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REFERENCE LINKING PLATFORM OF KOREA S&T JOURNALS
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Journal DOI :
Korean Society for Biotechnology and Bioengineering
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Volume & Issues
Volume 11, Issue 6 - Dec 1996
Volume 11, Issue 5 - Oct 1996
Volume 11, Issue 4 - Aug 1996
Volume 11, Issue 3 - Jun 1996
Volume 11, Issue 2 - Apr 1996
Volume 11, Issue 1 - Feb 1996
Selecting the target year
Detection of flavonoid compounds by cell culture of Ginkgo biloba L
KSBB Journal, volume 11, issue 1, 1996, Pages 1~7
Calli induced from Ginkgo bilha L. were cultured to investigate optimal culture conditions and identify the possibility production of useful compounds. Calli were obtained from leaves and stems of Ginkgo biloba seedlings and embryos on WP medium supplemented with 2mg/
NAA and 5mg/
kinetin. Chlorophyll-ricked green callus was inducted in MS liquid medium containing 1mg/
NAA and 0.1mg/
kinetin under light as 3 clones selected with origin. Embryo derived callus showed the highest growth rate. Analysis for flavonoids and their precursor was performed by TLC and EMS. A specific precursor of flavonoid was identified in callus, not in natural leaves. These findings indicate that tissue culture may produce rlavonoids.
The Overexpression of Subtilisin Enzyme Using Mutations on Transition State Regulatory Proteins of AprE Promoter and Development of Bacillus subtilis Host System
KSBB Journal, volume 11, issue 1, 1996, Pages 8~14
Bacillus subtillis strains with transition state regulator mutations and a spore mutation were developed for the overexpression of apsE and for the enhancement of expression level. Among the many regulator genes, degU and hpr were chosen as a representative positive and negative regulator for the aprE, respectively. Spo II G was used for the construction of asporogeneous strains. All the mutants were constructed from two protease-deleted strain DB104 and the apsE gene was transformed with an integration vector pMK101. DB104(deg
)::pMK101(Cm) and DB104(
her(Em))::pMKl01(Cm) show 7-fold and about 2-fold increase in aprE expression level, respectively. But the effect of transition state regulator mutation on the aprE expression was diminished when the integrated aprE gene was amplified by the high concentration of chloramphenicol, i. e. 30
spoIIG(Pm) degUh(32) his+)::pMK101(Cm) and DB104(
hpr(Em))::pMK101 double mutant show 10-fold and 3-fold increase in aprE expression level, respectively. The results suggest that sporulation mutation and transition state regulator mutation have independent and additive effect on the aprE expression, and the same gene dosage effect on the transition state regulator mutation was also identified.
Purification of Biosynthesized Hyaluronic Acid for Its Medical Application
KSBB Journal, volume 11, issue 1, 1996, Pages 15~21
Purification of hyaluronic acid produced by Streptococcus equi was carried out to obtain clinical grade hyaluronic acid. The removal method of the bacteria was selected as filtration because filtration was the most effective method in removing impurities such as protein and nucleic acid of the fermentation broth. The removal efficiencies of protein and nucleic acid of hyaluronic acid solution were increased to 75% and 67%, respectively, by filtration with adding 0.6% of activatied carbon and 1.0% colite. Hyaluronic acid solution was precipitated by mixing with 2 volumes of ethanol. Effects of pH and conductivity on ethanol preciptation of hyaluronic acid were investigated. Protein and nucleic acid of hyaluronic acid were remained almost constant regardless of pH and conductivity, and the recovery of hyaluronic acid was optimum as about 85% at pH 7 and l00mS of conductivity Protein of hyaluronic acid was completly removed by three serial filtration and ethanol precipitation, however, nucleic acid was not removed. Hyaluronic acid solution was passed through a column of Duolite A7 to remove its nucleic acid, where 65% of nucleic acid was removed at pH 7 and 40mS of conductivity. The residual nucleic acid of hyaluronic acid solution was completly removed by treatment of 0.2% hydroxyapatite and the clinical grade hylauronic acid could be obtained.
