Single-position prone lateral lumbar interbody fusion (LLIF) has emerged as an alternative to the conventional two-position technique requiring patient repositioning from the lateral decubitus (LD) to the prone position. Because LLIF utilizes a retroperitoneal approach, the spatial relationship between the surgical corridor and retroperitoneal structures is a critical safety consideration. This study evaluated positional changes of retroperitoneal structures between the LD and prone positions using positional MRI in patients with degenerative lumbar spine disease. Seventy patients scheduled for LLIF were enrolled in this multicenter study. Lumbar MRI was performed in the right LD and prone positions using a platform simulating an operating table. At the L4–5 level, distances were measured from the anterior and middle vertebral lines (AVL and MVL) to the posterior border of the descending colon, the centers of the abdominal aorta and inferior vena cava, and the lateral and posterior borders of the left kidney. Paired statistical comparisons were performed, and the influence of body mass index (BMI) was assessed using simple linear regression. At L4–5, the descending colon shifted significantly more dorsally in the prone position than in the LD position, although the displacement was small (approximately 10 mm). The abdominal aorta showed a slight rightward shift in the LD position, whereas no significant positional change occurred in the inferior vena cava. The lateral border of the left kidney was significantly more lateral in the prone position. Higher BMI correlated with increased skin-to-disc distance and anterior displacement of the colon, particularly in the prone position. Overall, prone positioning produced only minimal positional changes in retroperitoneal structures relative to the standard LD position. This suggests that prone LLIF does not substantially increase the risk of visceral and vascular injuries. However, obesity significantly deepens the surgical corridor, particularly in the prone position, which warrants careful consideration during surgical planning.