"Andrew has been experiencing some discomfort in his left ankle and did not bowl during practice today," said the England and Wales Cricket Board in a statement on Saturday.
"Andrew suffered a reaction following Thursday's match (in Manchester)
The statement added a decision on his fitness for Sunday's game would be made after further assessment.
Flintoff has had surgery on the ankle three times in his career, most recently in June.
England are 3-1 up in the seven-match series.