Eucalyptus nitens is a fast-growing wood species with a relevant presence in countries like Australia and Chile. The sustainable construction goals have driven the search of structural applications for Eucalyptus nitens; however, this process has been complicated due to the defects usually presented in these timber boards. This study aims to evaluate the dynamic elasticity modulus(Exd) of Eucalyptus nitens timber boards through non-destructive vibration-based tests. Thirty-six timber boards with different levels of knots and cracks were instrumented and tested in a simply supported condition by measuring longitudinal and transverse vibrations. In the first stage, the Exd was calculated globally through simplified normative formulas. Then, in a second stage, the local variability of the Exd was estimated using operational modal analysis (OMA), finite element numerical simulations (FEM), and regional sensitivity analysis (RSA). The positive correlation found between the global static modulus of elasticity and Exd suggests that non-destructive techniques could be used as a reliable and fast alternative for the assessment of bending stiffness. Finally, the proposed method to estimate the local variability of Exdt based on the combination of OMA, FEM,and RSA techniques was useful to improve the structural selection process of timber boards forlightweight social housing floors.