The Spanish tennis star said his lawyers lodged the defamation suit in Paris against Roselyne Bachelot because of her "offensive remarks" on French television last month.
Bachelot, France's former minister for health and sport, said on TV show Le Grand 8 that the Spaniard's seven-month injury hiatus in 2012 was "probably due to a positive doping test."
"Through this case, I intend not only to defend my integrity and my image as an athlete but also the values I have defended all my career," Nadal said in a statement.
He said that if he wins the case, any compensation awarded by the judge will be "paid back in full to an NGO or foundation in France."
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Bachelot's remarks upset Nadal and outraged Spaniards, including many fellow athletes who quickly came out to defend the tennis star. The Spanish Olympic Committee and members of the Spanish government also loudly criticized the former French minister.
Nadal's coach and uncle, Toni Nadal, called Bachelot "an imbecile," according to Spanish media.
The president of Spain's Olympic committee said he would like to see Nadal as the country's flag bearer at the Rio de Janeiro Games, in part to make a statement after the recent accusations made by Bachelot.
Alejandro Blanco said Nadal "is a reference" and deserves the honor "after how he has been treated" by the former French minister. Nadal had been selected as Spain's flag bearer for the 2012 London Olympics but missed the games because of an injury.
Bachelot's allegations came in the wake of Maria Sharapova's failed doping test.