A joint research team at University College London systematically investigated the influence of substrate microstructure on defect formation during laser powder bed fusion of aluminum alloys. Using a high-performance Al-Ni-Ce-Mn-Fe alloy (PA1) specially designed for additive manufacturing, single-track printing experiments were conducted on as-cast PA1 and rolled Al6061 substrates, tracked by high-speed synchrotron X-ray imaging. Results showed that large columnar intermetallic compounds in the as-cast PA1 substrate strongly induced defects during the initial printed layers, but this effect diminished as microstructure homogenized with increasing layer count. The study also identified a new defect type—"intermetallic-induced porosity"—originating from the interaction between laser keyholes and pre-existing intermetallic compounds in the substrate. These findings offer direct reference value for verifying powder batch consistency and the adaptability of consumable process windows.