This study involved optimisation of acid hydrolysis in ethanol production from prosopis juliflora. A two level full factorial design with four factors, two levels and two replicas (24 2=32 experimental runs) was applied to optimize acid hydrolysis and study the interaction effects of acid hydrolysis factors, namely, acid concentration, solid fraction, temperature, and time. The screening of significant acid hydrolysis factors were done by using the two-level full factorial design using design expert® 7 software.