Sang-Mok Lee

Simple Synthesis of Functionalized Superparamagnetic Magnetite/Silica Core/Shell Nanoparticles and their Application as Magnetically Separable High‐Performance Biocatalysts

Uniformly sized silica-coated magnetic nanoparticles (magnetite@silica) are synthesized in a simple one-pot process using reverse micelles as nanoreactors. The core diameter of the magnetic nanoparticles is easily controlled by adjusting the w value ([polar solvent]/[surfactant]) in the reverse-micelle solution, and the thickness of the silica shell is easily controlled by varying the amount of tetraethyl orthosilicate added after the synthesis of the magnetite cores. Several grams of monodisperse magnetite@silica nanoparticles can be synthesized without going through any size-selection process. When crosslinked enzyme molecules form clusters on the surfaces of the magnetite@silica nanoparticles, the resulting hybrid composites are magnetically separable, highly active, and stable under harsh shaking conditions for more than 15 days. Conversely, covalently attached enzymes on the surface of the magnetite@silica nanoparticles are deactivated under the same conditions.

Purification of l(+)-lactic acid from fermentation broth with paper sludge as a cellulosic feedstock using weak anion exchanger Amberlite IRA-92

Lactic acid promises to be an important commodity chemical in the future as a monomer for the production of biodegradable polylactic acid. With the increase of the lactic acid demand, the need to explore alternative feedstock sources and purification processes that are inexpensive and efficient is becoming more important. This paper first reports the purification results of lactic acid from the fermentation broth with paper sludge as a cellulosic feedstock using weak anion exchanger Amberlite IRA-92. Some factors such as flow rate, sample volume loaded, pH, and column were systematically examined to improve the purity, yield and productivity in lactic acid purification. Adsorption isotherm of standard lactic acid and lactic acid in the fermentation broth by anion exchanger IRA-92 were also investigated. Results indicate that in purification process the increase of pH of the fermentation broth ranging from 5.0 to 6.0 can significantly enhance the recovery yield, purity and productivity. The decrease of flow rate and sample volume loaded can also improve the recovery yield and purity but apparently reduce the productivity. In addition, the scale-up of purification process in laboratory size has little influence on the recovery yield and purity. After optimization, the yield, purity and productivity are found to be about 82.6%, 96.2% and 1.16 g LA/(g-resin day), respectively.

Enzyme-catalyzed reactions in ionic liquids

Ionic liquids have been suggested as potential “green solvents” due to their unique properties such as non-volatility, nonflammability, and a wide temperature range for liquid phase. This review describes recent advances of biocatalyst reactions in ionic liquids. Enzyme-catalyzed reactions in ionic liquids-transesterification, synthesis, conversion, ammoniolysis, hydrolysis, epoxidation, resolution, and oxidation are presented. The use of ionic liquids for protein folding/refolding and the toxicity of ionic liquids are also discussed.

Influence of ionic liquids on the growth ofEscherichia coli

Ionic liquids are compounds that composed only of ions and are liquid at room temperature. Thus, it is normally named room temperature ionic liquid (RTIL). In this study, the application of RTILs to the extractive fermentation of biomaterials was investigated as a substitute of organic solvents. The relative toxicity of the RTILs on the growth ofE. coli was tested. The inhibition of cell growth in the presence of various ionic liquids was measured using solid and liquid culture, and EC50 of each RTILs was calculated. The number of viable and total cells was measured by the number of colonies and optical density, respectively. Effective concentrations of toxicity (EC50) in these tested systems were similar with conventional solvents, such as acetone, acetonitrile, and ethanol. The viability ofE. coli was affected by the polarity and ionic properties of ionic liquids. The resistance of the microorganisms against ionic liquids was different with the cations and anions composing ionic liquids. No general influence of the anionic compound of the ionic liquids was found on toxicity comparing with distinctive influence of cationic moiety.

Highly stable trypsin-aggregate coatings on polymer nanofibers for repeated protein digestion

A stable and robust trypsin‐based biocatalytic system was developed and demonstrated for proteomic applications. The system utilizes polymer nanofibers coated with trypsin aggregates for immobilized protease digestions. After covalently attaching an initial layer of trypsin to the polymer nanofibers, highly concentrated trypsin molecules are crosslinked to the layered trypsin by way of a glutaraldehyde treatment. This process produced a 300‐fold increase in trypsin activity compared with a conventional method for covalent trypsin immobilization, and proved to be robust in that it still maintained a high level of activity after a year of repeated recycling. This highly stable form of immobilized trypsin was resistant to autolysis, enabling repeated digestions of BSA over 40u2005days and successful peptide identification by LC‐MS/MS. This active and stable form of immobilized trypsin was successfully employed in the digestion of yeast proteome extract with high reproducibility and within shorter time than conventional protein digestion using solution phase trypsin. Finally, the immobilized trypsin was resistant to proteolysis when exposed to other enzymes (i.e., chymotrypsin), which makes it suitable for use in “real‐world” proteomic applications. Overall, the biocatalytic nanofibers with trypsin aggregate coatings proved to be an effective approach for repeated and automated protein digestion in proteomic analyses.