Studies on Mass Production of Intracellularly-Produced Secondary Metabolite, Cyclosporin A by Use of Immobilized Fungal Cells in Stirred-Tank Immobilized Perfusion Reactor System(IPRS)
KSBB Journal, volume 11, issue 1, 1996, Pages 22~29
Immobilized bioprocess was carried out for continuous production of cyclosporin A (CyA) produced intracellularly as a secondary metabolite by a filamentous fungus, Tolypocladium inflatum. Immobilization procedure for entrapping conidiospores of the producer was significantly simplified by use of a modified immobilization technique. A newly-designed immobilized perfusion reactor system (IPRS) showed good process benefits as demonstrated by the role of the high density immobilized cells as an efficient biomass generator, continuously supplying highly active CyA-producing free cells (1.0g/
/hr) even at very high dilution rate (
). IPRS bioprocess was possible since efficient decantor system developed in our laboratory separated the sloughed-off free cells from the immobilized biomass effectively, thus overcoming wash-out phenomenon frequently encountered in continuous free cell cultures. Furthermore the released-free cells remaining in the bulk solution did not appear to cause substrate mass transfer limitation which was often experienced in suspended mycelial fungal cell fermentations. The primary reason for this was that the suspension broth of the IPRS mainly consisted of roundshaped short mycelial fragments and conidiospores, still remaining Newtonian even at high cell density. In parallel with IPRS bioprocess development, other key factors to be considered necessarily for significant increase in CyA productivity would be strain improvement and medium optimization for the immobilized cells.
Determination of Optimum Bead Size by Calculating Effectiveness Factors in Cyclosporin A Fermentation by Immobilized Cells
KSBB Journal, volume 11, issue 1, 1996, Pages 30~36
Based on fermentation data for cyclosporin A production, simple Monod kinetics was proposed for both immobilized and suspended cultures. Higher value of
mas and lower value of Km suggest better catalytic activity of the immobilized cells than the parallel suspended cells. Furthermore, lower Km value in the immobilized cell system indicates higher affinity of the immobilized cells for carbon substrate as compared with the suspended cells. For immobilized cell cultures, these parameters were also utilized for the estimation of effectiveness factor, an indicator for intraparticle mass transfer resistance. Based on simulation studies, optimum radius of celite beads was turned out
In this simulation work, we examined the influence of biosupport size and immobilized biomass density on diffusional resistance of substrate inside the bead matrix. In order to maintain uniformly distributed cell activities in biosupport, it was essential to determine optimum slze of particle, and then to estimate the most economic loaded biomass content.
Purification of Extracellular Agarase from Marine Bacterium (Pseudosmonas sp. W7) and Molecular Cloning of the Agarase Gene
KSBB Journal, volume 11, issue 1, 1996, Pages 37~45
Marine bacterial strain, highly effective agar degrading, was isolated from south sea of Korea and was identified as Pseudomonas sp. This strain was named Halophilic Pseudomonas sp. W7 and accumulated an extracellular agarase which showed a high level of enzyme activity in the presence of agar and agarose. This extracellular agarase was purified by anion-exchange chromatography and gel filtration. Purified agarase showed a single protein band upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis and its molecular weight was estimated to be about 89KDa. The agarase gene was cloned into Escherichia coli JM83 using the plasmid vector pUC19. DNA fragments(3.7, 3.0Kb) of Hind III-digested chromosomal DNA of Pseudomonas sp. W7 was inserted into the Hind III site of pUC19. Selected transformants, E. coli JM83/pSWl 000000and E. coli JM83/pSW3, produced agarase and this agarase was accumulated In the cytoplasmic space.
Continuous Foam Separation of Yeast Cells
KSBB Journal, volume 11, issue 1, 1996, Pages 46~51
Cell separation by means of continuous foam separation of Saccharomyes formosensis without additive was investigated. The yeast separation ratio was improved at low feed rates, high nitrogen rates, optimum pH and temperature for ethanol production, dilution of cultivation medium and addition of 0.5g/
. Percentage of yeast removal and yeast separation ratio were more than 85 when continuous foam separation was operated in optimum condition..
