The 31-year-old defended her title on Valegro after holding off strong competition from Germany’s Isabell Werth (89.071%) and Kristina Broring-Sprehe (87.142%).
Britain’s Carl Hester, a team-mate of Dujardin in the dressage team event when they won silver, came seventh (82.482%).
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It is Dujardin's third gold medal, with her team dressage title from London making up the trio, and she becomes the first British woman to retain an individual Olympic title.
After the display, she told the BBC: "I’m actually a little bit speechless for once in my life. It means the absolute world to me.
"At London, I had no expectations or pressure to go out there. Today I felt a huge amount of pressure and expectation.
"For me it could be one of the last rides on Valegro. For me, to finish it this way, it’s a really emotional time. I’m so happy."
Ian Stark, the Olympic silver medallist and BBC equestrian pundit, said of Dujardin's display: "That was incredible. We can just abandon the rest of the day - I'm happy now!"
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