MEASUREMENT OF CO2 AND HCO3- FLUXES IN CYANOBACTERIA AND MICROALGAE DURING STEADY-STATE PHOTOSYNTHESIS

A mass spectrometric procedure is described which allows the simultaneous estimation of both CO2 and HCO3- fluxes associated with cyanobacteria and green algae during steady-state photosynthesis. This technique utilizes the chemical disequilibrium which exists between CO2 and HCO3- during photosynthesis in cell suspensions which lack carbonic anhydrase activity. The kinetic equations which are derived for flux determinations are based on models of photosynthesis in both cyanobacteria and green algae which seem most reasonable given our present level of understanding, together with direct measurement of [CO2], estimation of [HCO3-] and application of the kinetic rate constants for the interconversion Of CO2 and HCO3-. From measurements made in the light, net uptake of both CO2 and HCO3- can be readily determined. In addition, analysis of the dark phase immediately following light-off provides the possibility of also determining the CO2 evolution which is occurring during photosynthesis, and thus also the gross CO2 uptake rates in the light. Results are presented for the response of dissolved inorganic carbon (C(i)) flux rates to external C(i) in low-C(i) grown cells of both Synechococcus PCC7942 and Chlamydomonas reinhardtii and these are consistent with previous studies showing that such cells possess capacities to utilize both CO2 and HCO3- for photosynthesis. The advantages and potential errors which are inherent in this technique are discussed together with its potential for future studies on C(i) transport under various experimental conditions.