It was previously revealed that the total CO2 concentration in seawater decreased during the Late Archean. In this paper, to assess the secular change of total CO2 concentration in seawater, we focused on the Paleoproterozoic era when the Earth experienced its first recorded global glaciation. The 2.4 Ga Ongeluk Formation outcrops in the Kaapvaal Craton, South Africa. The formation consists mainly of submarine volcanic rocks that have erupted during the global glaciation. The undeformed lavas are mostly carbonate-free but contain rare disseminated calcites. The carbon isotope ratio of the disseminated calcite (delta(13)Ccc vs. VPDB) ranges from -31.9 to -13.2 %. The relatively low delta(13)Ccc values clearly indicate that the carbonation was partially contributed by C-13-depleted CO2 derived from decomposition of organic matter beneath the seafloor. The absence of delta(13)Ccc higher than -13.2% is consistent with the exceptionally C-13-depleted CO2 in the Ongeluk seawater during glaciation. The results suggest that carbonation occurred during subseafloor hydrothermal circulation just after the eruption of the lavas. Previously, it was reported that the carbonate content in the uppermost subseafloor crust decreased from 3.2 to 2.6 Ga, indicating a decrease in total CO2 concentration in seawater during that time. However, the average CO2 (as carbonate) content in the Ongeluk lavas (<0.001 wt%) is much lower than those of 2.6 Ga representatives and even of modern equivalents. This finding suggests that the total CO2 concentration in seawater further decreased during the period between 2.6 and 2.4 Ga. Thus, the very low content of carbonate in the Ongeluk lavas is probable evidence for the extremely low CO2 concentration in seawater during the global glaciation. Considering that the carbonate content of the subseafloor crusts also shows a good correlation with independently estimated atmospheric pCO(2) levels through the Earth history, it seem highly likely that the low carbonate content in the Ongeluk lavas reflects the low atmospheric pCO(2) at that time. We conclude that the continuous decrease in CO2 concentration of seawater/atm. from 3.2 Ga was one of the contributing factors to the Paleoproterozoic global glaciation.