To develop new remediation methods for acidic soils polluted by heavy metals, the chemical fractions of Cu(II), Pb(II) and Cd(II) in an Ultisol with and without rice straw biochar were compared and the effect of biochar incorporation on the mobility and bioavailability of these metals was investigated. In light of the decreasing zeta potential and increasing CEC, the incorporation of biochar made the negative soil surface charge more negative. Additionally, the soil pH increased markedly after the addition of biochar. These changes in soil properties were advantageous for heavy metal immobilization in the bulk soil. The acid soluble Cu(II) and Pb(II) decreased by 19.7-100.0% and 18.8-77.0%, respectively, as the amount of biochar added increased. The descending range of acid soluble Cd(II) was 5.6-14.1%, which was much lower than that of Cu(II) and Pb(II). When 5.0 mmol/kg of these heavy metals was added, the reducible Pb(II) for treatments containing 3% and 5% biochar was 2.0 and 3.0 times higher than that of samples without biochar, while the reducible Cu(II) increased by 61.6% and 132.6% for the corresponding treatments, respectively. When 3% and 5% biochar was added, the oxidizable portion of Pb(II) increased by 1.18 and 1.94 times, respectively, while the oxidizable portion of Cu(II) increased by 8.13 and 7.16 times, respectively, primarily due to the high adsorption affinity of functional groups of biochar to Cu(II). The residual heavy metal contents were low and changed little with the incorporation of biochar.