Alex Santos has joined Akira Kaji on Japan's injury list ahead of the World Cup opener with Australia on 12 June.
Santos limped off the field against Malta on Sunday
The Brazilian-born winger picked up the injury during the 1-0 win over Malta on Sunday, in Japan's final warm-up match.
He said: "I've got the foot iced up but I should be okay for the Australia game. I'll see how it is on Tuesday."
Kaji, 26, remained a fitness doubt with a bruised ankle, and coach Zico said: "It is difficult to say when he will back in action. It may take some time."
Zico added: "He is still in a lot of pain."
Right-back Kaji said: "I'm getting better but I am not sure about the opening match."
First-choice strikers, Naohiro Takahara and Atsushi Yanagisawa were left out of the Malta game because they were "not 100 percent" fit due to muscle pains.
"Yanagisawa has recently come back from a broken metatarsal and his muscles are a bit tender and Takahara tweaked his knee so we kept him out to be on the safe side," Zico said.