In this study, plant (SAP, TAN) and microbial (RAM) biosurfactants were applied to two soils, in single or sequential soil washing. Soil 1 was moderately acidic, with higher content of organic matter and HMs (Cd, Cu, Ni, Pb and Zn), and was aged for 24 months. Soil 2 was alkaline, with lower content of organic matter and HMs, and was aged for 1 month. The stability of the HMs (as I R), except for Pb, was higher in soil 1 than in soil 2, but the rankings of HM stability were similar: Cu > Ni ≈ Pb > Zn > Cd (soil 1) and Cu > Pb > Ni > Zn > Cd (soil 2). The HMs were removed more efficiently from soil 2 than from soil 1, mainly from the readily available fraction and partially from the reducible fraction. Cu (0.3-0.5 times), Ni (0.4-1.1 times) and Zn (0.5-1.0 times) showed the largest increases in stability after soil washing with SAP. With a single washing, SAP and TAN were the least effective for Pb removal (18-31% and 11-35%, respectively), while RAM was the most effective for Cd (69-73%) and Pb (58-63%) removal. Sequential soil washing, especially with SAP followed by RAM, was the best choice for simultaneous removal of multi-HMs.