Ryder Cup stars Sergio Garcia, Ian Poulter and Lee Westwood will all be ineligible to play in this year's event after resigning from the DP World Tour.
The trio, along with Richard Bland, had been sanctioned for breaking tour rules after joining breakaway tour LIV Golf.
The players say they have "no option" given punishments being imposed after they joined the Saudi Arabia-funded tour.
Bland has also quit the European circuit.
All four will now be unable to play for Europe against the United States in Rome.
In a statement, the DP World Tour confirmed the resignations saying they had been "sanctioned for serious breaches of the tour's conflicting tournament regulations".
The players were hit with £100,000 fines for playing in LIV's opening tournament at the Centurion Club in Hertfordshire last June. They lost an appeal held by the arbitration service Sport Resolutions and had until 3 May to pay their fines.
The DP World Tour thanked the players for the contribution they have made, adding: "In particular Sergio, Ian and Lee for the significant part they have played in Europe's success in the Ryder Cup over many years".
Garcia is Europe's all-time record points scorer while Westwood played in a record 11 matches for Europe.
World number one Jon Rahm has made no secret of his desire to partner Garcia in this year's match after both Spaniards combined successfully when Europe lost at Whistling Straits in 2021.
"The resignations, however, along with the sanctions imposed upon them, are a consequence of their own choices," the DP World Tour statement added.
"As we have consistently maintained throughout the past year, the tour has a responsibility to its entire membership to administer the member regulations which each player signs up to.
"These regulations are in place to protect the collective interests of all DP World Tour members."
Under the tour's rules the players cannot serve as future captains or vice captains. Westwood, Poulter and Garcia were seen as certainties to skipper Europe in future Ryder Cups until they defected to LIV.
Westwood called it "a sad day" according to the Daily Telegraph. The Englishman has been a member for more than 30 years after joining as a 19-year-old.
"I could not see any other option with all the punishments hanging over me," Westwood said.
"I don't agree with the direction [chief executive] Keith [Pelley] and his board have taken and I want to move on." Westwood, who signed a multi-million dollar deal with LIV, confirmed he has paid his £100,000 fine.