De Jong, 21, has been strongly linked with a move to the Spanish champions over the last few months, and it was claimed last weekend that the Catalan outfit were prepared to pay £44m for his services.
However, Overmars has said that neither De Jong nor Matthijs de Ligt - who has been strongly linked with a move to England - will be allowed to leave Ajax ahead of the new season.
"De Jong to Barcelona? He is not for sale. And I have communicated this to all parties involved. It's the same for Matthijs. Clubs that are interested are free to come back next year," Overmars told De Telegraaf.
"Of course, Davinson Sanchez's move last year showed that it is not always easy to stand firm. But we have now reached a point where we simply say that we are not selling these players."
De Jong scored once and registered nine assists in 26 appearances for Ajax last term.