Effect of Shear Stress on Bovine Aortic Smooth Muscle Cell Growth
KSBB Journal, volume 11, issue 1, 1996, Pages 52~57
Bovine aortic smooth muscle cells cultured on the slide glass were exposed to sheared flow up to 120 hours in flow chamber to see the effect of shear stress on cell growth in wall shear stresses of 0 to 26dyn/
. From lactate dehydrogenase concentration measurement of the circulating medium, it was shown that sheared flow in the shear stress range did not remove additional smooth muscle cells from the slide glass compared with cells in stationary condition. According to smooth muscle cell counting per
of the surface, smooth muscle cells grew fastest in the stationary condition. As the wall shear stress increased, the growth of cells became slower. When the wall shear stress increased over 17dyn/
, cell growth was not observed throughout the experiment
Continuous Production of Sorbitol with Zymomonas mobilis in a Packed Bed Reactor
KSBB Journal, volume 11, issue 1, 1996, Pages 58~64
The purpose of this study is to develop a continuous process for sorbitol production using Zymomonas mobilis immobilized in K-carra-geenan. The glutaraldehyde cross-linking of toluene-treated cells immobilized in alginate or chitin showed high enzyme stability for long period. However, loss of enzyme activity was observed at 23% during 210h. In order to investigate the stability of glucose-fructose oxidoreductase of cethyltrimethylammoniumbromide (CT AB) treated cells, the long term continuous process was carried out with Z. mobilis immobilized in K-carrageenan in the continuous stirred tank reactor(CSTR) and the packed bed reactor. The continuous production of sorbitol with the immobilized CT AB permeabilized cells in packed bed reactor was more stable than in CSTR. Two stage continuous process with CT AB treated cells of Z. mobilis immobilized in K-carrageenan was carried out at various dilution rates. At the first stage, the productivity was increased up to 15 g/
-h as dilution rate increased and decreased over O.32
of dilution rate. Similarly, maximum productivity obtained at the second stage was 22g/
-h at O.32
The Production of Sodium Gluconate by Aspergillus niger
KSBB Journal, volume 11, issue 1, 1996, Pages 65~70
Sodium gluconate was produced by neutralization of gluconic acid formed during the submerged culture fermentation of glucose with Aspergillus niger ACM 7. The fermentation characteristics of Aspergillus niger ACM 7 were investigated quantitatively according to the change of the initial glucose concentrations and the initial pHs of fermentation broth. The maximum specific growth rate was estimated to be
of initial glucose concentration. The maximum fermentability of sodium gluconate was 95% at the initial glucose concentration of 26g/
. However, the maximum sodium gluconate productivity was 1.18g/
/hr when the initial glucose concentration was 110g/
. The optimum pH was found to be 5.5 for both the cell growth and the sodium gluconate production. With optimized culture conditions, the productivity of sodium gluconate in a fed-batch culture(production fermentor, 16,000
) increased up to 7.1g/
Studies on the supercritical fluid extraction of taxol from yew tree
KSBB Journal, volume 11, issue 1, 1996, Pages 71~76
Studies were carried out to examine some factors affecting the supercritical carbon dioxide extraction of taxol from the bark of Taxus cuspidata using a continuous packed bed extractor. The factors investigated in this study were pressure, temperature, volume of carbon dioxide, and co-solvent. It was found out that the amount of taxol extracted was not significantly affected by the operating pressure in the absence of a co-solvent although it increased by about 20% at 5500 psig. With
of carbon dioxide the saturated amount of taxol was extracted at 318K and 5500psig. Methanol was found to be the most effective co-solvent in terms of amount of taxol extracted among six different co-solvents used. When methanol was used as a co-solvent the effect of operating pressure became significant; approximately 50% increase in the amount of taxol extracted was observed at 3000 psig as compared to at 2500 or 3500psig. The optimum methanol concentration in supercritical fluid was 13% (w/w)at 308K, 3000psig.
Development of Alginate-Celite Immobilization Technique for the Improvement of Ethanol Productivity
KSBB Journal, volume 11, issue 1, 1996, Pages 77~85
The optimal initial pH for the ethanol production by Saccharomyces K35 was found to be 5.0, and about 80% of yield was obtained when 200g/
of glucose was used as a substrate, which showed sugar tolerant. As the additives and cross-linking agent, the addition of 1.67%(w/v) Celite R-634 together with 0.33%(v/v) of glutaraldehyde(ACG bead) resulted in better stability, ethanol productivity and cell viability than Ca-alginate bead. Also, ACG bead seemed to be more resistant to phosphate ion than Ca-alginate bead, considering outgrowing cell concentration in the media. Scanning electron microscopic observation depicted that the surface of ACG bead was almost similar to the original state but not for Ca-alginate bead. When repealpd-batch culture was performed with Ca-alginate bead for 60 days in a 500m1 Erlenmeyer flask, ethanol and cell concentration were maintained about 138g/
-gel and 29~30g/
-gel, respectively, up to 40 days(7th run number), and then both were rapidly decreased. In the case of ACG bead, ethanol and cell concentration were maintained about 130~150g/
-gel and 32~35g/
-gel, respectively, up to 60days(10th run number). Cell viability was maintained about 70%, and outgrowing cell concentration was below 5.8% of total cell concentration.
