Anemia due to iron deficiency is a global problem that requires serious awareness. Iron deficiency can be caused by the lack of iron intake, and low absorption of iron. It is known that peptides obtained from protein hydrolysates can generally act as iron chelating. Peptides can increase iron solubility, bioavailability, absorption and stability. BPPT (Agency for the Assessment and Application of Technology) developed soybean hydrolysates, which was obtained by thermal and enzymatic hydrolysis. Soybean hydrolysates had been tested for iron binding using in vitro test. The aim of this study was to characterize peptides from soybean hydrolysates which had iron binding activity. This research was conducted with two stages of separation, which were using 10 kD Molecular Weight Cut Off ultrafiltration membrane and gel filtration chromatography used the Superdex 30 column (GE Healthcare, USA) was coupled with Aktaprime. Furthermore, to find out the character of the peptide which acts as a promoter of iron binding, analysis of protein molecular weights using SDS-PAGE and analysis of composition amino acids using HPLC were performed. The results showed that the peptides from soybean hydrolysate which act as iron binder had a molecular weight of 10 kDa. Composition of amino acids in the fraction were dominated by arginine, aspartic acid and glutamic acid.