The 32-year-old reportedly made the inflammatory comment at a pre-Super Bowl concert before singing the track, which was inspired by his split from Spears.
But "Friends With Benefits" star has insisted he did not disrespect his former girlfriend, reported Daily Star.
Introducing the song, he had said, "Sometimes in life, you think you found the one. But then one day you find out that she is just some bitch."
After coming under fire for the comment on micro blogging site Twitter, Timberlake wrote, "Ok. I see you. Wouldn't disrespect ANYONE personally. Ever. #Relax #ItsBritneyBitch I do love that saying though *with accent* #Respect (sic)"
The actor, who is now married to Jessica Biel, called off his three-year relationship with Spears in 2002 after she allegedly cheated on him with her then choreographer, Wade Robson.