From an engineering standpoint, investigating the effects of rotation speed and fulvic acid concentration on the development of secondary high-iron minerals is crucial for treating acid mine drainage. The Fe2+ oxidation mechanism by Acidithiobacillus (A.) ferrooxidans to synthesise secondary high-iron minerals was examined in this study using shaking flask tests under various conditions: fulvic acid concentrations of 0, 0.2, or 0.4 g/L and rotation speeds of 180 r/min or 100 r/min. The pH, Fe2+ oxidation rate, total iron precipitation rate, secondary high-iron mineral functional groups and ore equivalent indicators were also investigated. The results demonstrated that at a fulvic acid concentration of 0 g/L, the pH decreased from 2.5 to 2.17 at 180 r/min. At 0.2 g/L, it decreased from 2.5 to 2.05. Finally, at 0.4 g/L, it decreased from 2.5 to 2.07. Fe2+ was completely oxidised after 48 h, and the final total iron precipitation rate ranged from 26.2% to 33.4%. The synthesised secondary high-iron minerals were uniformly dispersed in the solution. When the rotation speed was 100 r/min, the pH reduced from 2.5 to 2.25 at a fulvic acid concentration of 0 g/L, from 2.5 to 2.14 at 0.2 g/L, and from 2.5 to 2.19 at 0.4 g/L. Notably, Fe2+ was completely oxidised within 72 h. The experiment’s final iron precipitation rate ranged from 23.6 to 29.6%. The synthesised secondary high-iron minerals were blocky and adhered to the bottom of the shaking flask. In summary, at a rotation speed of 180 r/min or 100 r/min, the Fe2+ oxidation rate and total iron precipitation rate of the experimental group with a fulvic acid concentration of 0.2 g/L were higher than those of the control group and the experimental group with a fulvic acid concentration of 0.4 g/L. A fulvic acid concentration of 0.2 g/L enhanced the activity of A. ferrooxidans. The minerals obtained from these experiments were characterised and identified as schwertmannite and jarosite.