The 22-year-old Real Madrid defender has injured his left thigh and will not be available to coach Vicente del Bosque for the Group C clash.
Carvajal played the full 90 minutes of Spain's 1-0 friendly defeat to France on Thursday.
A team statement released on Monday read: "Dani Carvajal is sidelined from the national team due to a injury in his left thigh.
"Injury was detected with an MRI scan carried out on Monday 8th September in Valencia…all the tests carried out by the medical staff will be sent to Real Madrid."
Carvajal's injury could also rule him out of Saturday's Madrid derby against Atletico.
The Liga clash – a repeat of last season's UEFA Champions League final – will take place at Santiago Bernabeu.
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