Italian Midfielder Marco Materazzi put Italy in front with a towering first-half header in the Group E match in Hamburg before his fellow substitute Filippo Inzaghi finished a breakaway attack in the 87th minute to ensure Italy finished top of the group.
Italy only needed a draw to advance and their task was made easier in first-half stoppage time when the Czech Republic's Jan Polak was sent off for receiving a second yellow card.
The Czechs, who needed to win to progress, enjoyed spells of pressure but when they did breach the Italian defence they found keeper Gianluigi Buffon in imperious form. They waned in the second half and lacked the impetus to turn the game around.
Czech coach Karel Brückner said: "It's not a big disappointment. We knew from the start that we weren't in an easy group. It's over, what can I do about it?" The Czechs had suffered a major blow in their first Group E match against the USA when they lost their key striker, Jan Koller, through a leg injury that put him out of the tournament.
The Czechs threatened first after a sublime pass from Pavel Nedved opened up a statuesque Italian defence. The former European Footballer of the Year, who has played in Italy for the last 10 years, clipped the ball over Fabio Cannavaro's head and into the path of Baros whose angled shot was blocked by Buffon.
Baros was recalled to the Czech's starting line-up after missing the first two matches with a foot injury. Buffon had to be alert again moments later, diving low to gather Nedved's speculative drive.
The Czechs maintained the pressure and almost made the breakthrough after a robust run by Nedved. The Czech playmaker pulled the trigger from 20 yards and Buffon got down low to palm the ball away, but only as far as Marek Jankulovski.
The marauding left-back, who also plays in Italy, tried to steer the ball across goal but Buffon snuffed out the danger with his shins. The Italians were forced into making a substitution in the 17th minute when central defender Alessandro Nesta limped off feeling his groin.
Materazzi came on and nine minutes later he was on the scoresheet. Francesco Totti swung over a corner from the right and Materazzi jumped highest to plant a firm, downward header past Petr Cech.
Karel Poborsky rolled a shot harmlessly wide as the Czechs tried to muster a reply, but the hill they had to climb became a mountain when Polak was sent off in first-half stoppage time after clattering into Totti.
Totti almost extended Italy's lead in the 53rd minute, but his rasping effort was kept out by Peter Cech's right hand.
The Czechs responded immediately with a swift counter-attack which put Nedved through on goal, but his shot was too close to Buffon.
In the first match of the knockout phase, Italy will face the team that comes second in Group F. That will be determined later on Thursday at 2100 CET when Brazil meets Japan and Australia plays Croatia.