Introduction: Laryngeal stroboscopy (LS) is crucial while studying the mucosal wave (MW) and its characteristics. However, there may be limitations of the technique causing changes in the dynamics of the free edges (FEs), which could lead to uncertainty when establishing clinical judgements. This investigation aimed to determine if there is a different biomechanical model in phonation and MW during LS exploration using a flexible endoscope. Methods: This descriptive study was conducted on 137 adult participants divided into the following groups: control, functional FEs pathology and organic FEs pathology. Biomechanical analysis was conducted using the Voice Online Lab® App with voice recordings in natural pitch and volume, before and simultaneously with LS. Distribution and mean difference values were obtained according to phonation condition, group, and gender. Student’s t-test was used to assess significant differences and test the hypotheses. Results: During LS, the following was evident in the entire population with significant values (P < 0.05): an increase in the mean values of fundamental frequency, a decrease in tension and force of glottic closure, an increase in glottal gap and a decrease in all MW correlate values in both phases of the vocal cycle. These differences were greater in men (P < 0.05) and were significant in the organic pathology group. Conclusion: LS conducted with a flexible endoscope plays a major role on the dynamics of FEs, and these changes are more pronounced in men. There may be a more pronounced compensatory laryngeal overstrain in FEs with a consolidated pathology compared with the functional pathology and control groups. Professionals employing LS in their clinical practice should consider these factors.