Cyanobacteria culture in bicarbonate system to optimize biomineralization

The calcification phenomenon has been extensively studied for calcifying microorganisms, and seems to be dependant on seawater composition in marine environment (Stanley, 2006; Stanley et al., 2005) as well as on the morphology and physiology of these calcifying microorganisms (Lowenstam and Weiner, 1989). In modern cyanobacteria calcification, Merz (1992) and Merz-Preiß and Riding (1999) suggested two calcification styles, i.e. impregnation and encrustation, responding to environmental factors, and also restricting calcification to certain taxa, cyanbacteria could be distinguished as « calcifying and no calcifying strains ». According to Lee et al. (2006), species of cyanobacteria Synechococcus are known to be responsible for CaCO3 precipitation in the presence of calcium via the carbonate formation in the culture medium due to an alkaline environment development during cell growth. Different mechanisms have been proposed and classified into biologically induced and biologically controlled. Moreover, the situation is probably more complex, Obst et al., (2009) suggested a passive surface-induced mechanism for CaCO3 nucleation by cyanobacteria, the cells membrane contributed to reduce the specific interfacial free energy of the calcite nuclei. The main accepted fact is that cyanobacterial calcification is clearly related to photosynthetic carbon uptake (Altermann et al., 2006; Riding, 2006). The inference of a relationship between CO2 concentrating mechanisms (CCM)-stimulated photosynthesis and calcification has been rapidly suggested. In cyanobacteria, whatever the mineral carbon source entering the cell, only bicarbonate is present in the cytosol. The CO2 entering the cell is converted to HCO3

(Price et al., 2008; Jansson and Northen, 2010). Under these conditions, the CO2 fixation tends probably to be globally pH-neutral. Consequently, the high CO2 transfer will tend to lower the pH at the cell-surface microenvironment generating unfavourable conditions to calcification. Moreover microalgae growth (Chlorella for example) inhibition has been reported with 10% and 15% CO2 concentrations in the flue gas stream (Chiu et al., 2008). In these conditions with 2 to 15 % CO2 (v/v) in gaz flux, the average pH was 7.6 to 6.8. More surprisingly, weak Ci fixation into biomass (2% of the total CO2 sequestered) by cyanobacteria Aphanothece microscopica Nägeli using 15% CO2 air has been reported (Jacob-Lopes et al., 2009), suggesting the existence of other routes of CO2 conversion. Developments are needed to evaluate more precisely the potentiality of cyanobacteria to capture CO2 (via photosynthesis and CaCO3 precipitation) and to optimize the process with CO2 rich flue gas as main carbon source. In this study, we cultivated Synechococcus sp. PCC 8806 in an open-system with atmospheric CO2 as carbon source (limiting CO2 supply condition as in major natural environments). Non-limiting CO2 supply condition was also operated through a pH regulation, using the carbonate system (bicarbonate/carbonate) to capture CO2 (0.003 v/v in air) injected. In these two approaches, very low calcium concentration was used to examine the photosynthetic response to the culture conditions with no CaCO3 precipitation interference.

The marine strain of cyanobacteria Synechococcus sp. PCC 8806 was used for the present study and was maintained on ASN-III growth medium (Rippka et al., 1979) with atmospheric CO2 as carbon source. The culture was transferred to fresh medium with low CaCl2 concentration (0.38 mM) every 4 weeks. All cultures were grown at 25°C in an Incubator/Shaker (Snijders, ECD01E) under an intermittent light regime (12/12h light/dark A detailed observation of the pH evolution during the dark and light periods is presented in the Figure 2. During the dark phase, the photosynthesis ceased. The CO2 entry in the carbonate system was responsible for the decrease of pH observed over the 12 hours of night period and was estimated by the evolution of hydroxyl, bicarbonate and carbonate concentrations (see Table 1) according to equations 2 and 3. CO2 supply before and after 70 days have been respectively estimated to 42 and 90 μmol of CO2 over a period of 12 h.