Tiger Roll was downed late on by stablemate and same-owned Delta Work – with the winning horse being booed as it passed the post and as it passed the stands afterwards.
Tiger Roll heroically took it up after travelling well but Delta Work - a previous winner at Cheltenham and also owned by Ryanair supremo Michael O'Leary - battled past him on the run-in to head him late home.
These two were a long way clear of Plan Of Attack, who finished third.
Many racegoers looked unhappy that the winner, from the same stable, was the party pooper for Tiger Roll's final race.
Trainer Gordon Elliott bought Tiger Roll in 2013 after he had been bred in Ireland, before selling him on to Ryanair founder Michael O'Leary.
He has become a legend of the sport over his nine-year career, winning the Grand National twice.
While he has only won 13 of the 44 races he has ridden, more than half of them have come at Cheltenham and Aintree. In total, he has earned winnings of more than £1.4m for his owners, with Elliott having bought him as an 'afterthought'.
But his career has now come to an end with the stallion set to be put out to pasture following the Cross Country Chase on Wednesday.
Former jockey Ruby Walsh told Paddy Power: "He's a wonderful horse. He's won two Grand Nationals. He's won a Triumph Hurdle, a National Hunt Chase and now he's trying to win his fourth Cross Country Chase.
"Whatever accolades have been thrown at the horse, you'd have to say his trainer deserves them every bit as much.".
