Very high cycle fatigue (VHCF) data and experiments, 107–109 cycles to failure, have traditionally been both a cumbersome and costly task to perform. However, characterizing VHCF behavior of material systems is critical for the design and sustainability of turbine engines as outlined in the turbine engine structural integrity program (ENSIP). With recent advancements, ultrasonic fatigue (UF) test systems have become increasingly available to generate VHCF fatigue data. A primary consideration for ultrasonic fatigue testing is the frequency of loading, the resulting thermal evolution, and its effect on fatigue life. To mitigate the heat generation within the specimen during experiments, cooling air is directed to the specimen and cyclic loading is performed by selecting an appropriate test frequency or defining a duty cycle rather than continuously subjected to fatigue. However, standardization of experimental test procedures remains ongoing and continues to be developed. In this study, a Shimadzu USF-1000A ultrasonic fatigue test system is used to characterize VHCF behavior of Ti-6Al-4V to understand the effect of duty cycle and thermal evolution on fatigue life for ultrasonic fatigue testing. Titanium 6Al-4V test specimens are subjected to fully reversed axial fatigue at 20 kHz exciting resonance in an axial mode to better characterize the experimental process. Three duty cycle-cooling air configurations and their effect on fatigue life due to self-generated heat during the experiment are investigated. Heat generation is monitored in situ via a single-point optical pyrometer, and in situ mechanical and thermal data are collected and compared to standardized servohydraulic fatigue test data performed in this study as well as from data found in the literature.