In this study, an experimental design was employed to optimize the fabrication of the YCa2Cu3O7 superconductor composite. The superconductors were fabricated using biocompatible calcium carbonate nanoparticles extracted from cuttlebone, Sepia pharaonis, of the Persian Gulf in the Bushehr coastal area, and their effects on YCa2Cu3O7 superconductor’s properties were investigated. Ball milling process and solid-state calcination reaction were used for obtaining the natural calcium carbonate nanoparticles and fabrication of the superconductors, respectively. The effects of three factors, milling time and the excess amount of the natural calcium carbonate as well as yttrium oxide, were investigated on the superconductivity properties of the products. The Taguchi statistical design of experiments was conducted to reveal the sensitivity analysis of the products’ properties due to the variation in each variable, finding optimal production conditions, and reduction in the total number of required experiments. In this method, the oxygen content of the fabricated superconductor was considered as the targeted YCa2Cu3O7 property. Moreover, the morphology and structural properties of the products were investigated to find the relationship between physicochemical properties and superconductivity. The YCa2Cu3O7 superconductors were characterized by the Meissner effect test, XRD analysis, and FESEM imaging. The sensitivity analysis was shown that by increasing milling time and the excess amount of the reactants in the solid-state reaction, the connectivity and crystallization of the grains were improved, and the optimum operating conditions for the production were achieved. Besides, the variation in milling time causes more signal-to-noise ratio, which indicates the highest efficacy of this factor on the superconductivity of the products in comparison with the other variables.