Armstrong reportedly fell around 20 kilometers from the finish and was seen pointing to his collarbone, CNN's Al Goodman reported from northern Spain.
Organizers said Armstrong was taken to hospital and had withdrawn from the race.
The 37-year-old Armstrong, who ended a three-year retirement to return to competitive cycling in January, walked to the ambulance unaided, television pictures from the scene showed.
A group of 15 to 20 riders fell, according to Bartosz Huzarski, a cyclist racing for the Italian team ISD. Huzarski, who saw the fall, said he did not know what had caused it. Only Armstrong appeared to indicate he was hurt, the Polish cyclist said.
An official with Armstrong's team, Astana, said he did not know the extent of the cyclist's injuries.
The fall took place a on a sunny day on a stretch of two-lane highway, Goodman said, as riders tackled the 104-mile (168-km) opening stage.
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