Abstract The objective of the present study is to improve the biohydrogen production from marine macroalgae (Ulva reticulate) by acidic - hydrogen peroxide (H2O2) induced microwave (AHMW) pretreatment. Higher soluble chemical oxygen demand (SCOD) release of 1450 mg/L and its liquefaction rate of 30.2% was achieved in microwave (MW) pretreatment with treatment time period of 15 mins. Varying concentration of H2O2 from 0.003 to 0.03 g/g TS were used in the optimal microwave power (40%) to enhance the organic release in H2O2 induced microwave pretreatment (HMW). Maximum liquefaction of 33.9% was obtained at the H2O2 concentration of 0.024 g /g TS. The combined HMW pretreatment under acidic (pH 4–6.5) show synergistic effect on organic release. At optimal pH 5, AHMW pretreatment shows the SCOD release of 1850 mg/L with its liquefaction of 38.5% at time of 10 min. Therefore, AHMW pretreatment significantly reduce the treatment time and increase liquefaction when compared to MW and HMW. The maximum biohydrogen production was observed as 92.5 mL H2/g COD in AHMW pretreatment.