Comparative Effect of Rhamnolipid and Sophorolipid on the Bioavailability of Residual Hydrocarbons for Biohydrogen Production via Dark Fermentation

Abstract

Abstract Limited hydrocarbon bioavailability remains a key bottleneck in coupling bioremediation with biohydrogen production through dark fermentation. This study examined the effects of two biosurfactants; rhamnolipid and sophorolipid on the solubilization of residual hydrocarbons and subsequent hydrogen generation by mixed anaerobic cultures. Surfactants were applied at 0.05–0.3% (w/v) under thermophilic (80 °C) conditions. Both treatments enhanced hydrocarbon solubilization compared to the control, with rhamnolipid achieving a 31% increase and sophorolipid a 24% increase in soluble hydrocarbon fractions. Correspondingly, cumulative hydrogen yields reached 52 ± 2 mL H₂/g chemical oxygen demand (COD) for rhamnolipid and 44 ± 3 mL H₂/g COD for sophorolipid, against 38 ± 2 mL H₂/g COD in the control. Kinetic analysis showed higher substrate degradation rates in rhamnolipid systems (k = 0.072 h⁻¹) than sophorolipid (k = 0.061 h⁻¹). Microbial profiling indicated enrichment of Clostridium butyricum and Bacillus subtilis under rhamnolipid treatment, correlating with improved hydrogen productivity. Overall, rhamnolipid demonstrated superior performance in enhancing hydrocarbon bioavailability and biohydrogen yield, suggesting its greater compatibility with dark fermentative consortia for integrated bioremediation and bioenergy recovery from depleted hydrocarbon reservoir.