Detecting the initiation of mechanical injury to biological tissue, and not just its ultimate failure, is critical to enable a sensitive and specific characterization of tissue tolerance, to develop quantitative relationships between macro-and microstructural tissue responses, and to appropriately interpret physiological responses to loading. We have developed a novel methodological approach to detect the onset and spatial location of structural damage in collagenous soft tissue before its visible rupture, via the identification of atypical regional collagen fiber kinematics during loading. Our methods utilize high-speed quantitative polarized light imaging and identify the onset of tissue damage in ligament regions where mean collagen fiber rotation significantly deviates from its behavior during noninjurious loading. This technique was validated by its ability to predict the location of visible rupture (p=0.0009). This fiber rotation-based metric of damage identifies potential facet capsular ligament injury beginning well-before rupture, at 51±12% of the displacement required to produce tissue failure. While traditional macro-scale strain metrics fail to identify the location of microstructural damage, initial injury detection determined by altered fiber rotation was significantly correlated (R=0.757; p=0.049) with tissue yield (defined by a decrease in stiffness), supporting the capabilities of this method. Damaged regions exhibited higher variance in fiber direction compared to undamaged regions (p=0.0412).