3.2.2. Co-Precipitation Process with Controlled pH (MpH)

SP_pH nanoparticles were obtained based on the strategy of Saraiva et al. [20], which consists of a standard co-precipitation method at room temperature, using as precursors ferrous and ferric chlorides in a molar ratio [Fe³⁺/Fe²⁺] = 1:2. In a typical reaction, two aqueous solutions of FeCl₃.6H₂O (0.01 M) and FeCl₂.4H₂O (0.02 M) were mixed. The resulting solution was de-aerated with nitrogen gas (N₂) for 10 min to ensure the removal of oxygen during the co-precipitation process. To promote the precipitation of Fe₃O₄, NH₃ aqua (25% w/w) was slowly added until the pH of the solution was close to 9.6. The instantaneously formed SPION suspension was filtered through a 0.22 μm membrane, washed with Milli-Q water, and the obtained precipitated SP_pH were freeze–dried.