This study aimed to compare different protocols (Protocol 1: P1; Protocol 2: P2; Protocol 3: P3; Protocol 4: P4) for the extraction of spongin-like collagen (SC) from marine sponges. The SEM micrographs demonstrated a fibrillar structure for the extracts from Chondrilla caribensis and the nodular/particulate aggregates for Aplysina fulva. FTIR showed for all samples peaks similar to collagen for both species. For C. caribensis, the extracts obtained using P2, P3, and P4 protocols presented higher values of extraction yield, TPQ, and GAGs. P2 and P4 showed higher values of SC concentration and for antioxidant analysis. For A. fulva, P2, P3, and P4 provided a higher extraction yield besides an increase in the antioxidant assay. For both species, no difference was observed for Col quantification and TPQ analysis; also, higher values of GAGs were found using P2 and P4. Fibroblast proliferation observed for C. caribensis was lower for P1 on day 1 and for P2 and P3 on day 3 (for 50%) compared to the control group. There was a significant reduction in fibroblast cell proliferation for all A. fulva extracts evaluated. It can be concluded that protocols P2 and P4 were more efficient for extracting SC from C. caribensis.