There is a growing acceptance worldwide for the application of dispersants as a marine oil spill response strategy. The development of more effective dispersants with less toxicity and higher biodegradability would be a step forward in improving public acceptance and regulatory approvals for their use. By applying advances in environmental biotechnology, a bio-dispersant agent with a lipopeptide biosurfactant produced by Bacillus subtilis N3-1P as the key component was formulated in this study. The economic feasibility of producing biosurfactant (a high-added-value bioproduct) from fish waste-based peptone as a nutrient substrate was evaluated. Protein hydrolyzate was prepared from cod liver and head wastes obtained from fish processing facilities. Hydrolysis conditions (i.e., time, temperature, pH and enzyme to substrate level) for preparing protein hydrolyzates were optimized by response surface methodology using a factorial design. The critical micelle dilution (CMD) value for biosurfactant produced from the fish liver and head waste generated peptones was 54.72 and 47.59 CMD, respectively. Biosurfactant product generated by fish liver peptone had a low critical micelle concentration of 0.18 g L-1 and could reduce the surface tension of distilled water to 27.9 mN/m. Structure characterization proved that the generated biosurfactant product belongs to the lipopeptide class. An alternative to the key surfactant dioctyl sulfosuccinate sodium (DOSS) used in Corexit 9500 has been proposed based on a binary mixture of lipopeptides and DOSS that exhibited synergistic effects. Using the standard baffled flask test, a high dispersion efficiency of 76.8% for Alaska North Slope oil was achieved at a biodispersant composition of 80/20 (v/v) of lipopeptides/DOSS. The results show that fish waste can be utilized to produce a more effective, environmentally acceptable and cost-efficient biodispersant that can be applied to oil spills in the marine environment.