| Summary: | 碩士 === 國立中興大學 === 食品科學系 === 93 === Previous study regarding determination of heterotrophic cultivation conditions in shaking flasks for a useful algal strain, Chlorella pyrenoidosa NCHU-6, was continued. Medium optimization using experimental designs was performed further in this study. Moreover, the optimum medium obtained was used for subsequent semicontinuous cultivation of the alga to improve further the algal growth and cellular chlorophyll content by using the table-top fermentor without illumination.
In the preliminary tests of the response surface methodology (RSM),
the original medium was stepwise examined. Minerals or salts such as A5 solution, KH2PO4, MgSO4．7H2O, FeSO4．7H2O and EDTA-Na2 were found to have almost no influence on the algal growth expressed as packed cell volume (PCV), and chlorophyll production. The optimal glucose concentration for the algal production was 8 %, which led to the highest PCV (130 ml/L) and chlorophyll production (50.68 mg/L). The urea concentration for peak PCV (110 ml/L) was 10 g/L, and 12 g/L for chlorophyll production (67.84 mg/L). The optimal potassium nitrate concentrations for PCV and chlorophyll production were 10 g/L and 15 g/L, respectively. The optimal initial pH was 9.0.
The optimal medium composition for algal growth (PCV) of C. pyrenoidosa NCHU-6 was thereafter determined by RSM as follows (per liter): glucose, 60.26 g; KNO3, 13 g; Urea, 10.17 g; MgSO4.7H2O, 1.5 g; KH2PO4, 1.5 g; FeSO4.7H2O, 0.12 g；EDTA-Na2,0.12 g; A5 solution,6ml; deionized water; initial pH︰8.7. The maximal PCV obtained was 110 ml/L after 7 days of cultivation. Concentrations of chlorophyll, crude proteins, and Chlorella growth factor (CGF) were 63.75 mg/L, 66.27%, and 1.35 A260/mg, respectively.
Similarly, the optimum medium composition for chlorophyll production of C. pyrenoidosa NCHU-6 was also determined by RSM as follows (per liter): glucose, 63.94 g; KNO3, 15.13 g; Urea, 10.24 g; MgSO4.7H2O, 1.5 g; KH2PO4, 1.5 g; FeSO4.7H2O, 0.12 g; EDTA-Na2, 0.12 g; A5 solution, 6 ml; deionized water; initial pH︰8.9. The maximal chlorophyll content was 70.28 mg/L after 7 days of cultivation. The PCV was 90.5 ml/L, and contents of crude protein and CGF were 65.43% and 1.36 A260/mg, respectively.
Since we had determined that 60%(v/v) culture broth and replacing with fresh medium in the middle of cultivation was good for the semi-continuous cultivation of C. pyrenoidosa NCHU-6 in a 7L-fermentor in the previous study . Taking 40% (v/v) of remaining culture broth as starter, and fed the medium which was optimal for chlorophyll produciton at day 3 or day 4. The highest PCV (70 mg/L) was obtained at day 10 if feeding was performed at day 4, which the peak chlorophyll production, 169.32 mg/L, reached at day 9 if feeding was done at day 3. Algal cells harvested from that of feeding at day4 had the highest crude protein content.
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