Messi's left knee collided with the right hand of Benfica's Artur as he ran onto Gerard Pique's long pass and tried to round the goalkeeper in the 85th minute.
Messi took a left-footed shot from an angle, then fell to the field in pain. He rolled onto his back and held the knee, then was loaded onto a cart.
''Messi has a bruise on the outside of his left knee,'' Barcelona said on its Twitter feed. ''He'll undergo further tests to determine the exact extent of the injury.''
A three-time FIFA Player of the Year, Messi has 84 goals for Barcelona and Argentina this year, one shy of Gerd Mueller's 40-year-old record for most goals in a calendar year for club and country.