The current study has been done with the goal of biodiesel synthesis of sour cherry kernel oil (SCKO). SCKO is a potential novel and free source for biodiesel production. SCKs have an oil content of similar to 25%, so the SCKs can be considered as a suitable source for producing fuel from wastes. SCKO has a low acid value (0.9 mg KOH/g). Therefore, the transesterification reaction performed to produce biodiesel from SCKO in one step. Then, the physicochemical properties of biodiesel analyzed. The transesterification reaction performed in the constant condition of temperature of 65 degrees C, cycle of 1 s, and amplitude of 100%. The effects of process parameters including catalyst amount, time, and methanol: oil molar ratio on biodiesel yield investigated and the reaction process optimized by using the response surface methodology (RSM). The results showed that the maximum biodiesel yield of 91.9% obtains in the condition of the methanol: oil molar ratio of 8.21:1, catalyst amount of 1.35 w/w%, and reaction time of 2.5 min. Moreover, the result of the Physicochemical properties of SCKO biodiesel are as follows: density of 0.881 gr/cm(3), kinematic viscosity of 5.03 mm(2)/s, flash point of 158 degrees C, cloud point of 5 degrees C, and pour point of (-7) degrees C. All properties of the produced biodiesel of SCKO complies with standard ASTM 6751 and EN 14214. Moreover, in comparison of the diesel fuel, biodiesel obtained from SCKO enhance properties including cloud point, flash point, and pour point. Hence, SCKO is an attractive and valuable source for biodiesel production.