This study aims to analyze the impact of different 3D printed biomimetic midsole structures on gait phase and gait ratio to optimize running shoe design. Participants wore shoes with various 3D printed midsoles while gait ratio(heel rotation, foot balance, forefoot balance, hallux stiffness, medial forefoot balance, and meta loading) and gait phase(forefoot off phase, foot flat phase, forefoot contact phase, and initial contact phase) data were collected using a gait analysis system. These metrics reflect foot pressure distribution, gait symmetry, and balance, providing insights into how midsole structures affect gait. The results showed that different 3D printed biomimetic midsoles significantly impacted gait phase and gait ratio. The 1TS structure, improved the stability of the stance phase and reduced swing phase imbalance, demonstrating its potential for optimizing gait balance and efficiency. This indicates that 3D printed biomimetic midsoles can notably influence running gait, with the 1TS structure enhancing stability and balance, offering a new direction for optimizing running shoe design to improve performance and comfort.