Czech Kvitova will face Svetlana Kuznetsova in the final after she surprisingly knocked out defending champion Maria Sharapova.
Williams had not lost since October but her mood appeared negative and she was slow around the court against fourth seed Kvitova who broke the American’s first service game and never looked back.
“I’m just exhausted today. This has been a rough week for me,” Williams told a news conference.
“It’s good to be really exhausted now going into another tournament and then getting ready for Roland Garros.
“So this is all great preparation that I actually really, really needed, something to push me to the wall and to the limit.”
Kvitova broke serve three times in the first set and there was little resistance in the second set from a despondent Williams.
It was a setback for Williams who won in Madrid in 2012 and 2013, and this was her first claycourt tournament of the season as she prepares for the French Open looking to win her 20th grand slam title.
Sharapova crashed out after failing to match the solid hitting of unseeded Kuznetsova who won 6-2 6-4.
She looked tense from the start against her fellow Russian, whose consistency from the back of the court helped her break twice to win the first set.
Sharapova came to the tournament in poor form and struggled to get through her previous two matches in the Spanish capital with Caroline Garcia and Caroline Wozniacki taking her to three sets.
She failed to convert a break point at 2-1 in the second set and lost her serve in the following game. The Russian battled hard to win her next service games but went down 6-4.
“I felt my shots were a little too flat and right in her striking zone,” Sharapova told reporters.
“She was able to push me back and when we got into a rally she was the one that found the right ball to be the more aggressive player."
Kuznetsova is a former grand slam winner at the U.S. Open and Roland Garros but has struggled to reach those heights in recent years and is now ranked 29th in the world.
In the men’s tournament Rafa Nadal faces 10th seed Grigor Dimitrov in the quarter-finals as he continues his bid for a third consecutive Madrid title.
Second seed Andy Murray meets fifth-seeded Milos Raonic.
(This version of the story changes dateline)
(Reporting by Tim Hanlon in Barcelona; Editing by Ed Osmond and Toby Davis)
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