Kerber became the first female top seed in Roland-Garros professional-era history to fall in the opening round, following a 6-2, 6-2 defeat on the Philippe-Chatrier Court.
The world No.1, who also lost in the opening round last year, continued her turbulent form in 2017, while Makarova flourished with punchy groundstrokes to nullify Kerber’s weapons with her own power in a ruthless display.
The Russian, a former world No.8, has proved a problematic opponent for Kerber in the past.
“I'm so happy. It was quite a tough first round,” stated Makarova, who held a 4-7 head-to-head record against the German ahead of the contest. “We have played a lot of matches in the past with plenty of tight moments.
“It's always tough to play against Angelique, and especially when she's No.1. It's a totally different opponent, so I'm very happy that I could control emotions and my good tennis today.”
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World number 1 Angelique kerber was shown the door by Russian Ekaterina Makarova in the opening round at Roland-Garros on Sunday
Also a first-round casualty here last year when she wasn’t ranked No 1, Kerber could now lose her ranking crown to either Karolina Pliskova or Simona Halep.
Olympic champion Monica Puig advanced 6-3 3-6 6-2 over Italian 31st seed Roberta Vinci, while Swiss 30th seed Timea Bacsinszky thumped Spain’s Sara Sorribes Tormo 6-1 6-1.
A former semi-finalist in Paris, Bacsinszky next plays American Madison Brengle, a 1-6 6-3 13-11 victor over German Julia Georges.
And in a record 20th appearance at Roland Garros, Venus Williams has eased into the second round with a straight sets victory over Qiang Wang of China.
Svetlana Kuznetsova struggled to keep up with the score at the end of her first-round victory over Christina McHale.
Eighth-seeded Kuznetsova, who won the tournament in 2009, triumphed 7-5 6-4 but had to check the score with the umpire toward the end.