Measurement for Determining the Biodegradation of Starch-Filled Polyethylene Film by
KSBB Journal, volume 11, issue 1, 1996, Pages 86~91
Optimal reaction condition for the starch hydrolysis by
-amylase was determined and the sugar produced under the optimal condition was measured for estimating the biodegradation of strach-filled polyethylene film. Optimal ranges of temperature and pH were 70~
and 6.3~7.3, respectively. The 100 units of
-amylase per mg starch were enough for the enzyme reaction. Reaction with polyethylene film containing 5%, 10%, 15% and 20% starch in the above condition showed that the sugar produced was proportional to the starch content in film. This relationship provides a calibration curve for determining the starch content of search-filled polyethylene film. The average amount of hydrolyzed starch was about 40% of total starch in film. The rest of the starch is considered to be still dispersed in the film and not to be attacked by
-amylase. In this experiment, we could obtain the higher biodegradability through the
-amylase reaction in the above optimal condition than the reported one which had been Improved by adding surfactant.
Isolation and Characterization of Serratia sp. JM Producing Chitinase
KSBB Journal, volume 11, issue 1, 1996, Pages 92~98
A chitinase-producing bacterium was isolated from seashore mud around Beobseongpo in Chunnam province by selective enrichment culture, and among it, one isolate which was the best in producing of chitinase was selected. Nutrient or MacConkey medium was confirmed with secreting of prodigiosin pigment by Serratia sp. JM, and it was performed by the production of clear zone on medium containing chitin. Serratia sp. JM was almost same compared with Serratia marcescens ATCC 27117 in respect of its morphological, physiological and biochemical characteristics except succinic, urea and pyruvic acid. Serratia sp. JM was resistant to tetracycline but was not resistant to kanamycin and chloramphenicol. The optimal temperature and pH for the production of chitinase from Serratia sp. JM were
and 7.5, respectively. Production of chitinase and pH in the medium increased until the cultivation of 120 hours, but after 120 hours, they were decreased due to the acetic acid accumulated from degradation of chitin by Serratia sp. JM.
Extraction of Taxol and Baccatin III from Needles of Taxus Cuspidata by Using Supercritical Carbon Dioxide with Cosolvents
KSBB Journal, volume 11, issue 1, 1996, Pages 100~106
The extraction of taxol and baccatin III from the ground needles of Taxus cuspidata were carried out by using supercritical carbon dioxide with and without cosolvents such as ethylacetate and methanol at 300 bar and 313K. Taxol is a promising anticancer agent and baccatin III is a precursor of semisysthesis of taxol. The taxol and baccatin III contents in the extracts were determined by a HPLC. The highest yields of taxol and baccatin III could be obtained by the supercritical extraction with 3wt% methanol and ethylacetate, respectively, as cosolvents. It was also found that the selectivity of taxol and baccatin III were 0.117 and 1.245wt%, respectively, with 0.7wt% ethylacetate, which demonstrated that the selectivity of taxol and baccatin III were increased about 1.8 and 19 times than those of conventional organic solvent extraction.
Extracellular compounds can enhance development of carrot somatic embryos
KSBB Journal, volume 11, issue 1, 1996, Pages 107~114
The enhancing effect of excreted cell factors on the production of somatic embryos from suspension cultures of Daucus carota was studied as a function of factors including molecular size, harvesting time, injection period, and concentration of the extracellular compounds. The production of late-stage embryos was increased up to 1, 500 embryos/ml compared with control cultures when high molecular size and extracellular factors, extracted from newly established embryo culture at early stationary phase, were added at the starting time. The stimulating effect on the production of somatic embryos can be attributed to the presence of high molecular size(> 10 kDa) compounds.
Zoolan Gene Cloning of Zoogloea ramigera 115
KSBB Journal, volume 11, issue 1, 1996, Pages 115~123
Two kinds of mutants were isolated to clone a cluster of genes essential for zooglan biosynthesis. Zoogloea ramigera 115 strains produce capsular polysaccharide. To achieve conjugation in strain 115 and to facilitate recovery of product, a capsule non-forming strain was isolated via successive centrifugation and screening. The other kind of mutants devoid of or producing altered exopolysaccharides were obtained using classical transposon(Tn5) technique and screened for altered colony morphology and celluflour binding properties. Complementation of these mutants was achieved with Z. ramigera 115 slime gene library constructed in a broad host range cosmid vector and helper plasmid by triparental conjugation.