Olympique Lyon but their Serie A form is good, especially at home. They moved to joint third with Inter Milan by beating Livorno 1-0 on Saturday despite the turmoil of president Andrea Della Valle quitting last week.
Prandelli faces Liverpool as a coach after playing in the Juventus side that beat the English club in the 1985 European Cup final on the night of the Heysel tragedy.
"The word Heysel implies particular things that have nothing to do with Liverpool or the game of football," Prandelli said.
"Meeting Liverpool again does not take me back to that dramatic experience. Now I want to transmit the joy of playing against a great team like Liverpool."
