In an unspecified future (the video release states it is the year 40,000), Barbarella (Jane Fonda) is assigned by the President of Earth (Claude Dauphin) to retrieve Doctor Durand Durand (Milo O'Shea) from the planet Tau Ceti. Durand Durand is the inventor of the Positronic Ray, a weapon. Earth is now a peaceful planet, and weapons are unheard of. Because Tau Ceti is an unknown region of space there is the potential for the weapon to fall into the wrong hands. Donning the first of many outfits, Barbarella sets out to find the missing scientist. She crashes on Tau Ceti, on an icy plain.
Barbarella is soon knocked unconscious by two mysterious girls who hit her on the forehead with a snowball and then capture her. Barbarella is taken to the wreckage of a spaceship called the Alpha 1. Inside the wreckage, she is tied up and several children emerge from within the ship. They set out several dolls which have razor sharp teeth. As the dolls begin to bite her, Barbarella faints but is rescued by Mark Hand (Ugo Tognazzi), the Catchman, who patrols the ice looking for errant children.
While taking her back to her ship, Barbarella offers to reward Mark and he asks to make love to her. She reveals that people on Earth no longer have penetrative intercourse, but make love by consuming exaltation transference pills, and pressing their palms together when their "psychocardiograms are in perfect harmony". Hand prefers the bed, and Barbarella agrees. Hand's vessel makes long loops around Barbarella's crashed vessel while the two make love, and when it finally comes to a stop, Barbarella is blissfully humming. Hand repairs her ship, and Barbarella departs, promising to return, and agreeing that doing things the old-fashioned way is sometimes best.
Barbarella's ship burrows through the planet, coming out next to a vast labyrinth. Upon emerging from her ship, she is knocked unconscious by a rockslide. She is found by a blind angel named Pygar (John Phillip Law). He states he is the last of the ornithanthropes, and that he has lost the ability to fly. Barbarella discovers the labyrinth is a prison for people cast out of Sogo, the City of Night. Pygar introduces her to Professor Ping (Marcel Marceau), who offers to repair her ship. Ping also notes that Pygar is capable of flight, but merely lacks the will.
After Pygar saves her from the Black Guards, Barbarella shows her thanks by making love to him. As she hums to herself in his nest afterwards, Pygar soars overhead, having regained his will to fly. Pygar flies Barbarella to Sogo, using some of the weaponry she brought with her to destroy the city's guards. Sogo is a decadent city ruled over by the Great Tyrant and powered by a liquid essence of evil called the Mathmos.
Barbarella is briefly separated from Pygar, and meets a one-eyed wench who saves her from being assaulted by two of Sogo's residents. Barbarella soon reunites with Pygar and the two are taken by the Concierge (O'Shea) to meet the Great Tyrant. Pygar is left to become the Great Tyrant's plaything, while Barbarella is placed in a cage, to be pecked to death by birds (something Barbarella says is "much too poetic a way to die").
Barbarella is rescued by Dildano (David Hemmings), leader of the resistance to the Great Tyrant. Barbarella eagerly offers to reward him. As she begins to remove her torn suit, Dildano says he has the pill, and wants to experience love the Earth way. Dildano offers to help Barbarella find Durand Durand in exchange for her help in deposing the Great Tyrant. Barbarella is given an invisible key to the Tyrant's Chamber of Dreams, the only place she is vulnerable.
Barbarella is captured by the Concierge and she is placed inside the Excessive Machine. As the Concierge begins to play, Barbarella experiences increasing pleasure, and her clothing is expelled from the machine. The Concierge tells her when they reach the crescendo, she will die of pleasure. He then begins to play faster and more furiously, while Barbarella writhes in ecstasy inside the machine. Eventually, the machine overloads and burns out, unable to keep up with her. Barbarella then discovers the Concierge is none other than Durand Durand, aged thirty years due to the Mathmos.
Durand Durand traps Barbarella in the Tyrant's Chamber of Dreams, taking both keys and locking them inside. As he prepares to crown himself lord of Sogo, Dildano launches his revolution. Durand Durand uses his Positronic Ray to decimate the rebels. The Great Tyrant then releases the Mathmos, which consumes all of Sogo and Durand Durand with it. Barbarella and the Great Tyrant are protected from the Mathmos by Barbarella's innate goodness. They emerge from the Mathmos to find Pygar. Pygar then flies Barbarella and the Tyrant away from the Mathmos. When asked by Barbarella why he saved the Tyrant after everything she had done to him, Pygar responds, "An angel has no memory."
- Jane Fonda as Barbarella
- John Phillip Law as Pygar, the angel
- Anita Pallenberg (dubbed by Joan Greenwood) as The Great Tyrant, Black Queen of Sogo
- Milo O'Shea as Durand Durand / Concierge
- Marcel Marceau as Professor Ping
- David Hemmings as Dildano
- Claude Dauphin as President Dianthus of Earth
- Ugo Tognazzi as Mark Hand
- Fabienne Fabre as the tree woman