Green microalgae are likely to play an important role in bioregenerative systems for producing food and converting CO2 to O2 in a controlled ecological life support system (CELSS). In the present study, a method for evaluating the effects of environmental variables on the multiplication rate of microalgal cells was developed to determine the optimum culture condition for a microalgal culture system that can function effectively in the CELSS. The microalga, Euglena gracilis, was cultured in water droplets (3 microliters in liquid volume each) in a vessel (25 ml in air volume) in which the CO2 and O2 concentrations were controlled. The number of Euglena cells cultured at CO2 concentrations ranging from 2% to 6%, O2 concentrations ranging from 5% to 20%, and PPF levels ranging from 50 to 100 micromoles m-2 s-1 was monitored by using a video camera and a microscope. The multiplication rate of cells was highest and the cell number increased by 8.3 times during 48 h under a condition of 4% CO2, 21% O2 and 100 micromoles m-2 s-1 PPF. The multiplication rate of the cells was highest at 4% CO2, followed by 6% and 2% CO2, and it decreased with decreasing O2 concentration and decreasing PPF.