Purpose: To evaluate and compare intraoperative pain during upper eyelid blepharoplasty (UEB) between on first and second operated eyelids. Methods: In this cross-sectional observational study, the patients were divided into two groups, with group 1 representing 40 patients whose surgery was first started on the right and group 2 representing the other 40 patients who started surgery on the left first. UEB was performed to all patients by the same surgeon under the same and equal amount of local anesthesia. Degree of pain felt during surgery on first and second operated eyelid was evaluated with the visual analoge scale (VAS) and Wong-Baker Facial Pain Expression Scale (WBFPES) of all patients immediately after surgery and was compared using paired t-test. Results: There were 20 female and 20 male patients in both groups. In group 1, the VAS value was 2.8 ± 1.5 and WBFPES value was 2.7 ± 1.6 UEB in the first operated eyelid; the VAS value was 4.1 ± 1.8 and WBFPES value was 3.9 ± 1.8 UEB in the second operated eyelid. A significant difference was found between pains felt during first and second operated eyelid UEB regarding VAS and WBFPES values in group 1 (p = 0.003 and p = 0.002, respectively). In group 2, the VAS value was 1.9 ± 1.0 and WBFPES value was 2.0 ± 1.0 UEB in the first operated eyelid; the VAS value was 3.0 ± 1.5 and WBFPES value was 2.8 ± 1.6 UEB in the second operated eyelid. A significant difference was found between pains felt during first operated and second operated eyelid UEB regarding VAS and WBFPES values in group 2 (p = 0.009 and p = 0.005, respectively). Conclusions To the best of our knowledge, this study is the first to evaluate which eyelid is more painful during surgery in UEB. Patients tend to feel more pain during surgery on the second operated eyelid. Therefore, surgeons should consider using local anesthetics with more volume or longer duration in the second operating eyelid in light of this information and patients should be given detailed information about pain.