The 28-year-old striker has been in sensational form of late, and the Rio de Janeiro-based club to remain in the race for the league title with two more goals
The 28-year-old scored twice at the Engenhao to take his overall tally in the competition to 44 goals, breaking the previous record owned by Magno Alves, who netted 43 times in the league for the club between 1998 and 2003.
Fred, who signed for Fluminense in 2009 after four years at Lyon, has been a big hit for the Rio de Janeiro-based club, scoring 87 goals in 129 matches.
He had already broken the club's record for the most goals in a single edition of the Brasileiro after netting 22 times last season.
"This was a blessed night," Fred told Lancenet. "I knew that with a goal I would be part of Fluminense's history, but you do not think about that on the field.
"We had a had time in the first half, but then we made the most of our chances and things got easier for the strikers. I hope this is the first of many records that I will break."
Fluminense are currently third in the Brasileiro standings on 22 points, three points adrift of leaders Atletico Mineiro.