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Mitsuyasu Okabe,Tiejun Liu,Shigenobu Miura,Tomohiro Arimura,Min-Yi Tei,Enoch Y. Park 한국생물공학회 2005 Biotechnology and Bioprocess Engineering Vol.10 No.6
Various processes which produce L-lactic acid using ammonia-tolerant mutant strain, Rhizopus sp. MK-96-1196, in a 3 L airlift bioreactor were evaluated. When the fed-batch culture was carried out by keeping the glucose concentration at 30 g/L, more than 140 g/L of L-lactic acid was produced with a product yield of 83%. In the case of the batch culture with 200 g/L of initial glucose concentration, 121 g/L of L-lactic acid was obtained but the low product yield based on the amount of glucose consumed. In the case of a continuous culture, 1.5 g/L/h of the volumetric productivity with a product yield of 71% was achieved at dilution rate of 0.024 h-1. Basis on these results three processes were evaluated by simple variable cost estimation including carbon source, steam, and waste treatment costs. The total variable costs of the fed-batch and continuous cultures were 88% and 140%, respectively, compared to that of batch culture. The fed-batch culture with high L-lactic acid concentration and high product yield decreased variable costs, and was the best-suited for the industrial production of L-lactic acid.
Cellulose Production from Gluconobacter oxydans TQ-B2
최두복,차월석,Shiru Jia,KiAn Cho,Mitsuyasu Okabe,Hongyu Ou,Guibing Chen 한국생물공학회 2004 Biotechnology and Bioprocess Engineering Vol.9 No.3
Gluconobacter oxydans that produces the cellulose was isolated. In order to confirm the chemical features of cellulose, various spectrophtometeric analysis were carried out using electron microscopy, X-ray diffractogram, and CP/MAS 13C NMR. The purified cellulose was found to be identical to that of Acetobacter xylinum. For effective production of cellulose, the various carbon and nitrogen sources, mixture of calcium and magnesium ions, and biotin concentration were investigated in flask cultures. Among the various carbon sources, glucose and sucrose were found to be best for the production of cellulose, with maximum concentration of 2.41 g/L obtained when a mixture of 10 g/L of each glucose and sucrose were used. With regard to the nitrogen sources, when 20 g/L of yeast extract was used, the maximum concentration of bacterial cellulose was reached. The concentration of cellulose was increased with mixture of 2 mM of each Ca2+ and Mg2+. The optimum biotin concentration for the production of cellulose was in the range of 15 to 20 mg/L. At higher biotin concentration (25~35 mg/L), the bacterial cellulose production was lower.
Liu, Tiejun,Miura, Shigenobu,Arimura, Tomohiro,Tei, Min-Yi,Park, Enoch Y.,Okabe, Mitsuyasu The Korean Society for Biotechnology and Bioengine 2005 Biotechnology and Bioprocess Engineering Vol.10 No.6
Various processes which produce L-lactic acid using ammonia-tolerant mutant strain, Rhizopus sp. MK-96-1196, in a 3L airlift bioreactor were evaluated. When the fed-batch culture was carried out by keeping the glucose concentration at 30g/L, more than 140 g/L of L-lactic acid was produced with a product yield of 83%. In the case of the batch culture with 200g/L of initial glucose concentration, 121g/L of L-lactic acid was obtained but the low product yield based on the amount of glucose consumed. In the case of a continuous culture, 1.5g/L/h of the volumetric productivity with a product yield of 71% was achieved at dilution rate of $0.024\;h^{-1}$. Basis on these results three processes were evaluated by simple variable cost estimation including carbon source, steam, and waste treatment costs. The total variable costs of the fed-batch and continuous cultures were 88% and 140%, respectively, compared to that of batch culture. The fed-batch culture with high L-lactic acid concentration and high product yield decreased variable costs, and was the best-suited for the industrial production of L-lactic acid.
Expression of Mouse $\alpha-Amylase$ Gene in Methylotrophic Yeast Pichia pastoris
Uehara Hiroyuki,Choi Du Bok,Park Enoch Y.,Okabe Mitsuyasu The Korean Society for Biotechnology and Bioengine 2000 Biotechnology and Bioprocess Engineering Vol.5 No.1
The expression of the mouse $\alpha-amylase$ gene in the methylotrophic yeast, P pastoris was investigated. The mouse $\alpha-amylase$ gene was inserted into the multi-cloning site of a Pichi a expression vector, pPIC9, yielding a new expression vector pME624. The plasmid pME624 was digested with SalI or BglII, and was introduced into P. pastoris strain GSl15 by the PEG1000 method. Fifty-three transformants were obtained by the transplacement of pME624 digested with SaiII or BglII into the HIS4locus $(38\;of\;Mut^+\;clone)$ or into the AOX1 locus $(15\;of\;Mut^s\;clone)$. Southern blot was carried out in 11 transformants, which showed that the mouse $\alpha-amylase$ gene was integrated into the Pichia chromosome. When the second screening was performed in shaker culture, transformant G2 showed the highest $\alpha-amylase$ activity, 290 units/ml after 3-day culture, among 53 transformants. When this expression level of the mouse $\alpha-amylase$ gene is compared with that in recombinant Saccharomyces cerevisiae harboring a plasmid encoding the same mouse $\alpha-amylase$ gene, the specific enzyme activity is eight fold higher than that of the recombinant S. cerevisiae.
Naoyuki Moriya,Yukiko Moriya,Hideo Nomura,Kisato Kusano,Yukoh Asada,Hirofumi Uchiyama,Enoch Y. Park,Mitsuyasu Okabe 한국생물공학회 2013 Biotechnology and Bioprocess Engineering Vol.18 No.6
β-(1→3)-D-glucans with β-(1→6)-glycosidiclinked branches are known to be immune activation agentsand are incorporated in anti-cancer drugs and healthpromotingsupplements. β-Glucan concentration was 9.2 g/Lin a 200-L pilot scale fermentor using mutant strainAureobasidium pullulans M-2 from an imperfect fungalstrain belonging to A. pullulans M-1. The culture broth ofA. pullulans M-2 had a faint yellow color, whereas that ofthe wild-type had an intense dark green color caused by theaccumulation of melanin-like pigments. β-Glucan producedby A. pullulans M-2 was identified as a polysaccharide ofD-glucose monomers linked by β-(1→3, 1→6)-glycosidicbonds through GC/MS and NMR analysis. When aconventional medium was used in the culture of A. pullulansM-2 in a 3-L jar fermentor, β-glucan concentration was1.4-fold that produced by the wild-type. However, when amedium optimized by statistical experimental design wasused with dissolved oxygen at 10%, the β-glucan concentrationwas 9.9 g/L with a yield of 0.52 (g β-glucan/g consumedsucrose), 2.9-fold that of the wild-type. This level ofproductivity was reproduced when the fermentation wasscaled up 200-L. The industrial production of high β-glucan without melanin-like pigments is highly expected,as a health-promoting supplement or functional food.