Background: Few original articles describe the perioperative outcomes of uniportal thoracoscopic segmentectomy using a unidirectional dissection approach. In this retrospective study, we evaluated the feasibility and safety of this procedure. Methods: This study included 119 patients who underwent uniportal thoracoscopic segmentectomy in our department between February 2019 and December 2022. The patients were divided into unidirectional (group U, n = 28) and conventional (group C, n = 91) dissection approach groups. While the dominant pulmonary vessels and bronchi were transected at the hilum without dissecting a fissure in the unidirectional (U) group, the dominant pulmonary artery was exposed and divided at a fissure in the conventional (C) group. Patient characteristics and perioperative outcomes were compared between groups U and C. Results: The proportions of simple and complex segmentectomies were statistically similar between the groups. The operating time was shorter (group U: 110 [interqurtile range: 90–140] min, group C: 135 [interqurtile range: 105–166] min, p = 0.012) and there was less blood loss (group U: 0 [interqurtile range: 0–0] g, group C: 0 [interqurtile range: 0–50] g, p = 0.003) in group U than in group C. However, there were no significant intergroup differences in other perioperative outcomes. Conclusions: The unidirectional dissection approach in uniportal thoracoscopic pulmonary segmentectomy is safe and feasible and enables a smoother operation.