The Spanish Footballers' Association (AFE) notified La Liga on Monday that it does not approve of plans to stage Girona's home game against Barcelona in January in the United States.
La Liga planned to host a regular-season game overseas annually as part of a 15-year marketing agreement with Relevent Sports and decided to move the Catalan derby, set for Jan. 26, to suburban Miami.
The players' union has had reservations about the plan from the start, and after receiving a detailed report from La Liga, it said in a statement it believes the decision to move the game violates several clauses in the collective bargaining agreement, specifically with regards to the players' well-being.
"The AFE reminds [La Liga] that for the time being, there are no mandatory authorisations that allow for the game to be played in the United States," it said. "Therefore, considering the lack of information mentioned, AFE has transmitted to La Liga its lack of support for that decision."
The union's decision comes three days after the Royal Spanish Football Federation (RFEF) indicated that it also does not intend to support La Liga's plans.
La Liga president Javier Tebas told ESPN FC on Friday that there was still a "90 percent chance" the game would be moved to Hard Rock Stadium.
Tebas believes not staging a league game in the United States would be a missed opportunity for Spain's top flight to continue its growth.
"In Europe, no league has initiatives like this one," he said while speaking at a football congress in Madrid. "We have to grow where we have to grow. We have to be different. This is an industry and it affects everyone.
"Since I've arrived, La Liga has grown 22 percent while the Premier [League] has dropped 12 percent, so we are not doing so bad."
On not having the support of the Spanish FA and the Spanish Footballers' Association, Tebas added: "Perhaps now the chances of playing the game are not 90 percent but 89 percent. We have a clear aim to play the game there this season and we will continue trying."
In addition to the RFEF and AFE, the league would also need the support of Spain's sports council, UEFA, CONCACAF and U.S. Soccer in order to move the game from Girona's Estadi Montilivi.