This study was devoted to the comprehensive thermodynamic modeling of biohydrogen production using the light‐dependent bacterium Rhodospirillum rubrum. Conventional exergy and eco‐exergy concepts were employed for decision making on the carbon source concentration and light intensity. Several thermodynamic indicators were applied to identify optimal experimental variables with respect to the sustainability issues. The results obtained using both concepts were remarkably different from each other because of the incorporation of the work of the information embodied in the genomes of the living microorganisms considered in the eco‐exergetic computations. However, both analyses identified 1 g L−1 substrate concentration and 1000 lux light intensity as optimal fermentation conditions. Under these conditions, the normalized exergy destruction as a decision making parameter was 179.7 and 171.54 kJ kJnormalH2
−1 using the conventional exergy and eco‐exergy concepts, respectively.