Leading off the fifth inning against right-hander Randall Delgado, Polanco hit a ground ball to third base and pulled up as he jogged down first-base line, but his discomfort was earlier in the at-bat when he took a weak swing and whiffed at a 94.9 mph fastball.
"I tried to go through today with that, but when I went to hit, I tried to stay back, I couldn't hold," he said. "It was painful."
But Polanco actually tweaked his hamstring two innings earlier while making an excellent running catch in right field. Polanco covered a lot of ground and sprinted to track down a fly ball hit by Jake Lamb for the second out of the inning.
"That's what was weird. I was feeling good," Polanco said. "I ran too much. I ran a long way. … That's when I felt it."
John Jaso replaced Polanco in right field in the bottom of the fifth.
Polanco is considered day-to-day as the Pirates continue to treat and evaluate him, the club announced. It is unclear how long the injury will linger.
"I don't know. I have to see when I wake up tomorrow how I feel," Polanco said. "I'll talk to the trainers. Hopefully it'll feel better because right now I'm feeling a lot better in the box."
And that is perhaps only the second-cruelest thing about the timing of Polanco's early exit. The Pirates just returned to full strength on Friday, when Adam Frazier and David Freese came off the disabled list following time down to heal strained hamstrings.
Losing Polanco for any length of time would be another blow for a team already without outfielder Starling Marte (80-game suspension for performance-enhancing drugs), third baseman Jung Ho Kang (awaiting a work visa in South Korea) and starter Jameson Taillon (surgery for suspected testicular cancer).
After a rough start to the season, Polanco has 11 hits in his last 33 at-bats over the past 10 games.
"That's hard for me now that I'm feeling better hitting and everything, I'm back in right field, it's hard for me that that happened," Polanco said. "It is what it is. That's baseball. Just trying to get better as soon as I can."
Adam Berry has covered the Pirates for MLB.com since 2015. Follow him on Twitter and Facebook, read his blog and listen to his podcast. This story was not subject to the approval of Major League Baseball or its clubs.