"Jennifer is not pregnant," the 44-year-old's rep told the Mail Online.
The rumour mill went into overdrive when Jennifer stepped out on 1 August for the New York premiere of her new film We're The Millers showing off a slightly rounded tummy. The Friends star was wearing a figure-hugging purple satin Burberry dress, which revealed a fuller figure.
She has since laughed off speculation that she is pregnant, putting her slightly curvier figure down to gaining "a couple of pounds".
"There's nothing to announce. That's just a couple of pounds," she said during an interview on Australia's Kyle and Jackie O Show.
The latest denial comes after Us Weekly published Jennifer on their front cover on Wednesday with the coverline, Jen's Big Secret, Pregnant!
The publication detailed how Jennifer has been hiding her baby bump during the promotion of her two new films We're The Millers and Life Of Crime.
According to Us Weekly, the Emmy winner has even had to rethink her wardrobe after fuelling the pregnancy rumours with her previous figure-hugging outfits.
"Her stylist had to redo her whole press tour wardrobe," a source tells the publication.
Another source tells the magazine of Jennifer’s reaction to being pregnant with Justin Theroux’s child: "She's thrilled!"
Jennifer has expressed her frustration at the world's interest in her private life. "It does feel a bit like they've exhausted the question, and also the speculations," she said.
"It's all been pretty much exhausted. I just think, 'Why not just wait until we have the fun of announcing something instead of playing the guessing game', whether it be family, weddings, this, that, the other thing," she added.