The statement came as the team's new coach, Montse Tome, was poised to announce the lineup for two upcoming international matches, in the new UEFA Nations League, which federation sources said would now be delayed.
"As we have informed the RFEF today, the changes put in place are not enough for the players to feel they are in a safe place where women are respected, where women's soccer is supported and where we can give our maximum performance," they said in a statement released by the Futpro union.
The statement was signed by 39 players, among them 21 of the 23 players on the women's national squad.
Spanish soccer has been in turmoil since RFEF boss Luis Rubiales, who has since resigned, forcibly kissed midfielder Jenni Hermoso on the lips after Spain defeated England in the Women's World Cup final on Aug. 20.
On Aug. 25, 81 Spain players, including the 23 world champions, joined a mass strike, saying they would not play for the national team without significant changes at the head of the federation.
Since then, the RFEF sacked the team's controversial coach Jorge Vilda and named Tome as his replacement, the first woman to hold the post, as well as pledging further internal reforms. And Rubiales resigned earlier this week.
In the statement, the players expressed their "enormous dissatisfaction" with the events after the World Cup and at an RFEF meeting on Aug. 25 at which a belligerent Rubiales said he would not resign, to widespread applause.
"The events, which unfortunately everyone saw, were not a one-off, and they go beyond the realm of sport. We must have zero tolerance for such incidents for the sake of our colleague [Hermoso], for our sakes, and those of all women," they said.
The players said they had met several times with the RFEF to set out "the essential changes to move forward and build a system that does not tolerate or participate in such degrading acts."
Demanding "fundamental changes in the RFEF's leadership" they called for the "resignation of the RFEF president" in a statement widely understood to refer to interim leader Pedro Rocha, who took over when FIFA suspended Rubiales on Aug. 26.
Despite weeks of resistance, Rubiales finally resigned late on Sunday and was in court in Madrid earlier on Friday on charges of sexual assault and coercion over the kiss and his subsequent behaviour towards Hermoso.
Tome had been due to announce the lineup for Spain's Nations League matches against Sweden and Switzerland on Sept. 22 and 26.
Athenea del Castillo and retiring Claudia Zornoza are the two World Cup champions who did not sign the players' statement.
© Agence France-Presse