Bong Hyun Chung

Concentration of tocopherols from soybean sludge by supercritical carbon dioxide

A supercritical fluid extraction method has been applied to test the feasibility of tocopherol concentration from soybean sludge with carbon dioxide at temperatures and pressures ranging from 35 to 70°C and 200 to 400 bar, respectively. The supercritical solubility of the esterified soybean sludge was over 4–6 times greater than that of the original soybean sludge. By a simple batch-type one-stage method the tocopherols in the esterified soybean sludge could be concentrated up to 40 wt%. The overall results of the present study show that soybean sludge initially containing about 13–14 wt% tocopherols may require a countercurrent multistage column to be highly and effectively concentrated.

High pressure three-phase equilibria for the carbon dioxide-ethanol-water system

Abstract The three-phase equilibria for the carbon dioxide-ethanol-water system were measured at 313.2 K and pressures of 8.14, 8.21 and 8.27 MPa. The experimental data were correlated by using the Patel-Teja equation of state with the mixing rule of Wilson.

Glucose-limited fed-batch culture of Escherichia coli for production of recombinant human interleukin-2 with the DO-stat method

Abstract Fed-batch culture employing the DO-stat method was carried out to produce recombinant human interleukin-2 in Escherichia coli . The concentration of interleukin-2 and final cell density in the fed-batch culture were 1.02 g/ l and 60 OD (optical density), respectively. This represents a significant improvement in interleukin-2 production and final cell density over the equivalent values of batach fermentation.

Role of Junctin Protein Interactions in Cellular Dynamics of Calsequestrin Polymer upon Calcium Perturbation

Background: In vitro studies have reported reversible calsequestrin polymerization and depolymerization. Results: Live cell imaging analysis revealed Ca2+-dependent decondensation of calsequestrin speckles, consistent with in vitro microscopic data. Conclusion: Calsequestrin depolymerization by calcium depletion requires coexistence of junctin. Significance: The role of calsequestrin in intracellular calcium homeostasis was explored. Calsequestrin (CSQ), the major intrasarcoplasmic reticulum calcium storage protein, undergoes dynamic polymerization and depolymerization in a Ca2+-dependent manner. However, no direct evidence of CSQ depolymerization in vivo with physiological relevance has been obtained. In the present study, live cell imaging analysis facilitated characterization of the in vivo dynamics of the macromolecular CSQ structure. CSQ2 appeared as speckles in the presence of normal sarcoplasmic reticulum (SR) Ca2+ that were decondensed upon Ca2+ depletion. Moreover, CSQ2 decondensation occurred only in the stoichiometric presence of junctin (JNT). When expressed alone, CSQ2 speckles remained unchanged, even after Ca2+ depletion. FRET analysis revealed constant interactions between CSQ2 and JNT, regardless of the SR Ca2+ concentration, implying that JNT is an essential component of the CSQ scaffold. In vitro solubility assay, electron microscopy, and atomic force microscopy studies using purified recombinant proteins confirmed Ca2+ and JNT-dependent disassembly of the CSQ2 polymer. Accordingly, we conclude that reversible polymerization and depolymerization of CSQ are critical in SR Ca2+ homeostasis.

Screening of a specific monoclonal antibody against and detection ofListeria monocytogenes whole cells using a surface plasmon resonance biosensor

In this study, a specific monoclonal antibody againstListeria monocytogenes was screened using an SPR biosensor Monoclonal antibodies were bound to protein L, after which theL. monocytogenes cells were subjected to an affinity assay. Protein L was immobilized on a carboxymethyl dextran (CM-Dex) surface via an amine coupling method and utilized repeatedly by regeneration. The monoclonal antibody, ‘A18’, was selected and employed for the high-sensitivity detection ofL. monocytogenes. Under optimized conditions, 103 cells/ml or 50 cells were detected by the SPR biosensor.

