Direct measurement of pCO(2) in cultures of marine phytoplankton: how good is the estimate from pH(NBS) and single point titration of alkalinity?

In physiological studies of marine phytoplankton in culture, the CO2 equilibrium is typically calculated from measurement of PHNBS (calibrated in dilute National Bureau of Standards buffers) and a single point titration of alkalinity (A(T)) This approach has widespread appeal because it is simple, inexpensive, and requires very low sample volumes. However, its continued application ignores advances in analytical and theoretical oceanic CO2 chemistry that suggest fundamental flaws in the assumption PHNBS = -log[H+] as a consequence of the high ionic strength of seawater and liquid residual junction errors in the electrode. Here we compare directly measured pCO(2) in sterile artificial seawater with calculated values, adopting the usual assumptions and commonly used equilibrium constants. Calculated values either overestimated (ca +15 to +23 % error) or underestimated (-9 to -11% error) directly measured pCO(2) depending upon whether constants were derived, respectively, on the PHNBS Or 'seawater' (PHSWS) scales. With the currently accepted equilibrium constants on the 'total hydrogen ion' pH scale (PHTOT), and converting PHNBS to PHSWS using an apparent activity coefficient f(H) (optimum value 0.85) and then to PHTOT, excellent agreement was achieved between calculated and measured pCO(2), both in sterile seawater and in cultures of the diatom Thalassiosira pseudonana. However, f(H) is generally unknown and is specific to electrode and electrode condition, making calculated pCO(2) a rather nebulous concept. Without knowledge of f(H), calculated pCO(2) had a total uncertainty of an order (similar to 120 ppmv at atmospheric equilibrium 360 ppmv) similar to the variation in atmospheric pCO(2) between glacial periods and the present (similar to 160 ppmv). This method therefore clearly lacks the resolution required to address the biogeochemical significance of key physiological questions. Future studies should measure pCO(2) directly, and then if required calculate CO2(aq) from pCO(2) and the solubility coefficient. Alternatively, since analytical precision of calculated pCO(2) was excellent, and accuracy is potentially good when f(H) is known, we advocate improved interdisciplinary collaboration in order to improve this pH & AT approach as a simple but effective tool for the study of marine phytoplankton physiology under controlled conditions.