The 18-year-old swam the opening, backstroke leg in the hilltop pool in Barcelona and the U.S. triumph followed her titles this week in 100 and 200 backstroke, 200 freestyle and the 4x100 and 4x200 freestyle relays. She was fourth in Friday's 100 freestyle.
"It hurt really, really bad but now we're all done and we're all super excited," a still-dripping Franklin, who is due to sign on at University of California, Berkeley later this year, told reporters.
"I think I'm going to take my time off until I get to Berkeley so I think that gives me about two and a half weeks. We're all pretty excited about it."
Franklin's sixth gold is one more than the previous record of five she jointly held with compatriot Tracy Caulkins and Australia's Libby Trickett.
The only other woman to win six golds at a major swimming meet was East German Kristin Otto at the 1988 Olympics in Seoul.
After her three world titles at the 2011 championships in Shanghai, Franklin is also tied for the most golds overall with Trickett on nine.
The British quartet of Lauren Quigley, Sophie Allen, Jemma Lowe and Fran Halsall were the final Brits to compete in Barcelona, finishing sixth. They improved on their heat time to clock 3:58.67.
It was not an American double on the day, however, as France were handed a shock victory in the men's 4x100 metres medley relay when gold-medal favourites United States were disqualified.
The Americans touched first in three minutes 30.06 seconds before the stadium announcer drew gasps from the crowd and prompted wild celebrations among the French when he announced the U.S. had been disqualified "for an early exchange".
Official results showed 19-year-old Kevin Cordes, swimming the second, breaststroke leg, had jumped early by 0.04 seconds.
"A relay disqualification is not a particular individual's fault," U.S. team member Nathan Adrian told reporters.
"It's Team USA's fault and it falls on all of our shoulders," he added.
"It will really motivate him. In the next couple of years we're going to have the fastest breaststroker in the world."
France clocked a time of 3:31.51 to take gold, with Australia winning silver in 3:31.64 and Japan bronze in 3:32.26.
The Americans were denied what would have been a 12th triumph in the event in 15 championships since the inaugural edition in Belgrade in 1973.
The drama echoed an incident at the world championships in Melbourne in 2007.
Michael Phelps' bid to win eight golds was ruined when the U.S. team was disqualified from the heats of the medley relay.
Phelps had already won a record-equalling six titles and was expected to win his final two events on the last day of competition to make it eight from eight.
He safely qualified for the 400 individual medley but lost his chance to compete in an eighth final when Ian Crocker left the starting blocks one-hundredth of a second too early.