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Biofilms play a pivotal role in the progression of periodontitis and they can be treated with antiseptics (i.e. chlorhexidine) or antibiotics, but these therapeutic alternatives are unable of ameliorating periodontal alveolar bone loss, which has been, on the other hand, successfully treated with bone-preserving agents. The improved bone formation achieved in animal models by the combination of two such agents: bioactive glass (BAG) and bisphosphonates has attracted the interest for further exploring dental applications. However, the antimicrobial effects that may result from combining them have not been yet investigated. Here, our aim was to explore the anti-biofilm effects that could result from combining BAG with bisphosphonates, particularly in a dental biofilm model. The experiments were performed with an oral cavity single-specie (Aggregatibacter actinomycetemcomitans) biofilm assay, which was optimized in this contribution. Risedronate displayed an intrinsic anti-biofilm effect, and all bisphosphonates, except clodronate, reduced biofilm formation when combined with BAG. In particular, the anti-biofilm activity of risedronate was significantly increased by the combination with BAG. Since it has been proposed that some of the antimicrobial effects of BAG are caused by local pH changes, studies of pH variations were performed to gain a mechanistic understanding. However, the observed anti-biofilm effects could not be explained with lowered pHs. Overall, these results do provide further support for the promising use of bisphosphonate-BAG combinations in dental applications. These findings are particularly relevant for patients undergoing cancer chemotherapy, or osteoporotic patients, which are known to be more vulnerable to periodontitis. In such cases, bisphosphonate treatment could play a double positive effect: local treatment of periodontitis (in combination with BAG) and systemic treatment of osteoporosis, prevention of hypercalcemia and metastases.