Growth parameters of Chlorella sp. MTF-7 aerated with flue gas

In our previous study (Chiu et al., 2008), microalgal cell growth was significantly

inhibited when wild-type microalgal Chlorella sp. cultures were aerated with gas containing a high concentration of CO2 (> 10% CO2). Given the high concentration of CO2 in flue gas (about 20-25% CO2), the growth potential of the isolated microalga, Chlorella sp. MTF-7, when aerated directly with flue gas was first evaluated. In indoor culture experiments, batch cultures of Chlorella sp. MTF-7 were incubated for 6 days at 25 ± 1°C under continuous cool white fluorescent light. The light intensity was approximately 300 mol m^-2 s^-1 at the surface of the photobioreactor. The flue gas generated from coke oven of a steel plant was collected in a gas storage bag, and the gas was continuously introduced into the photobioreactor by an air blower.

Figure 13 shows the growth curves of Chlorella sp. (wild-type, WT) and Chlorella sp.

MTF-7 aerated with flue gas or CO2-enriched gas (2, 10, or 25% CO2 aeration) for 6 days.

The growth potential of Chlorella sp. MTF-7 was significantly higher than that of Chlorella sp. WT when aerated with flue gas or CO₂. The maximum biomass concentrations in

Chlorella sp. MTF-7 cultures aerated with 2, 10 or 25% CO₂ were 1.67, 1.50 and 1.32 g L^-1, respectively. The maximum biomass concentration was 2.40 g L^-1 in the batch cultures of Chlorella sp. MTF-7 aerated with flue gas. The average growth rates of the Chlorella sp.

MTF-7 cultures aerated with flue gas or 2, 10 or 25% CO2 were 0.37, 0.25, 0.15 and 0.19 g L^-1 d^-1, respectively. The growth rates of the Chlorella sp. MTF-7 cultures aerated with flue gas were approximately 1.5-, 2.5- and 2.0-fold higher than those of the Chlorella sp. WT cultures aerated with 2, 10 or 25% CO2, respectively. These results indicated that Chlorella sp.

MTF-7 could be cultured with flue gas aeration; the maximum biomass productivity was 0.64 g L^-1 d^-1 in the batch culture aerated with flue gas. The growth potential of Chlorella sp.

MTF-7 cultures aerated with flue gas from the coke oven of a steel plant, which contained approximately 25% CO2, 4% O2, 80 ppm NO and 90 ppm SO2, was higher than that of the cultures aerated with 2, 10 or 25% CO2-enriched gas without pH control. The high growth capacity of microalgae aerated with flue gas has been reported previously (Douskova et al., 2009). The volumetric concentration of CO2 provided to the control culture was the same as the average concentration in the flue gas (11%). However, the growth rate of C. vulgaris cultures aerated by flue gas from an incinerator was 48% higher than that of the control culture. The high concentration of CO₂ in flue gas was a major factor in microalgal growth (Yoo et al., 2009). In our previous study, a high initial density of Chlorella sp. could

overcome the environmental stress induced by high CO₂ aeration and grow rapidly (Chiu et al., 2008). In this experiment, an initial high-density culture was used, and a gas-switching cycle operation was also introduced. In a high-density culture, the growth inhibition caused by the high CO2 concentration in the flue gas is reduced, and the pH value of the culture can be stably maintained.

The NOX present in flue gas inhibits microalgal growth (Lee et al., 2000). However, the toxic effect of NO can also be overcome by high-density cultures, and NO can be a nitrogen source for microalgal cultures. NO absorbed in the medium can be converted to NO2–

and then oxidized to NO3

-, which can be utilized as a nitrogen source (Nagase et al.-, 2001).

Gaseous NO can dissolve in the broth of microalgal cultures and can be taken up directly by algal cells through diffusion (Nagase et al., 2001). The flue gas, which contains CO2 and NO, could provide not only a carbon source for microalgal growth but also an additional nitrogen source.

SOX in flue gas is also an inhibitor of microalgal growth (Lee et al., 2000). The main form of SOX in the flue gas generated from a coke oven is SO2. Lee et al. (2000) reported that the growth of Chlorella KR-1 aerated with simulated flue gas containing 150 ppm SO₂ was suppressed because of cellular toxicity when a low-density initial biomass concentration (0.1 g L^-1) was used, but Chlorella KR-1 exhibited good growth when a high-density initial

biomass concentration (0.5 g L^-1) was used. The toxic effect of SO2 could be overcome by acidophilic microalgal isolation (Kurano et al., 1995; Lee et al., 2002). Hauck et al. (1996) reported that an acidophilic microalga, Cyanidium caladrium, grew well in the presence of 200 ppm SO2 in simulated flue gas aeration. Considering that growth of most algal strains reported was completely inhibited, when the cultures aerated with flue gas which contained SO2 concentration higher than 50 ppm (Kurano et al., 1995; Kauck et al., 1996). In our study, the isolated mutant strain, Chlorella sp. MTF-7, showed remarkably excellent tolerances to SO2 and grew well in cultures supplied with gas containing approximately 90 ppm SO2 when an initial biomass concentration of at least 0.5 g L^-1 Chlorella sp. MTF-7 was used.

