In studies of the inorganic carbon (Ci) concentrating mechanism of cyanobacteria glycolaldehyde is often used to inhibit photosynthetic CO2fixation. As a partial inhibition of active CO2transport by glycolaldehyde under steady-state Ciflux conditions has been recently suggested, in this paper, using mass spectrometry, we investigate in detail the relationship between glycolaldehyde concentration ([GLY]) and CO2transport both during the initial period following illumination of a cyanobacterial cell suspension and during the so-called steady state when an internal Cipool has been allowed to develop. Carbon dioxide uptake following illumination of a cyanobacterial cell suspension was progressively reduced by increasing [GLY] in the medium, both in the absence or in the presence of carbonic anhydrase. The same features were found when either Ciin the form of bicarbonate was injected to a pre-illuminated cell suspension in the presence of carbonic anhydrase in the medium or a pulse of CO2was provided to the cells in the light without carbonic anhydrase in the extracellular medium. Although photosynthesis was completely abolished with 10 mM glycolaldehyde, CO2uptake was only inhibited 20% but was decreased further by higher [GLY]. Double reciprocal plots of CO2uptake versus initial [CO2] determined with a range of [GLY] showed the inhibition to be noncompetitive. To simultaneously study CO2uptake and CO2efflux, H13C18O3was used with cells in which CO2transport alone, or both CO2and HCO3-transport system were allowed to proceed. Cells were inhibited with either iodoacetamide or glycolaldehyde and the time course of the various CO2species in the medium demonstrated that CO2uptake was inhibited partially with glycolaldehyde resulting in a greater CO2efflux from the cells. When the cell suspension was again illuminated, in the presence of glycolaldehyde, initial CO2uptake was greatly reduced, and the extracellular [CO2] eventually rose to a level higher than that which would be expected to be in equilibrium with the amount of Cipresent in the medium. Results are discussed in terms of glycolaldehyde use in Citransport mechanism studies in cyanobacteria.Key words:SynechococcusUTEX 625, blue-green algae, cyanobacteria, CO2transport, HCO3-transport, CO2efflux, inhibitor, Ciconcentrating mechanism.