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The motion of the skeletal estimated from skin attached marker-based motion capture(MOCAP) systems is known to be affected by significant bias caused by anatomical landmarks mislocation but especially by soft tissue artifacts (such as skin deformation and sliding, inertial effects and muscle contraction). As a consequence, the error associated with this bias can propagate to joint kinematics and kinetics data, particularly in small rodents. The purpose of this study was to perform a segmental kinematic analysis of the rat hindlimb during locomotion, using both global optimization as well as segmental optimization methods. Eight rats were evaluated for natural overground walking and motion of the right hindlimb was captured with an optoeletronic system while the animals walked in the track. Three-dimensional (3D) biomechanical analyses were carried out and hip, knee and ankle joint angular displacements and velocities were calculated. Comparison between both methods demonstrated that the magnitude of the kinematic error due to skin movement increases in the segmental optimization when compared with the global optimization method. The kinematic results assessed with the global optimization method matches more closely to the joint angles and ranges of motion calculated from bone-derived kinematics, being the knee and hip joints with more significant differences.