The satisfactory growth of Chlorella sp. MTF-7 in cultures supplied with gas containing approximately 90 ppm SO2 may be due to its ability to tolerate highly oxidative molecular species. Bisulfite (HSO₃^-) and sulfite (SO₃^2-) are microalgal growth inhibitors that are formed in water from SO₂ (Yang et al., 2004). As SO₂ dissolves in the culture broth, HSO₃^- is formed: HSO₃^- can be converted to SO₃^2- and SO₄^2- at appropriate pH values. As HSO₃^- is converted to SO₄^2-, highly oxidative molecular species are formed, such as superoxide anions, hydrogen peroxide and hydroxyl radical. These highly oxidative molecular species can cause the peroxidation of membrane lipids and the bleaching of chlorophyll; thus, microalgal

growth is inhibited by the processing of HSO₃^- to SO₄^2- (Ranieri et al., 1999; Noji et al., 2001).

The inhibitory effect of SO2 on Chlorella sp. MTF-7 growth might be eliminated by screening specific mutant strains and using a high concentration of inoculum.

To assess the potential of Chlorella sp. MTF-7 to be cultured by the side of the stack of a coke oven for the on-site bioremediation of flue gas without a cooling system, the growth of Chlorella sp. WT and Chlorella sp. MTF-7 when aerated with flue gas at different culture temperatures was also evaluated. Figure 14 shows the growth curves of Chlorella sp. WT and Chlorella sp. MTF-7 when aerated with flue gas at 25, 30, 35 or 40ºC. The average growth rates of the Chlorella sp. WT cultures that were aerated with flue gas at 25, 30, 35 or 40ºC were 0.23, 0.21, 0.14 and 0.11 g L^-1 d^-1, respectively. The average growth rates of the

Chlorella sp. MTF-7 cultures that were aerated with flue gas at 25, 30, 35 or 40ºCwere 0.37, 0.39, 0.32 and 0.24 g L^-1 d^-1, respectively. The optimal growth temperature for Chlorella sp.

MTF-7 was 30ºC, and the maximum biomass productivity of Chlorella sp. MTF-7 cultured at 30ºC and aerated with flue gas was 0.70 g L^-1 d^-1. However, the growth rate and biomass productivity of Chlorella sp. MTF-7 that was cultured at higher temperatures (35 and 40°C) remained high and were significantly greater than those of Chlorella sp. WT, even when the

wild-type microalgal cells cultured at 25 and 30ºC.

7.2.1.2. Outdoor culture experiments

To evaluate microalgal growth performance during on-site flue gas aeration, a Chlorella sp. MTF-7 culture system was installed next to the smokestack of a coke oven at the China Steel Corporation in southern Taiwan (Figure 15). The flue gas from the coke oven was

introduced into the microalgal cultures by suction pump, and air was supplied by an air pump.

The gas was provided with either continuous flue gas aeration or intermittent flue gas aeration controlled by a gas-switching cycle operation (Figure 16). For continuous flue gas aeration, the flue gas was supplied continuously for 9 h during the day. For intermittent flue gas aeration, the flue gas was supplied in 30-min intervals every hour from 07:30 to 16:30; a gas-switching cycle was performed with a flue gas inlet load for 30 min followed by an air inlet load for 30 min (30 min flue gas/30 min air) for 9 h during the day.

Figure 17 shows the growth curves that resulted when different initial biomass concentrations (0.5, 0.75, 1.0and 1.25 g L^-1) of the Chlorella sp. MTF-7 inoculum were aerated with continuous (Figure 17A) and intermittent flue gas (Figure 17B) at 0.05 vvm.

The growth profiles of Chlorella sp. MTF-7 aerated with flue gas were stable and linear with respect to the initial biomass concentration of the inoculum, whether the flue gas supply was continuous or intermittent. The average growth rates of Chlorella sp. MTF-7 when the initial biomass inoculum was 0.5, 0.75, 1.0 or 1.25 g L^-1 were 0.13, 0.11, 0.11 and 0.05 g L^-1 d^-1 with continuous flue gas aeration, and 0.30, 0.36, 0.29 and 0.28 g L^-1 d^-1 with intermittent flue gas aeration, respectively. The growth rates of the cultures aerated with intermittent flue gas were 2.3-, 3.1-, 2.6- and 5.2-fold higher than those of the cultures aerated with continuous flue gas when initial biomass concentrations of 0.5, 0.75, 1.0 or 1.25 g L^-1 were used, respectively.

During a 6-day cultivation in which the initial biomass concentration of Chlorella sp. MTF-7 was 0.75 g L^-1, the maximum biomass growth rate was 0.52 g L^-1 d^-1, and the average biomass growth rate was 0.36 g L^-1 d^-1. The growth potential of Chlorella sp. MTF-7 cultures aerated with intermittent flue gas was significantly higher than that of Chlorella sp. MTF-7 cultures continuously aerated with flue gas. The intermittent flue gas aeration strategy for cultivation could enhance microalgal growth and also increase the utilization of the CO2 in the flue gas.

These results demonstrate that Chlorella sp. MTF-7 can grow well in an outdoor photobioreactor aerated directly with flue gas from a coke oven.

7.2.2. Flue gas bioremediation by continuous flue gas aeration