Kupka worked as an illustrator of books and posters and, during his early years in Paris, became known for his satirical drawings for newspapers and magazines. In 1906, he settled in Puteaux, a suburb of Paris, and that same year exhibited for the first time at the Salon d'Automne. Kupka was deeply impressed by the first Futurist Manifesto, published in 1909 in Le Figaro. Kupka’s 1909 painting Piano Keyboard/Lake marked a break in his representational style. His work became increasingly abstract around 1910 – 11, reflecting his theories of motion, color, and the relationship between music and painting (orphism). In 1911, he attended meetings of the Puteaux Group (Section d'Or). In 1912, he exhibited at the Salon des Indépendants in the Cubist room, although he did not wish to be identified with any movement. Creation in the Plastic Arts, a book Kupka completed in 1913, was published in Prague in 1923.

In 1931, he was a founding member of Abstraction - Création. In 1936, his work was included in the exhibition Cubism and Abstract Art at the Museum of Modern Art in New York City, and in an important show with another excellent Czech painter Alphonse Mucha at the Jeu de Paume in Paris. A retrospective of his work took place at the Galerie Mánes in Prague in 1946. The same year, Kupka participated in the Salon des Réalités Nouvelles, where he continued to exhibit regularly until his death. During the early 1950s, he gained general recognition and had several solo shows in New York.

Between 1919 and 1938 Kupka was financially supported by his good friend, art collector and industrialist Jindřich Waldes who accumulated a substantial collection of his art. Kupka died in 1957 in Puteaux, France.

Kupka had a strong interest in color theory; around 1910 he began developing his own color wheels, adapting a format previously explored by Sir Isaac Newton and Hermann von Helmholtz. This work in turn led Kupka to execute a series of paintings he called "Discs of Newton" (1911 - 12). Kupka was interested in freeing colors from descriptive associations. His work in this area is thought to have influenced other artists like Robert Delaunay. Important works include Planes by Colors, The Colored One, Reminiscence of a Cathedral, Blue Space. Works in the Peggy Guggenheim Collection, Venice, Italy include Study for Woman Picking Flowers (Femme cueillant des fleurs), cca 1910, Study for Amorpha, Warm Chromatics, Chromatique chaude and for Fugue in Two Colors (Fugue a deux couleurs), cca 1910 - 1911, Vertical Planes (Plans verticaux), 1911 - 1912, Study for Organization of Graphic Motifs I (Localisations de mobiles graphiques I), cca 1911 - 12, Around a point (Autour d'un point), cca 1920 - 1925.