Liverpool had led at the Amex through Harvey Elliott, but have now won just once in their last six games.
Lewis Dunk levelled before the break for Brighton and they were well worthy of the victory given to them by a moment of magic from Mitoma.
The Japanese winger cushioned Pervis Estupinan’s cross into his path, flicked the ball over Joe Gomez and fired high past Alisson Becker in the 92nd minute.
Liverpool are languishing in ninth in the Premier League, 21 points behind pacesetters Arsenal.
Their FA Cup exit leaves the Champions League as their only remaining shot at a trophy this season.
Last season Jurgen Klopp’s men were on the brink of a historic quadruple, which included their first FA Cup triumph in 16 years.
But Klopp is still searching for answers to halt their decline after a second defeat on the south coast in 15 days.
The German described Brighton’s 3-0 win when the sides last met as the worst performance of his managerial career.
Liverpool did at least put up a fight this time and will rue two big early chances missed by Mohamed Salah.
The Egyptian did at least have a hand in Liverpool’s goal as he released Elliott, whose strike had too much power for Steele to keep out.
Brighton hit back within nine minutes when Tariq Lamptey’s powerful shot was deflected past the helpless Alisson by Dunk.
The hosts were without Moises Caicedo after the Ecuadorian took to social media on Friday to try and force through a move to Arsenal.
Brighton have already lost Leandro Trossard, who scored a hat-trick against Liverpool earlier in the season, to the Gunners this month.
But those absences did not show as Liverpool were left riding their luck for most of the second period.
Ibrahima Konate and Fabinho were extremely fortunate to escape red cards for challenges on Alexis Mac Allister and Evan Ferguson.
Alisson threw himself in front of Solly March to make a brilliant save from Mitoma’s inventive cross and Konate did likewise to block from Ferguson.
But Brighton’s pressure finally got its reward when Mitoma struck his fourth goal in six games.