Third-placed Hoffenheim, whose 29-year-old coach Julian Nagelsmann is the youngest in the league, dominated the first half and deservedly won with a goal from Andrej Kramaric.
Bayern, who have 65 points from 27 games, are still 13 clear of second-placed RB Leipzig, who visit Mainz 05 on Wednesday.
Hoffenheim, who had never previously beaten Bayern in the league, are third with 51 and still on course for a Champions League place while Borussia Dortmund remain their closest rivals, remaining fourth after easing past Hamburg SV 3-0.
Bayern were outclassed in the first half and fell behind to a Kramaic half-volley which goalkeeper Sven Ulreich could only deflect into his net.
The Bavarians nearly levelled with their first real chance when Roberto Lewandowski's strike hit the crossbar.
Hoffenheim goalkeeper Oliver Baumann had more to do in the second half and Bayern nearly snatched an equaliser deep into stoppage time when Lewandowski's close-range shot was deflected wide of the goal.
Gonzalo Castro, Shinji Kagawa and Pierre-Emerick Aubameyang shared the goals in Dortmund's win, which left Hamburg -- the only ever-present club in the Bundesliga's 54-year history -- just one point clear of Augsburg in the relegation playoff spot.
(Writing by Brian Homewood in Berne,; Editing by Neville Dalton)
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