Fluminense celebrated the title it clinched a week ago, receiving the league trophy in front of home fans in Rio de Janeiro despite a 2-0 loss to Cruzeiro at Engenhao stadium.
Palmeiras, the team with the most national titles and the defending Brazilian Cup champion, had its demotion to the second division sealed after Portuguesa drew 2-2 with second-placed Gremio in Sao Paulo later in the day.
After the draw with Flamengo, it needed Portuguesa to lose the match at Caninde Stadium.
Victory escaped Palmeiras in Volta Redonda after former player Vagner Love scored in the 89th minute with a shot that deflected off a defender before going into the net.
The eight-time Brazilian champion was demoted for the first time since 2002.
With two rounds left, Palmeiras is seven points behind Portuguesa, the first team out of the relegation zone. It is 18th in the 20-team standings with 34 points from 36 matches. The bottom four clubs are relegated. Portuguesa, Bahia and Sport remain in danger, while Figueirense and last-placed Atletico Goianiense have already been demoted.
Before the Fluminense loss to Cruzeiro, world No.1 tennis player Novak Djokovic and former three-time French Open champion Gustavo Kuerten both scored in a charity football match at the stadium.
The exhibition was organised by Djokovic's Serbian countryman Dejan Petkovic, who starred for several Brazilian clubs in the past decade. Former Brazil stars Zico and Bebeto were among the participants.