Player had hoped to qualify for his 48th consecutive British Open, but will be unable to compete because of severely pulled rib muscles.
"I am just devastated and cannot put into words my deep disappointment," Player said. "I obviously aggravated what I thought was only a strain that would go away through exercise and practice. Muirfield was the site of my first major championship win and not to compete there this year is truly heartbreaking."
The 66-year-old Player is no longer exempt because the British has an age limit of 65 for past champions. The South African won in 1959 at Muirfield, 1968 at Carnoustie, and 1974 at Lytham.
Player, who has won 163 tournaments worldwide, will remain in Florida and seek physiotherapy for his injury. He also may be forced to withdraw from the British Senior Open later this summer.