Speaking to Empire magazine in an interview published on Wednesday, the Oscar-winning filmmaker said he would be focusing on his theatre projects for the foreseeable future.
The success of "Skyfall", the top-grossing Bond film having earned more than $1.1 billion at the global box office, had led to media speculation that Mendes would return for Bond 24.
"It has been a very difficult decision not to accept Michael and Barbara's very generous offer to direct the next Bond movie," said Mendes, referring to Bond producers Barbara Broccoli and Michael G. Wilson.
"Directing "Skyfall" was one of the best experiences of my professional life, but I have theatre and other commitments, including productions of "Charlie And The Chocolate Factory" and "King Lear", that need my complete focus over the next year and beyond."
Mendes has a long background in theatre direction and burst on to the movie scene by winning an Academy Award for his debut feature "American Beauty" starring Kevin Spacey.
His collaboration with Daniel Craig as James Bond and handling of key new cast including Ralph Fiennes and Ben Whishaw in "Skyfall" has been widely praised by critics and helped the film become a major commercial success.
The 47-year-old director left the door open for a return to the franchise in the future, however.
"I feel very honoured to have been part of the Bond family, and very much hope I have a chance to work with them again sometime in the future," he said.
Wilson and Broccoli said they understood Mendes' decision to turn down Bond 24 but added they "hope to have the opportunity to collaborate with him again."