In this study, a photobioreactor cultivation system and a calculation method for on-line monitoring of carbon and energy metabolism of microalgae were developed using Nannochloropsis sp. in nitrogen-repletion and nitrogen-limitation cultures. Only 30-60% of carbon fixed in Calvin cycle was used for biomass and the rest was lost in light respiration. The net fixed carbon was assumed to be incorporated into protein, lipids, carbohydrates, and nucleic acids, whose contents calculated on-line fitted well with the experimental measurements. Intracellular ATPs were quantitatively divided for biomass production and cell maintenance, and the result is in accordance with known reports. Due to light limitation induced by high cell concentration in batch cultures, the proportion of CO2 loss in light respiration and the proportion of energy for maintenance rapidly increased in culturing process. Nitrogen starvation reduced the light respiration, thus decreasing CO2 loss and maintenance energy, but no effect on ATP requirement for cell growth. (C) 2014 Elsevier Ltd. All rights reserved.