Mature coconut water (MCW) is a natural beverage and a main by-product of various coconut processing industries such as virgin coconut oil, coconut chips, coconut milk, etc. In spite of huge benefits, MCW’s short shelf life limits its market potential. In this context, the present study investigates the effect of ultrasonic processing parameters, such as amplitude (50%, 60% and 70%) and time (5 min, 10 min and 15 min), on microbial population and quality profile (pH, total soluble solids, total sugars, reducing sugars and non-reducing sugars) of MCW. Central composite design was used to create a multiple linear regression model for each response and to optimize ultrasound processing parameters. The optimal treatment parameters to ensure microbial safety and preserve the nutritional quality of MCW were 60% ultrasonic amplitude and treatment time of 10 min. Total sugars, reducing sugars, non-reducing sugars and microbial load of MCW determined at optimized conditions were 4.92%, 2.804%, 2.13% and 4.79 log cfu/mL, respectively. Ultrasonic treatment was found to be effective in inhibiting microbial growth and maintaining non-reducing sugars of MCW.