A Highly Sensitive Enzyme-Amplified Immunosensor Based on a Nanoporous Niobium Oxide (Nb2O5) Electrode

We report on the development of an enzyme-amplified sandwich-type immunosensor based on a thin gold film sputtered on an anodic nanoporous niobium oxide (Au@Nb2O5) electrode. The electrocatalytic activity of enzymatically amplified electroactive species and a stable electrode consisting of Au@Nb2O5 were used to obtain a powerful signal amplification of the electrochemical immunobiosensor. The method using this electrochemical biosensor based on an Au@Nb2O5 electrode provides a much better performance than those based on conventional bulk gold or niobium oxide electrodes. Our novel approach does not require any time-consuming cleaning steps to yield reproducible electrochemical signals. In addition, the strong adhesion of gold films on the niobium oxide electrodes offers a very stable substrate during electrochemical biosensing. Cyclic voltammetry measurements indicate that non-specific binding of proteins to the modified Au@Nb2O5 surface is sufficiently low to be ignored in the case of our novel system. Finally, we demonstrated the ability of the biosensor based on an Au@Nb2O5 offering the enhanced performance with a high resolution and sensitivity. Therefore, it is expected that the biosensor based on an Au@Nb2O5 has great potential for highly efficient biological devices.

Fermentation Strategy to Enhance Plasmid Stability during the Cultivation of Escherichia coli for the Production of Recombinant Levansucrase

Abstract To produce levansucrase, a fructosyltransferase enzyme, in a recombinant Escherichia coli harboring the levU gene of Zymomonas mobilis, fermentation strategies were examined in terms of induction methods and plasmid stability. Although the recombinant levansucrase was induced rapidly by IPTG, high instability of the plasmid and formation of inclusion bodies were observed. Segregational instability was aggravated by the excretion of β-lactamase into the culture broth during subculturing, which caused an overgrowth of plasmidfree cells. A combination of methicillin (2, 6-dimethoxyphenyl-penicillin) and ampicillin to provide selective pressure was effective in preventing the growth of plasmid-free cells. The population of plasmid-harboring cells was maintained above 95% of the total cells for more than 100 generations under this condition. In order to replace IPTG, which is toxic and too expensive for use in a large scale, d -lactose was tested and found to be favorable as an alternative inducer.

Production of recombinant human interleukin-z by Escherichia coli with computer controlled temperature induction

The amount of alkali added to the fermentation broth of a recombinant Escherichia coli strain for pH control was monitored on-line by an electronic balance interfaced to a computer. It was successfully correlated with the cell mass, and consequently the cell growth could be well estimated. Using this cell growth estimation technique, an automatic temperature induction was successfully carried out to produce human interleukin-2 in a high yield from E. coli harbouring a temperature controlled expression vector system.

Hollow Fiber Bioreactors with Internal Aeration Circuits

Abstract New hollow fiber bioreactors for aerobic culture were introduced and Aspergillus niger for citric acid production was cultivated as a model system. These reactors consisted of a bundle mixed of hydrophilic membranes for liquid nutrient transport and hydrophobic membranes for gaseous nutrient transport. The cells were successfully cultivated. However, the polymeric hollow fiber membranes were compressed and blocked by excessive fungal cell growth. Citric acid was produced with a high volumetric productivity compared with that of shake-flask fermentation, but the long-term operation was not successful due to a rapid decrease of the production rate.

Correlation of redox potential with state variables in cultures under controlled dissolved oxygen concentration and pH

In batch cultures for l‐ornithine production in which dissolved oxygen concentration and pH were closely controlled, time changes of redox potential were observed in connection with the profiles of cell, glucose, and ornithine concentrations. It was found that the redox potential profile had four different phases reflecting the physiological state of the culture and that it was closely related to cell concentration change. Effects of glucose and ornithine on the redox potential were identified in a separate series of experiments. On the basis of the experimental results, a correlation of redox potential to glucose, cell, and ornithine concentrations has been proposed. The proposed correlation can be used for on‐line estimation of ornithine concentration from on‐line data of redox potential, glucose concentration, and cell concentration.