β-Glucosidase coating on polymer nanofibers for improved cellulosic ethanol production

Abstractβ-Glucosidase (βG) can relieve the product inhibition of cellobiose in the cellulosic ethanol production by converting cellobiose into glucose. For the potential recycled uses, βG was immobilized and stabilized in the form of enzyme coating on polymer nanofibers. The βG coating (EC-βG) was fabricated by crosslinking additional βG molecules onto covalently attached βG molecules (CA-βG) via glutaraldehyde treatment. The initial activity of EC-βG was 36 times higher than that of CA-βG. After 20xa0days of incubation under shaking, CA-βG and EC-βG retained 33 and 91% of each initial activity, respectively. Magnetic nanofibers were also used for easy recovery and recycled uses of βG coating. It is anticipated that the recycled uses of highly active and stable βG coating can improve the economics of cellulosic ethanol production so long as economical materials are employed as a host of enzyme immobilization.

Modeling of Typical Microbial Cell Growth in Batch Culture

A mathematical model was developed, based on the time dependent changes of the specific growth rate, for prediction of the typical microbial cell growth in batch cultures. This model could predict both the lag growth phase and the stationary growth phase of batch cultures, and it was tested with the batch growth ofTrichoderma reesei andLactobacillus delbrucckii.

Continuous cultivation ofLactobacillus rhamnosus with cell recycling using an acoustic cell settler

Continuous production of lactic acid from glucose byLactobacillus rhamnosus with cell recycling using an acoustic cell settler was carried out. The performance of the system, such as the concentration of cell and product were compared with the control experiment without recycling. The acoustic settler showed cell separation efficiency of 67% during the continuous operation and the cell concentration in the fermentor with recycle exceeded that of the control by 29%. Compared with the control, lactic acid production was increased by 40%, while glucose consumption was only increased by 8%. The higher value of lactic acid production to substrate consumption (YP/S, product yield coefficient) achieved by cell recycling is interpreted to indicate that the recycled cell mass consumes less substrate to produce the same amount of product than the control. Within system environmental changes due to the longer mean cell residence time induced the cells maintaining the metabolic pathways to produce less by-product but more product, lactic acid.

Modeling and simulation of lactic acid fermentation with inhibition effects of lactic acid and glucose

An unstructured mathematical model for lactic acid fermentation was developed. This model was able to predict the inhibition effects of lactic acid and glucose and was confirmed to be valid with various initial concentrations of lactic acid and glucose. Simulation of energy production was made using this mathematical model, and the relationship between the kinetics of energy metabolism and lactic acid production was also analyzed.

Lipase-catalyzed transesterification in several reaction systems: An application of room temperature lonic liquids for bi-phasic production ofn-butyl acetate

Organic solvents are widely used in biotransformation systems. There are many efforts to reduce the consumption of organic solvents because of their toxicity to the environment and human health. In recent years, several groups have started to explore novel organic solvents called room temperature ionic liquids in order to substitute conventional organic solvents. In this work, lipase-catalyzed transesterification in several uni-and bi-phasic systems was studied. Two representative hydrophobic ionic liquids based on 1-butyl-3-methylimidazolum coupled with hexafluorophosphate ([BMIM][PF6]) and bis[(trifluoromethylsulfonyl) imide] ([BMIM] [Tf2N]) were employed as reaction media for the transesterification ofn-butanol. The commercial lipase, Novozym 435, was used for the transesterification reaction with vinyl acetate as an acyl donor, The conversion yield was increased around 10% in a water/[BMIM][Tf2N], bi-phasic system compared with that in a water/hexane system. A higher distribution of substrates into the water phase is believed to enhance the conversion yield in a water/[BMIM][Tf2N] system. Partion coefficients of the substrates in the water/[BMIM][Tf2N] bi-phasic system were higher than three times that found in the water/hexane system, while n-butyl acetate showed a similar distribution in both systems. Thus, RTILs appear to be a promising substitute of organic solvents in some biotransformation systems.