Simulated moving bed (SMB) process is a continuous chromatographic technology used to separate a large amount of petrochemicals, fine chemicals, pharmaceuticals, and so on, drawing a great attraction of related industries. With the recent development of biotechnology, the SMB process has been adopted for the separation of various useful bio-products. Attempts to understand the separation mechanism of the SMB process in many aspects are reported in many publications. These researches have dealt with the improvement of SMB for easier operation and solving problem in process. The feed mixture fed into the SMB process may be of different concentration batch by batch rather than in uniform concentration. Retention behaviors of feed (psicose (A) and fructose (B) mixture) existing in the SMB unit in dynamic steady-state and feed (psicose (C) and fructose (D) mixture) newly injected into the SMB were analyzed. It was observed that the existing components, (A) and (B), were eluted relatively faster as the injection time of new feed was earlier during the port-switching period. In addition, the components (C) and (D) were eluted earlier as fresh feed was injected earlier in a port-switching time.