A process for producing L-proline by fermentation, comprising culturing a microorganism belonging to the genus selected from Corynebacterium, Arthrobacter, Brevibacterium, Microbacterium and Saccharomyces and capable of producing L-proline in a culture medium to accumulate L-proline in the cultured liquor and recovering L-proline therefrom, which process is characterized by the use of a culture medium containing at least one member selected from L-glutamic acid and D-, L- and DL-pyrrolidonecarboxylic acid in association with a carbon source, nitrogen source and inorganic compounds.