Won Jae Choi

Kinetic resolution for optically active epoxides by microbial enantioselective hydrolysis

Resolution of several racemic epoxides was accomplished using an epoxide hydrolase activity of whole cells of the newly isolated Aspergillus niger. (S)-Styrene oxide, for example, was obtained from its racemates with optical purity of 100% ee and 32% yield.

Enhanced production of cis,cis-muconate in a cell-recycle bioreactor

For enhanced production of cis,cis-muconate via biocatalytic conversion from benzoate, a high cell density culture of a mutant Pseudomonas putida strain, BM014 in a cell-recycle bioreactor has been investigated. The volumetric productivity in a continuous bioconversion process was dependent on the biomass concentration, specific growth rate, and environmental conditions including dissolved oxygen tension (DOT) and cis,cis-muconate concentration. Biotransformation was started in a cell-recycle continuous culture mode for obtaining a high cell density of 40 g/l and then carried out in a fed-batch mode for accumulation of cis,cis-muconate to up to 12 g/l at which induction of catechol 1,2-dioxygenase, a key enzyme catalyzing the rate-determining step in the biosynthetic pathway, was maximized. Since in the biosynthesis of cis,cis-muconate growing cells were utilized as catalysts, cell growth should be continued, but a low specific growth rate achieved under phosphate-limited conditions was indispensable for higher productivity. When the concentration of phosphate in the feed was lowered from 150 to 10 mM, the productivity could be enhanced three fold. It was also found that cobalt ions markedly increased the specific productivity and that DOT should be kept at above 20% of the saturation level for achieving optimum rates of cell growth and product formation. After a period of fed-batch operation continuous production with cell recycling was attempted with phosphate-limited minimal medium supplemented with cobalt ions, and benzoate medium. Enhanced productivity of 5.5 g/(l·h) and a cis,cis-muconate concentration of 13.5 g/l in the culture broth were obtained and continuous production in a cell-recycle bioreactor was maintained for more than 7 d.

Biocatalytic production of chiral epichlorohydrin in organic solvents.

Enantioselective hydrolysis of racemic epichlorohydrin was accomplished for the production of enantiopure epichlorohydrin using the whole cells of an isolated Aspergillus niger spps. To overcome the spontaneous chemical degradation of epichlorohydrin that occurs in aqueous buffer, organic solvents were employed in the reaction medium. The enantioselectivity was highly dependent on the solvent structure, water content of the medium, and the initial epichlorohydrin concentration. (S)-epichlorohydrin could be obtained from its racemates (60 mM) with an optical purity of 100% enantiomeric excess (ee) and 20% yield in cyclohexane supplemented with 2.0% (v/v) water.

Production of optically active ketoprofen by direct enzymatic esterification.

For the production of optically active ketoprofen, enzymatic resolution of racemic ketoprofen in an organic solvent has been accomplished via enantioselective esterification. Pharmacologically inactive (R)-ketoprofen is converted into the corresponding (R)-ester by this method. Enantioselectivity in lipase-catalyzed resolution of racemic ketoprofen was mainly dependent on the sources of lipase, alcohol moiety, organic solvent, and water content. Ethanol was used as the alkyl donor and the optimum water content required for highly efficient enzymatic resolution was determined to be 0.1-0.15% (v/v), which was maintained using salt hydrates such as Na2SO4 x 10H2O. (S)-Ketoprofen could be obtained with high enantioselectivity (E=15) in n-hexane supplemented with ethylene dichloride (20% (v/v)) using commercially available Candida antarctica lipase (Novozym 435).

Two-Stage Reconstruction of Bilateral Alveolar Cleft Using Y-Shaped Anterior-Based Tongue Flap and Iliac Bone Graft

OBJECTIVE When an alveolar cleft is too large to close with adjacent mucobuccal flaps or large secondary fistula following a primary bilateral palatoplasty exists, a one-stage procedure for bone grafting becomes challenging. In such a case, we have used the tongue flap to repair the fistula and cleft alveolus followed by bone grafting to the cleft defect performed several months later. The purpose of this article is to report on our experiences with the use of an anteriorly based Y-shaped tongue flap to fit the palatal and labial alveolar defects and on the ultimate result of the bone graft. PATIENTS A series of 14 patients were treated with this approach from January 1994 to December 1998. The average age of the patients was 15.8 years (range 5 to 28 years). The mean period of follow-up following the second stage bone graft operation was 45.9 months (range 9 to 68 months). In 9 of the 14 patients, the long-fork type of a Y-shaped tongue flap was used for extended coverage of the labial-side alveolar defects with the palatal fistula; in the remaining patients, the short-forked design was used. RESULTS All patients demonstrated a good clinical result after the initial repair of cleft alveolus and palatal fistula. There was no fistula recurrence, although partial necrosis of distal margin in long-forked tongue flap occurred in one patient. Furthermore, the bone graft, which was performed an average of 8 months after the tongue flap repair, was always successful. Occasionally, transferred tongue tissue bulging interfered with the hygienic care of nearby teeth; however, these problems could be solved with proper contour-plasty performed afterward. No donor site complications such as sensory disturbance, change in taste, limitations in tongue movement, normal speech impairments, or tongue disfigurement were encountered. CONCLUSION This two-stage reconstruction of a bilateral cleft alveolus using a Y-shaped tongue flap and iliac bone graft was very successful. It may be indicated for a bilateral cleft alveolus patient in which the direct closure of the cleft defect with adjacent tissue or the buccal flap is not easy because of scarred fibrotic mucosa and accompanied residual palatal fistula.

