Background: In radiotherapy (RT), the dose given to the organs at risk (OAR) is as important as the dose received by the tumor. FF (Flattening Filtered) and FFF (Flattening Filter Free) treatment techniques used in RT offer different advantages in terms of their effects on target and critical organs. Treatment duration varies depending on the number of monitor units (MU) delivered per unit time. The aim of this study was to investigate biochemically and histopathologically the damage in healthy laryngeal tissue exposed to 600 MU/min FF and 2400 MU/min FF ionizing radiation under different fractionation protocols. Also to investigate the radioprotective effect of melatonin against the radiological damage. Methods: The 78 rats included in the study were divided into 10 groups. Control and melatonin-only groups did not receive RT. The other 8 groups received FF and FFF ionizing radiation in different dose fractions. Half of the radiation group received melatonin 15 min before RT. Results: No histological and biochemical differences were observed in the control and MLT only groups. Radiobiological damage was statistically significantly lower in the RT+MLT group compared to the RT only group. Conclusions: It has been observed that melatonin has a protective effect against tissue damage caused by radiation exposure in healthy laryngeal tissue.