1.  Temperature

Figure 1. The effect of temperature to specific growth rate, maximum biomass production, biomass productivity, yield of biomass from carbonate

M standard medium

45°C, 40°C, 35°C, 25°C, 30°C, 9 flasks,

Inoculation rate: 5%,

2.  Inoculums size

M standard medium

35°C, 1%, 5%, 10%, 3 repeats,

Figure 2, effect of inoculation size and growth curve

3.  Statistical experiment on pH, sodium carbonate, and sodium chloride

Code level / -1.68 / -1 / 0 / 1 / 1.68
sodium bicarbonate / 0.16M / 0.5 M / 1.0M / 1.5M / 1.84M
sodium chloride / 0.16M / 0.5 M / 1.0M / 1.5M / 1.84M
pH / 7.82 / 8.5 / 9.5 / 10.5 / 11.18
Number / NaCO3 / NaCl / pH
1 / -1 / -1 / -1
2 / 1 / -1 / -1
3 / -1 / 1 / -1
4 / 1 / 1 / -1
5 / -1 / -1 / 1
6 / 1 / -1 / 1
7 / -1 / 1 / 1
8 / 1 / 1 / 1
9 / -1.68 / 0 / 0
10 / 1.68 / 0 / 0
11 / 0 / -1.68 / 0
12 / 0 / 1.68 / 0
13 / 0 / 0 / -1.68
14 / 0 / 0 / 1.68
15 / 0 / 0 / 0
16 / 0 / 0 / 0
17 / 0 / 0 / 0
18 / 0 / 0 / 0
19 / 0 / 0 / 0
20 / 0 / 0 / 0

Table 1, experiment design

Table 2, parameters and levels

Table 3, equation

Table 4 verification experiment result

Figure 3. Response surface plot

6/8-6/15, experiment, shaker at 35°C

6/30-7/5, verification, shaker at 35°C

4.  With optimized pH, carbonate concentration, temperature and NaCl concentration, test different nitrogen sources

7/5 -7/12

Figure 4. Nitrogen effects to cell growth

Yeast extract, corn steep solids, peptone, tryptone,

7/12-7/19

Figure 5. Nitrate concentration effect

0.5 g/L, 1.0 g/L, 1.5 g/L, 2.0 g/L, 2.5 g/L, three repeats, 15 flasks

7/19 – 7/26

Figure 6. Ammonium concentration effect (0.5 g/L nitrate)

0 g/L, 0.1 g/L, 0.2 g/L, 0.4 g/L, 0.8 g/L, 1.6 g/L, three repeats, 18 flasks

5.  Composition analysis of the produced biomass

7/1- 7/15

Table 5. Fatty acid profile of algal biomass

Table 6. Protein profile of algal biomass

Check references

Table 7. Compositions of algal biomass

6.  Repeat culture:

7/5- 9/30

Medium made from NaHCO3 is sparged with pure CO2 until equilibrium. 6 flasks, 150 ml, Harvest at 1 week culture, compensate the water evaporation. re-inoculate 5 % seed cells, keep culturing for another week, 5 ml supernatant for Ci analysis. Use the final spent medium as absorbent for CO2 absorption; check how much CO2 can be hold, then, culture with this medium again.

Until the cell growth turn into slow, i.e., the productivity is less than 20% of maximum productivity.

Figure 7. Biomass production, Ci consumption, gain, and loss, pH change in the culture process, yield of biomass from consumed Ci

7.  Semi-continuous culture, investigate different harvesting interval time

7/5-7/30

Culture a week at first, then start semi-continuous culture

Harvest every 1 day, 2 days, 3days, and 4 days, keep 50% of cells in the original culture

Total four weeks, or until the cell stop growth

Figure 8. Biomass production, Ci consumption, gain, and loss, pH change in the culture process, yield of biomass from consumed Ci

Discussion:

CO2 loss and absorption should be considered in the culture process.

Bioresource Technology

Modeling work add result in open system as another paper