Four late free throw conversions by Malik Smith in the final 30 seconds and one by Elliott Eliason with 33 seconds, enabled the Gophers (13-3, 2-1 Big Ten) to hang on after taking its first lead of the second half with 1:43 remaining.
Tim Frazier scored 20 points for Penn State (9-7, 0-3) but was unable to finish the game after injuring his right leg with 2:08 left and Penn State leading 59-57.
Mathieu forced back-to-back Penn State turnovers, including a strip of John Johnson and his ensuing basket gave the Gophers a 63-60 lead and forced Penn State to foul.
The Nittany Lions' Allen Roberts missed a game-tying 3-point shot with 1 second to go.
Minnesota, which has won 30 out of 40 games in its series with Penn State, also got 11 points and 12 rebounds from Eliason.
Gophers Can't Hold Off Penn State, Fall 52-50
Tony Carr drew a foul on a final drive to the basket and made two free throws with five seconds left to lift Penn State to a 52-50 win over No. 24 Minnesota on Saturday.
Preview: Gophers at Penn State
Minnesota closes out a grueling stretch of road games to begin conference season in Happy Valley Saturday.
Minnesota Drops Contest at Michigan State, 65-47
Michigan State build a huge lead it used to cruise to a 65-47 victory over No. 24 Minnesota on Wednesday night.