The 22-year-old Ukrainian led throughout the race to become only the third swimmer, and first woman, to achieve an Olympic 'double double', having also won the 200 and 400 individual medley events at the 2000 Sydney Games.
American Amanda Beard, fourth in Monday's 100 metres breaststroke, moved up to second in her speciality stroke -- third of the four medley legs -- but could not catch Klochkova, who won in two minutes 11.14 seconds.
Beard, double breaststroke silver medallist at the age of 14 at the 1996 Atlanta Games, took another silver in 2:11.70, while Kirsty Coventry of Zimbabwe scooped bronze in 2:12.72 to add to the silver she won in Monday's 100 backstroke.
Hungary's Tamas Darnyi won both men's medleys in 1988 and 1992, while Russia's Alexander Popov won the 50 and 100 metres freestyle both in 1992 and 1996.
World championship 400 medley silver medallist in 1998 at the age of 15, Klochkova suffered a rare defeat at the hands of American Maggie Bowen in the 200 medley at the 2001 world championships but won the 400 medley and 400 freestyle.
Two years later she completed the 200-400 medley double at the Barcelona world championships. She has now won four Olympic and four world championship titles.
She has also won 10 individual European golds -- a record she shares with Popov -- including medley doubles at the last four editions of the championships.
