Biohydrogen Production by Locally Isolated Purple Non-Sulfur Bacteria: Strain Characterization, OFAT Optimization, and Evaluation of Silver Nanoparticle Effects

Abstract

This study investigated biohydrogen production by locally isolated purple non-sulfur bacteria (PNSB), focusing on strain characterization, process optimization, and the effects of silver nanoparticles (AgNPs). Two isolates were obtained and identified, with one strain closely related to Rhodopseudomonas palustris (PNSB_P) demonstrating superior hydrogen-producing capability, while the second strain (PNSB_B) showed no detectable production. Growth kinetics revealed that PNSB_P exhibited efficient biomass accumulation despite a lower specific growth rate, indicating effective metabolic allocation toward hydrogen production. Optimization using a one-factor-at-a-time (OFAT) approach showed that carbon source and growth conditions significantly influenced hydrogen yield (p < 0.05), with maltose and strict light–anaerobic conditions producing the highest outputs. In contrast, inoculum size and temperature showed no significant effects (p > 0.05) and baseline conditions were retained. Under combined optimized conditions, cumulative hydrogen production reached 194.86 mmol L⁻¹. Supplementation with AgNPs resulted in a slight, non-significant enhancement, with the highest yield (198.28 mmol L⁻¹) observed at 0.08 g L⁻¹, while higher concentrations exhibited inhibitory effects. Overall, the findings highlight the importance of strain selection and process optimization in enhancing photofermentative biohydrogen production and demonstrate the limited but concentration-dependent influence of nanoparticles.