Natalia Ishchenko and Svetlana Romashina - in costumes decorated with puppets, their eyes framed by doll-like eyelashes - took gold after scoring 98.9 points out of a possible 100 in the free routine executed for the final.
That marked an improvement on Monday's already leading score for the same syncopated sequence and took their total points to 197.1 - more than four points clear of their nearest rivals Spain and China, who took silver and bronze respectively.
Russians, bringing a tradition of ballet to the pool, have taken home the Olympic gold for duets since Sydney in 2000, earning a reputation for seamless execution in a sport that demands precision, endurance and nerves of steel underneath the fixed smiles, gelatined hair and often garish costumes.
"We work very hard, we train for up to 10 hours a day, in the pool, in the gym," Ishchenko said.
"I don't want to reveal our secrets, but to be upside down for this length of time is no joke."
Russia's swimmers, favourites even before the Games began, said they battled in the days leading up to the competition to focus on their routines and ignore headlines that "hung gold medals around our necks days before".
Coach Tatiana Danchenko said the pair would not be resting on their laurels as duets from China, Spain and even Britain continue to climb the rankings. Britain was in its first final for 20 years, jumping from 14th in Beijing to 9th in London.
"(The worst) would be to stay in one place," Danchenko said. "Our competition are not asleep, they are preparing, working, training and they'd like to unseat us."
A tango-inspired routine, using intricate tilted legwork and set to one of the dance's most famous tunes - 'La Cumparsita', led the Spanish duet to silver, retaining the same medal Spain had won in Beijing four years ago and holding off China.
Veteran swimmer Andrea Fuentes, already a silver medallist in 2008 for Spain, was dancing with relative newcomer Ona Carbonell, after her previous partner retired earlier this year.
China's swimmers, with a dragon routine, came just 0.03 points behind to take bronze and win the country's first Olympic medal for the duets, marking its emergence as a contender for the top.
"We finally built our status within the three strongest countries in synchronised swimming with this medal," China's Japanese-born coach, Masayo Imura, said. "This time we were just beaten by the degree of difficulty in this event."
Russia, which won all seven synchronised swimming golds at the 2011 world championships, are also favourites to retain the title in the team segment in London, which begins Thursday.