• KSBB Journal
• /
• v.14 no.3
• /
• pp.365-370
• /
• 1999

To optimize the culture conditions for Beauveria bassiana 726, the effects of culture medium, pH, and temperature on mycelium and spore production were investigated. The optimum temperature and pH for the cultivation of B. bassiana 726 were 28 $^{\circ}C$ and 5.0, respectively. The optimized medium was composed of 1.0~2.0% total sugar from molasses, 0.5% corn steep liquor and 0.05% KH$_2$PO$_4$. In the cultivation of B. bassiana 726 with the optimum medium, the specific growth rate and substrate utilization were well-fitted with the proposed kinetic model in the shake flask and stirred tank reactor. When the fed-batch cultivation using carbon suorce, nitrogen source, and mineral salt as a feeding medium was compared with batch cultivation in stirred tank reactor, mycelium (12.7 g/L) and spore production (5.4$\times$$10^8/mL$) were enhanced up to 110% and 85%, respectively.

• KSBB Journal
• /
• v.5 no.3
• /
• pp.269-277
• /
• 1990

Methylobacillus sp. SK1, an obligate methylotroph, was cultivated in a fed-batch culture using DO as a methanol feeding indicator with a micro computer-aided control system. While 2.07g/l of cell density was obtained after 13 hr in the batch culture (initial methanol concentration: 1.0%(v/v)),45.3g/l of cell density was obtained after 17 hr by feeding methanol and metal ions in the fed-batch culture with oxygen supply. The high-density biomass was obtained in short cultuivation time by fed-batch culture with feedback control, and consequently the biomass productivity was significantly increased. It was mainly due to extension of logarithmic growth period by methanol feeding without methanol inhibition and intensive aeration without DO limitation with microcomputer-aided control system.

• Journal of Microbiology and Biotechnology
• /
• v.14 no.2
• /
• pp.225-230
• /
• 2004

A plasminogen kringle domain 1 to 3, rKl-3, was expressed in Escherichia coli under the control of T7 promoter. For the cost-effective production of rKl-3, the induction process was analyzed and optimized. Induction characteristics with lactose were analyzed in terms of induction time and inducer concentration in various culture conditions including batch and high-cell-density fed-batch cultures. In the fed-batch culture, the induction around 6 h after initiation of the DO-stat fed-batch culture resulted in the highest expression level of rKI-3 among the induction points examined. The highest demand of oxygen at this point was crucial for the maximum expression level of rKI-3. As the lactose concentration increased, the expression level also increased, though the expression level showed a plateau above a concentration of 14 mM of lactose. Lactose acted less specifically than IPTG since most of it was hydrolyzed to glucose and galactose. However, using lactose, the cell growth and the maximum expression level of rKl-3 increased by 20% and 24%, respectively, compared with those using IPTG in the fed-batch culture. The lactose seemed to be hydrolyzed by intracellular and extracellular $\beta$-galactosidase liberated by cell lysis at the same time. Residual concentration of glucose was maintained to a a limit of detection by high performance liquid chromatography, and galactose was not consumed by the host strain Escherichia coli BL2l(DE3).

• Biotechnology and Bioprocess Engineering:BBE
• /
• v.9 no.3
• /
• pp.184-190
• /
• 2004

The influences of impeller types on morphology and protein expression were investigated in a submerged culture of Aspergillus oryzae. The impeller types strongly affected mycelial morphology and protein production in batch and fed-batch fermentations. Cells that were cultured by propeller agitation grew in the form of a pellet, whereas cells that were cultured by turbine agitation grew in a freely dispersed-hyphal manner and in a clumped form. Pellet-grown cells showed high levels of protein production for both the intracellularly heterologous protein (${\beta}$-glucuronidase) and the extracellularly homologous protein (${\alpha}$-amylase). The feeding mode of the carbon source also influenced the morphological distribution and protein expression in fed-batch fermentation of A. oryzae. Pulsed-feeding mainly showed high protein expression and homogeneous distribution of pellet whereas continuous feeding resulted in less protein expression and heterogeneous distribution with pellet and dispersed-hyphae. The pellet growth with propeller agitation paralleling with the pulsed-feeding of carbon source showed a high level of protein production in the submerged fed-batch fermentation of recombinant A. oryzae.

• Journal of Microbiology and Biotechnology
• /
• v.9 no.5
• /
• pp.548-553
• /
• 1999

A production of $\beta-Carotene$was attempted in a fed-batch culture of Blakeslea trispora by controlling both foam and dissolved oxygen in the presence of surfactant, Span 20. Results obtained from the shake flask cultures indicated that a high concentration of dissolved oxygen was needed for both cell growth and $\beta-Carotene$ synthesis, and the optimal concentration of glucose was found to be in the range of 50-100 g/l. In order to maintain the dissolved oxygen concentration level at higher than 50% of air saturation, pure oxygen was automatically sparged into the medium with air. Foam was controlled by bypassing air from the submerged aeration to the headspace in response to the foam that was caused by Span 20. High agitation speed was found to be detrimental to the cell growth due to shear damage, even though it provided sufficient dissolved oxygen. On the other hand, a low aeration speed caused stagnant regions in the fermentor because of improper mixing. Thus, for the fed-batch operation, agitation speed was increased gradually from 300 to 700 rpm to prevent cell damage at the initial stage of fermentation and to give efficient mixing for a viscous culture broth as the culture proceeded. By controlling dissolved oxygen and foam, a high concentration of $\beta-Carotene$otene (1,190 mg/l) was obtained in 6 days of the fed-batch culture of B. trispora with 2.5% of the dry cell weight, which was approximately 5 times higher than that of the batch cultures.

