Aim Defining the origins of muscle injury has important rehabilitation and exercise applications. However, current knowledge of muscle damage mechanics in human remains unclear in vivo. This study aimed to determine the relationships between muscle–tendon unit mechanics during maximal eccentric contractions and the extent of subsequent functional impairments induced by muscle damage. Methods The length of the muscle–tendon unit, fascicles and tendinous tissues was continuously measured on the gastrocnemius medialis using ultrasonography, in time with torque, during 10 sets of 30 maximal eccentric contractions of plantar flexors at 45°s−1, in seventeen participants. Results Muscle–tendon unit, fascicles and tendinous tissues were stretched up to 4.44 ± 0.33 cm, 2.31 ± 0.64 cm and 1.92 ± 0.61 cm respectively. Fascicle stretch length, lengthening amplitude and negative fascicle work beyond slack length were significantly correlated with the force decrease 48 h post‐exercise (r = 0.51, 0.47 and 0.68, respectively; P < 0.05). Conclusions This study demonstrates that the strain applied to human muscle fibres during eccentric contractions strongly influences the magnitude of muscle damage in vivo. Achilles tendon compliance decreases the amount of strain, while architectural gear ratio may moderately contribute to attenuating muscle fascicle lengthening and hence muscle damage. Further studies are necessary to explore the impact of various types of task to fully understand the contribution of muscle–tendon interactions during active lengthening to muscle damage.