In this study, Ca2+ was applied to enhance the photolysis H-2 production of Chlamydomonas reinhardtii grown in pure culture or co-cultivated with bacterium. With addition of Ca2+, the chlorophyll and protein contents in algal cells were maintained at high levels and the reactive oxygen species (ROS) were scavenged to tolerable levels by activities of catalase (CAT) and superoxide dismutase (SOD). High chlorophyll and low ROS are benefit to protect the photosystem II (PS II) activity that responsible for the direct photolysis H-2 production. Moreover, addition of Ca2+ was able to promote the acetic acid assimilation as well as the starch synthesis. The starch degradation, in return, contributed to the H-2 production via the indirect photolysis pathway. Based on the improvements of direct and indirect photolysis pathways, the H-2 production of algae grown in co-cultivation adding with 5 mM Ca2+ reached a maximum value of 306 mL L-1, which was 2.4 times higher than the production without Ca2+ addition. This study gives an efficient and easy strategy to improve the algal photolysis H-2 production, and gets an insight into the mechanisms of the improvement. (C) 2018 Hydrogen Energy Publications LLC. Published by Elsevier Ltd. All rights reserved.