The invention discloses an industrial production method for preparing rhamnolipid by utilizing dual-phase carbon source fermentation, and belongs to the technical field of biosurfactant production. The method comprises steps: (1), breeding of a dual-phase carbon source dominant strain: screening out the dual-phase carbon source dominant strain after activation and habituated culture of pseudomonas aeruginosa CICC10204; (2), enlarge cultivation of strain: obtaining secondary seed liquor after activation and enlarge cultivation of the dominant strain; (3), fermentation and cultivation: carrying out dual-carbon source fermentation and cultivation onto the secondary seed liquor; and (4), separating and purifying: removing thallus and impurities of fermented liquor through a centrifugal method or membrane filtration, removing biomacromolecules and polysaccharide by a 6K Dalton ultrafiltration membrane; evaporating, concentrating and removing moisture for the collected filtrate and finally obtaining a rhamnolipid industrially-purified product. The industrial production method disclosed by the invention realizes industrial production of a rhamnolipid refined product, and concentration of finally purified rhamnolipid can be over 90% through concentration.