• Journal of Microbiology and Biotechnology
• /
• v.25 no.3
• /
• pp.358-365
• /
• 2015

ε-Poly-L-lysine (ε-PL) is a homopolymer of L-lysine molecules connected between the epsilon amino and alpha carboxyl groups. This polymer is currently used as a natural preservative in food. Insufficient biomass is a major problem in ε-PL fermentation. Here, to improve cell growth and ε-PL productivity, various nitrogen-rich nutrients were supplemented into flask cultures after 16 h cultivation, marking the onset of ε-PL biosynthesis. Yeast extract, soybean powder, corn powder, and beef extract significantly improved cell growth. In terms of ε-PL productivity, yeast extract at 0.5% (w/v) gave the maximum yield (2.24 g/l), 115.4% higher than the control (1.04 g/l), followed by soybean powder (1.86 g/l) at 1% (w/v) and corn powder (1.72 g/l) at 1% (w/v). However, supplementation with beef extract inhibited ε-PL production. The optimal time for supplementation for all nutrients examined was at 16 h cultivation. The kinetics of yeast-extract-supplemented cultures showed enhanced cell growth and production duration. Thus, the most commonly used two-stage pH control fed-batch fermentation method was modified by omitting the pH 5.0-controlled period, and coupling the procedure with nutrient feeding in the pH 3.9-controlled phase. Using this process, by continuously feeding 0.5 g/h of yeast extract, soybean powder, or corn powder into cultures in a 30 L fermenter, the final ε-PL titer reached 28.2 g/l, 23.7 g/l, and 21.4 g/l, respectively, 91.8%, 61.2%, and 45.6% higher than that of the control (14.7 g/l). This describes a promising option for the mass production of ε-PL.

• Journal of Microbiology and Biotechnology
• /
• v.9 no.6
• /
• pp.716-721
• /
• 1999

It has been known that continuous cultivation of hairy root is difficult to maintain for a long period of time compared to the microbial and callus cultures. Chemostat cultivation was successfully carried out in order to economically produce a plant-based colorant, betacyanin, from red beet hairy root for more than 85 days in a 14-1 fermentor. The result from the chemostat cultivation was compared to those of the batch and fed-batch cultivations of red beet hairy roots. It was shown that hairy root reached its steady state within 50 days of the cultivation, and then maintained for about 25-30 days in a wide range of dilution rates. Total betacyanin production from the continuous process was also calculated to be 2.65g at 0.28(l/d) of dilution rate, compared to 0.196g from fed-batch cultivation. It was found that betacyanin production was a partially growth related process, yielding 0.376 mg/g-fresh wt. cell and $1.89{\times}10^{-5}$ mg/g-fresh wt. cell/d, with 0.92 of correlation factor in a partial growth-product model. It was also shown that the cell growth required was relatively large for maintenance amount of energy at a low dilution rate. The growth of hairy root was inhibited by high light intensity in following a photo-inhibition model. The growth parameters were estimated to be 0.3(l/d), $10.56kcal/\textrm{m}^2/h$,{\;}and{\;}35.81kcal/\textrm{m}^2/h$ for the maximum specific growth rate, half saturation light intensity, and inhibition light intensity, respectively.

• KSBB Journal
• /
• v.15 no.2
• /
• pp.174-180
• /
• 2000

Pichia pastoris and Hansenula polymorpha, the methylotrophic yeasts have been widely used as a host for the production of e eudaryotic proteins due to the advantages related to their inherited characters. This paper describes the method to enhance t the productivity of recombinant proteins by P. pastoris and H. po$\psi$morpha. In the production of recombinant proteins using a f fed batch fermentation system, the effects of specific growth rate on the specific expression rate of re$\infty$mbinant proteins w were studied. In both species, the expression system of recombinant proteins using the fed batch fermentation was optimezed.

• Journal of Microbiology and Biotechnology
• /
• v.10 no.3
• /
• pp.321-326
• /
• 2000

To examine the feasibility of the long-term production of the human granulocyte colony stimulating factor (hG-CSF) using the $_{L}-arabinose$ promoter system of Escherichia coli, flask relay culture and cyclic fed-batch culture were performed. In the flask relay culture, it was found that the pismid was maintained stably up to about 170 generations in an uninduced condition, whereby the cells could also maintain the capability of expressing hG-CSF expression were maintained stably up to at least 100 generations. In contrast, in the cyclid fed-batch culture, segregational plasmid instability was observed within about 4 generations after induction, even though the cell growth and hG-CSF production reached their maximum balues, 78.0 g/l of dry cell weight and 7.0 g/l of hG-CSF, respectively. It would appear that, when compared to the flask relay culture, the high-cell density and high-level expression of hG-CSF in the cyclic fed-batch cultrure led to the segregational plasmid instability; in other words, a severe metabolic burden existe on the cells due to the high-level expression of hG-CSF. Accordingly, based on these long-term cultures, the segregational and structural plasmid instability was observed and a strategy to overcome such problems could be designed.

• KSBB Journal
• /
• v.15 no.2
• /
• pp.181-187
• /
• 2000

The production of heterologous protein using GAL promoter in conventional S. cerevisiae has several problems to s이ve for c commercialization. In this research, S. cerevisiae mutant(reg1-501, gaI1), which cannot use galactose and has alleviated g glucose repression level, is used as host for optimizing induction of GAL promoter. In this experiment, the effects of specific g growth rate on specific recombinant protein expression rate were tested in both cases and optimum fed batch fermentation m method was obtained in both cases. Through these experiments, optimum condition of recombinant protein production by G GAL promoter using S. cerevisiae mutant (reg1-501, gal1) were found.