A separation phenomenon occurring during the drop weight tear test of commercial thick-walled API (American Petroleum Institute) X80 strip steel was investigated in this work. Microstructural analysis showed that the band structure of bainite elongated along the rolling direction works as the initiation sites of separation. The propagation of separation can be promoted not only by the occurrence of the band structure of martensite/austenite constituent, prior austenite grain boundaries, and elongated bainite, but also by fine acicular ferrite and bainite. Wide separation formed in the former case, while the narrow one appeared in the latter case. Some methods were proposed to obtain fine and homogeneous acicular ferrite in thick-walled X80 pipeline steel in order to minimize the occurrence of separation.