Hosszu, who has already won two golds and destroyed the 400m individual medley world record, pulled out of the 200m butterfly heats to focus on the 200m IM final scheduled for 11.30pm on Tuesday.
Spain's Mireia Belmonte Garcia, a 400 IM bronze medallist on Saturday, was fastest with Natalie Adams of the United States second.
Hungarian swimming federation official Gergely Csurka said Hosszu had not left the venue until 1am on Tuesday after winning Monday's 100m backstroke gold.
The swimming finals have been scheduled for prime time U.S. audiences, and to maximise advertising revenues, with some races ending after midnight local time with media and doping controls to follow.
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Hosszu's withdrawal meant she cannot now equal Australian swimmer Shane Gould -- who won her three golds, a silver and a bronze in Munich in 1972.
She could, however, still match former East German swimmer Kristin Otto's 1988 achievement of four individual golds at a single Games.
Nathan Adrian, the men's 100 metres freestyle champion and U.S. team co-captain, also cited the late nights after just scraping through to the semi-finals as the 16th and last qualifier.
Adrian was a mere 0.03sec faster than Japan's Katsumi Nakamura, who failed to make the cut.
"Feeling a little rusty this morning, trying to get back into the meet after a late night," said Adrian, who had a late night on Sunday after winning the 4x100m freestyle relay in a lineup that included 19=times gold medal winner Michael Phelps.
Australian teenager Kyle Chalmers was fastest in the 100m heats -- his time of 47.90 was the third fastest this year -- with American Caeleb Dressel second in 47.91 and Britain's Scott Duncan third in 48.01.
While Adrian was on the right side of the cut-off, 2012 Olympic champion Daniel Gyurta of Hungary failed to go through to the men's 200m breaststroke semi-finals after ending up 17th.
That failure meant none of the 2012 medallists in the race will be in the semi-finals, unless another swimmer scratches and opens a place for Gyurta. Russian Anton Chupkov was fastest in the heats.
World champions Britain led the way into the men's 4x200 freestyle final, followed by defending Olympic champions the United States, and Russia.
France, the silver medallists in London four years ago, failed to qualify.
(Reporting by Alan Baldwin, editing by Neil Robinson)
