Several reducing agents had a positive effect on the elimination of photosynthetic O2 in culture medium. However, some chemicals, such as hydroquinone and ascorbic acid, had adverse effects on microalgae growth. Cysteine was the only reducing agent that did not inhibit the growth of C. gloeopara and S. obliquus. The depletion of photosynthetic O2 improved the anaerobic H2 production, because photosynthesis and carbohydrate metabolism were both active. H2 accumulation was between 3 and 5 times greater when dissolved O2 was scavenger than under sulfur deprivation. This study demonstrates the concept of scavenging photosynthetic O2 as soon as it is produced by microalgae which was enough for inducing the metabolism of H2 production.