The human population is growing rapidly—it is expected to reach 9 billion by 2050 [1]—and people are living for longer periods, which, in association with poor nutrition and sedentarism, has increased the incidence of age-related metabolic diseases such as osteoporosis [2]. Microalgae are a valuable source of bioactive compounds that affect bone formation and mineralization. Methanolic extracts of Chlorella sp., Dunaliella salina and Phaeodactylum tricornutum were screened for osteoactive compounds using a zebrafish operculum assay and a gilthead seabream vertebrae-derived VSa13 cell line. Three concentrations (10, 31.6 and 100 µg/ml) were evaluated for each extract. While no significant effect was observed on opercular bone mineralization for any of the extracts and concentrations used, the extracts prepared from Chlorella sp. at all concentrations tested and D. salina extract at the highest concentration decreased the mineralization of the VSa13 extracellular matrix when compared to the control (0.1% DMSO). This anti-mineralogenic effect was associated with a change in cell morphology, suggesting that it may be the consequence of altered cell differentiation. On the other hand, extracts from P. tricornutum increased extracellular matrix mineralization by around 4% at all concentrations tested. This pro-mineralogenic effect may be related to the presence of high amounts of polyphenols in methanolic extracts. Polyphenols are secondary metabolites with a wide range of biological activities [3,4], in particular with the ability to stimulate bone formation and mineralization, but also osteoblast proliferation and differentiation [5]. Our results suggest that extracts from P. tricornutum may represent a promising source of nutrients that can potentially be used for the treatment or prevention of osteoporosis.