Abstract Bamboo is a kind of biological composites reinforced by unidirectional long fiber. Once there exists crack, the propagation of delamination is controlled by the interlaminar fracture toughness instead of by strength. In this paper, the end notched flexure (ENF) beam specimen was used to test the Mode II interlaminar fracture toughness G IIC along grain of Moso bamboo internode and the fracture surface was analyzed. The results were obtained that the Mode II interlaminar fracture toughness G IIC calculated by the experiment parameter substitution method was more accurate and the value was 1303.18 J/m 2 (coefficient of variation = 8.96%) which was about three times higher than the value of Mode I interlaminar fracture toughness; the crack propagation of Mode II interlaminar fracture was mainly self-similar cracking, but the fracture surface was rougher. Ground tissue in the zone of Mode II crack propagation was characterized by hackle shearing deformation. The SEM photos showed that ground tissue separated from fiber along middle lamella under shear stress and as the increasing of the dislocation of upper and lower layer, the thin-walled ground tissue would fracture transversely by tension, while to thick-walled fiber cell, only middle lamella and primary wall were torn then debonded, fragments remained.