Limited studies had been conducted using water-in-diesel emulsion produced from ultrasonic homogenizing method. In this study, Water-in-Biodiesel Emulsions (WiBE) produced using ultrasonic homogenizer were characterized and studied for their physical and chemical properties through various laboratory investigations. The data were then compared with WiBE produced using mechanical homogeniser by the current researchers. Physical characterization tests were carried out on 24 WiBE emulsions produced using an ultrasonic bath, with water percentage of 9, 12 and 15, HLB value of 6, 7, 8 and 9, and surfactant dosage of 5 and 10. The water droplets produced using ultrasonic homogeniser were found to be evenly distributed and generally smaller in size. The density and viscosity values of these emulsions were found to be uniformly larger than WiBE produced using mechanical homogeniser. Emulsions with 15 water exceeded the density threshold, indicating the limits of the amount of water which can be added to the biodiesel fuel. Also, it was found that for viscosity, the HLB is limited to HLB 9 for higher surfactant dosage. © 2018 The Authors, published by EDP Sciences.