Zambrano, who pitched one inning for the Mesa Rookie League team on Monday, threw 15 pitches, 10 for strikes, in front of 12,949 at Principle Park in Des Moines, Iowa.
He entered with the game tied at 4 and got Oklahoma City's Guilder Rodriguez to ground out to second and Craig Gentry singled to right off a 1-1 pitch. Zambrano then got help from center fielder Sam Fuld, who made a leaping catch at the wall of Hernan Iribarren's fly ball for the second out. Mitch Moreland singled, and Zambrano got Jarrod Saltalamacchia to ground out to second.
Of the five batters Zambrano faced, he only threw a first-pitch strike to Saltalamacchia.
After he was finished with the inning, Zambrano, who was the Cubs' Opening Day starter, went to the bullpen to get more work in.
He has not pitched for the Cubs since June 25, when he started against the White Sox and then threw a tantrum in the dugout after the first inning, upset at the way his teammates were playing behind him.
Zambrano was suspended for three days and then placed on the restricted list and underwent treatment for anger issues. He will stay with the Iowa Cubs to pitch in relief, traveling this weekend to Round Rock, and was not expected to join the big league team until July 30 in Colorado at the earliest.
Carrie Muskat is a reporter for MLB.com. This story was not subject to the approval of Major League Baseball or its clubs.