Thermo-responsive copolymers with ionic group as novel draw solutes for forward osmosis processes

Thermo-responsive copolymers containing ionic groups (P(MTxEOy), where x and y are the feed molar percent of [2-(methacryloyloxy)ethyl]trimethylammonium chloride (MTAC) and 2-(2-methoxyethoxy)ethyl methacrylate (MEO), respectively) were synthesized via free radical polymerization using MTAC and MEO as monomers, in order to use the copolymers as a draw solute in a forward osmosis (FO) system. The osmotic pressure and lower critical solution temperature (LCST) could be controlled by changing the composition of the copolymers. For example, P(MT25EO75) shows a relatively high osmotic pressure of 0.909 Osmol/kg at 0.2 g/mL, and P(MT5EO95) shows a LCST close to room temperature at 32 °C. When P(MT20EO80) was used as a draw solute in the FO system, a reasonable water permeation flux value of 5.45 Lm−2h−1 at 0.1 g/mL was observed and the draw solute (P(MT20EO80)) could be recovered with a very large draw solute recovery value of 99.80% from the draw solutions by heating to 70 °C, above the LCST, followed by a microfiltration process. Therefore, it is expected that such copolymer system could be a prospective candidate as a draw solute in the FO system.

Production ofcis,cis-muconic acid from benzoic acidvia microbial transformation

For the production ofcis,cis-muconic acidvia biocatalytic conversion reactions from a toxic cosubstrate, benzoic acid, a fed-batch process using computer-controlled DO-stat feeding was developed. The mutant strain ofPseudomonas putida BM014 producedcis,cis-muconic acid from benzoic acid with high conversion yield. More than 32 g/L ofcis,cis-muconic acid was accumulated in 42h and a productivity of 1.4 g/(L·h) was achieved.

Enhancement ofcis,cis-muconate productivity by overexpression of catechol 1,2-dioxygenase inPseudomonas putida BCM114

For enhancement ofcis,cis-muconate productivity from benzoate, catechol 1,2-dioxygenase (C12O) which catalyzes the rate-limiting step (catechol conversion tocis,cis-muconate) was cloned and expressed in recombinantPseudomonas putida BCM114. At higher benzoate concentrations (more than 15 mM),cis,cis-muconate productivity gradually decreased and unconverted catechol was accumulated up to 10 mM in the case of wildtypeP. putida BM014, whereascis,cis-muconate productivity continuously increased and catechol was completely transformed tocis,cis-muconate forP. putida BCM114. Specific C12O activity ofP. putida BCM114 was about three times higher than that ofP. putida BM014, and productivity was enhanced more than two times.

Poly(1-oxotrimethylene) fibers prepared by different draw ratios for the tire cord application

AbstractPoly(1-oxotrimethylene) (POTE) was prepared by polymerization of ethylene and carbon monoxide gases in an autoclave using a palladium catalyst system. POTE fibers having different draw ratios were prepared and characterized using differential scanning calorimetry (DSC), X-ray diffraction, and a universal testing machine (UTM). Tensile strength and modulus of POTE fibers having a draw ratio of 16 obtained by drying at 220 °C were found to be 13.4 and 369.8 g/denier, respectively. These values are much larger than those of poly(ethylene terephthalate) (PET) and Nylon 6 fibers that are currently used as tire cords.

Effect of dissolved oxygen concentration on the metabolism of glucose inPseudomonas putida BM014

The effect of dissolved oxygen concentration on the metabolism of glucose inPseudomonas putida BM014 was investigated. Glucose was completely converted to 2-ketogluconatevia extracellular oxdative pathway and then taken up for cell growth under the condition of sufficient dissolved oxygen concentration. On the other hand, oxygen limitation below dissolved oxygen tension (DOT) value of 20% of air saturation caused the shift of glucose metabolism from the extracellular oxidative pathway to the intracellular phosphorylative pathway. Specific activities of hexokinase and gluconate kinase in intracellular phosphorylation pathway decreased as the DOT increased, while 2-ketogluconokinase activity in extracellular oxidative pathway increased under the same condition. This result can be usefully applied to microbial transformation of glucose to 2-ketogluconate, the synthetic precursor for iso-vitamine C, with almost 100% yieldvia extracellular oxidation by simple DOT control.