BIOREACTOR SYSTEM DESIGNS FOR LIPASE-CATALYZED SYNTHESIS OF SACCHARIDE- FATTY ACID ESTERS IN SOLVENT-FREE MEDIA

As nontoxic biobased surfactants derived from plant oils and cellulose or starch, saccharide-fatty acid esters are widely used in cosmetics, food, and pharmaceutical industries due to their biocompatibility, biodegradability as well as antimicrobial activity. Generally, saccharide-fatty acid esters are synthesized chemically under high pressure, temperature and the presence of alkaline or acid catalysts leading to low-quality products (chemo-degradation of double bonds and oxygenated moieties) and large amounts of byproducts. In contrast, biocatalytic synthesis enhances sustainability: near-ambient pressure and temperature, the absence of toxic, acids and bases catalysts, and improved selectivity of products. For lipase-catalyzed synthesis under nearly anhydrous conditions, the major hurdle to be overcome is the poor miscibility of the acyl donor and acceptor substrates, resulting in slow reaction rates. Although several approaches such as, the employments of organic solvents, complexation agents, and ionic liquids, have been reported in the literature, a robust solution is desperately needed. This study focused on employing immobilized lipases under completely solvent-free conditions to synthesize saccharide-fatty acid esters using the ester products to enhance miscibility. Experimentally, metastable saccharide particles with a diameter of 10-100 micron-sized suspensions of saccharide were formed in oleic acid-rich ester mixtures initially for synthesis of saccharide-fatty acid esters in packed bed bioreactor containing immobilized lipases. Water, a by-product that limits ester yield by promoting hydrolysis, was removed via free evaporation. In this dissertation, a bioreactor system was developed for the eco-friendly solvent-free, immobilized lipase-catalyzed synthesis of biobasaed surfactants utilizing suspensions as reaction medium with 88 wt% in 6 days; the performance of the bioreactor systems developed for Objective 1 was optimized through water concentration control and interval time with 91 wt% in 4.8 days; and to improve design of bioreactor system developed in Objective 1 by in-line filter and derive a mathematical model to describe the production of esters by the bioreactor systems developed. Finally, 84 wt% ester content was achieved in 8.4 days

Mass Spectrometric Characterization and Fluorophore-Assisted Light Inactivation of Human Excitatory Amino Acid Transporter

Glia-expressing excitatory amino acid transporter 2 (EAAT2) mediates the bulk of glutamate re-uptake in the human central nervous system (CNS) and is associated with a variety of neurological disorders. Our understanding of the structure and mechanism of this integral membrane protein is limited. The goal of this study was to use pharmacological, mass spectrometric (MS) and photochemical approaches to probe EAAT2. For MS characterization, a hexahis epitope was incorporated into the N-terminus of human EAAT2. The recombinant protein was functionally expressed in HEK 293T cells and purified through a single-step nickel column. In-gel and in-solution trypsin digestions were conducted on the isolated protein. Overall, eighty-nine percent sequence coverage of the protein was achieved. An 88-amino acid tryptic peptide covering the proposed substrate binding site was revealed after N-deglycosylation. This study provided an efficient and simple method to purify, digest and characterize integral membrane proteins by MS. In addition, the EAAT2 peptide fingerprint obtained by digestion offered a template for later protein modification studies. In an effort to design photoaffinity labels for hEAAT2, a series of aryl diaminopropionic acids and aryl aspartylamide compounds were synthesized and characterized as potent EAAT inhibitors. Compounds containing 9-fluorenone groups were found to be able to irreversibly inactivate EAATs under UV-A illumination. The mechanism underlying the photo-inactivation was shown to be singlet oxygen mediated protein oxidation. The specificity of the photo-inactivation was illustrated by the protection effects of inhibitors, as well as the proximity between the transporter and ligands. Trypsin digestion and MS analyses revealed a mass change of a peptide from hEAAT2 binding pocket in the photo-inactivated protein. Molecular docking results supported our speculation that a tryptophan residue was oxidized during the photo-inactivation. The identification of possible EAAT2 photo-inactivation site provided additional information for the location of the EAAT2 lipophilic interaction domain

Self-deflecting plasmonic lattice solitons and sur-face modes in chirped plasmonic arrays

We show that chirped metal-dielectric waveguide arrays with focusing cubic nonlinearity can support plasmonic lattice solitons that undergo self-deflection in the transverse plane. Such lattice solitons are deeply-subwavelength self-sustained excitations, although they cover several periods of the array. Upon propagation,the excitations accelerate in the transverse plane and follow trajectories curved in the direction in which the separation between neighboring metallic layers decreases, a phenomenon that yields considerable deflection angles. The deflection angle can be controlled by varying the array chirp. We also reveal the existence of surface modes at the boundary of truncated plasmonic chirped arraythat form even in the absence of nonlinearity.Comment: 5 pages, 6 figures, to appear in Optics Letter

Differentiation of Eight Commercial Mushrooms by Electronic Nose and Gas Chromatography-Mass Spectrometry

Volatile profiles of eight mushrooms were characterized by gas chromatography-mass spectrometry and electronic nose analysis. Volatile compounds including 11 alcohols, 11 ketones, 15 aldehydes, 3 sulfur compounds and alkenes, 8 terpenes, 7 acid and esters, 5 heterocyclic compounds, 20 aromatic compounds, and 4 other compounds were identified. The overall aroma properties of the mushrooms were analyzed by the electronic nose. Results indicated that the e-nose sensors have the ability to accurately respond to different mushrooms with similar fingerprint chromatograms. The relationship between the GC-MS data and e-nose responses of different mushrooms was modeled by principal component analysis and partial least squares regression. This combination for the volatile analysis with chemometric methods can be applied to distinguish different mushrooms successfully. Furthermore, it is concluded that the volatile composition of commercial mushrooms could benefit a finger spectrum by e-nose to identify the species of edible fungi