This study investigates the seismic performance of a full-scale masonry barrel vault initially tested and damaged in its unreinforced state through shaking table tests. The damaged vault was subsequently retrofitted with a hemp-rope-based bio-composite reinforcement applied to the extrados and re-tested under seismic dynamic loading. The response was quantitatively evaluated via modal parameter identification using the Enhanced Frequency Domain Decomposition (EFDD) method and vibration-based damage indicators, including frequency shifts and Modal Assurance Criteria (MAC). Results demonstrate that the bio-composite reinforcement significantly mitigates stiffness degradation and modal parameters variations during seismic excitation, delaying damage progression and improving structural residual capacity. The strengthened vault kept higher vibration frequencies and exhibited reduced eigenvector variations compared to the unreinforced counterpart when subjected to the same input level. This confirmed the effectiveness of the reinforcement system in enhancing the seismic capacity of previously damaged masonry structures. These findings provide valuable experimental evidence supporting the use of sustainable bio-composite materials for the conservation and seismic upgrading of historic masonry vaults.