The role of pH control on biohydrogen production by co-culture of dark-fermentative Clostridium acetobutylicum and photofermentative Rhodobacter sphaeroides was studied. Single stage dark fermentation, photofermentation and hybrid co-culture systems were studied at different values of controlled and uncontrolled pH. Increasing pH during dark fermentation resulted in lower hydrogen production rate (HPR) and longer lag time for both controlled and uncontrolled conditions. However, it only slightly affected cumulative H-2 volume. Results have shown that pH control at pH 7.5 increased photofermentative hydrogen production from 0.966 to 2.502 L H-2/L-medium when compared to uncontrolled process. Fixed pH value has proven to be an important control strategy also for the hybrid process and resulted in obtaining balanced co-culture of dark and photofermentative bacteria. Control of pH at 7.0 was found optimum for bacteria cooperation in the co-culture what resulted in obtaining 2.533 L H-2/L-medium and H2 yield of 6.22 mol H-2/mol glucose. (C) 2015 Elsevier Ltd. All rights reserved.