Unlike other antiresorptive medications, bisphosphonate molecules accumulate in the bone matrix. Previous studies of side-effects of anti-resorptive treatment focused mainly on systemic effects. We hypothesize that matrix-bound bisphosphonate molecules contribute to the pathogenesis of bisphosphonate-related osteonecrosis of the jaw (BRONJ). In this study, we examined the effect of matrix-bound bisphosphonates on osteoclast differentiation in vitro using TRAP staining and resorption assay, with and without pretreatment with EDTA. We also tested the effect of zoledronate chelation on the healing of post-extraction defect in rats. Our results confirmed that bisphosphonates bind to, and can be chelated from, mineralized matrix in vitro in a dose-dependent manner. Matrix-bound bisphosphonates impaired the differentiation of osteoclasts, evidenced by TRAP activity and resorption assay. Zoledronate-treated rats that underwent bilateral dental extraction with unilateral EDTA treatment showed significant improvement in mucosal healing and micro-CT analysis on the chelated sides. The results suggest that matrix-bound bisphosphonates are accessible to osteoclasts and chelating agents and contribute to the pathogenesis of BRONJ. The use of topical chelating agents is a promising strategy for the prevention of BRONJ following dental procedures in bisphosphonate-treated patients.