By integrating fiber-reinforced composites (FRCs) with Three-dimensional (3D) printing, the flexibility of lightweight structures was promoted while eliminating the mold’s limitations. The design of the I-beam configuration was performed according to the equal-strength philosophy. Then, a multi-objective optimization analysis was conducted based on the NSGA-II algorithm. 3D printing was utilized to fabricate I-beams in three kinds of configurations and seven distinct materials. The flexural properties of the primitive (P-type), the designed (D-type), and the optimized (O-type) configurations were verified via three-point bending testing at a speed of 2 mm/min. Further, by combining different reinforcements, including continuous carbon fibers (CCFs), short carbon fibers (SCFs), and short glass fibers (SGFs) and distinct matrices, including polyamides (PAs), and polylactides (PLAs), the 3D-printed I-beams were studied experimentally. The results indicate that designed and optimized I-beams exhibit a 14.46% and 30.05% increase in the stiffness-to-mass ratio and a 7.83% and 40.59% increment in the load-to-mass ratio, respectively. The CCFs and SCFs result in an outstanding accretion in the flexural properties of 3D-printed I-beams, while the accretion is 2926% and 1070% in the stiffness-to-mass ratio and 656.7% and 344.4% in the load-to-mass ratio, respectively. For the matrix, PAs are a superior choice compared to PLAs for enhancing the positive impact of reinforcements.