In this study, we demonstrated a new method of achieving long-term photodependent hydrogen production under nitrogen limiting conditions in a medium with natural seawater. Moreover, centrifugation stage in pretreatment of culture was avoided. Chlorella pyrenoidosa passed into deprivation conditions after exhaustion of nitrogen source. Microalgae began to release hydrogen at 144 h of cultivation after stages of evolution and consumption of oxygen. C. pyrenoidosa produced 45 ml (183 mu M) of hydrogen per liter of culture. A significant effect of the initial oxygen concentration in the gas phase on the release of hydrogen was found. When initial oxygen content was 3.5%, the final H 2 yields reduced by 20%, while under 9.7%-21% of O-2, it reduced by 50% (compared to argon gas phase). Perhaps this was due to the low content of cells. We also demonstrated a decrease in the release of hydrogen by C. pyrenoidosa by 85% when using double stress conditions: phosphate-deprivation and limiting nitrogen source. (C) 2019 Hydrogen Energy Publications LLC. Published by Elsevier Ltd. All rights reserved.