Thesis (Ph. D.)--University of Texas at Austin, 2006. / Vita. Includes bibliographical references.

The synthesis of the sodium salt of heptakis (2, 3-di-O-acetyl-6-O-sulfo)-β-cyclodextrin was modified to increase the isomeric purity to 98.5%. This salt was used to obtain the organic-solvent-soluble, single-isomer, charged tetrabutylammonium salt of heptakis (2, 3-di-O-acetyl-6-O-sulfo)-β-cyclodextrin. Its isomeric purity was higher than 99%, as determined by CE, and its structure was confirmed by NMR and ESI-MS analysis. The hydrophobic single-isomer cyclodextrin was utilized to separate the enantiomers of weak base analytes in aprotic media by capillary electrophoresis. The effective mobilities and separation selectivities follow trends observed with negatively charged cyclodextrins in amphiprotic solvents. The properties of the dissolved cyclodextrin are altered by its counter ion, thereby affecting the separations of enantiomers. The aprotic media allow the modification of the separation selectivity, since the binding strength of the enantiomers to the cyclodextrin is intermediate between that reported in aqueous and methanolic buffers.

Chromosomal mediated antimicrobial resistance in Neisseria gonorrhoeae can develop as a result of three main processes including the alteration of target enzymes, changes in transmembrane transport channels and active efflux pump function. Single nucleotide polymorphisms (SNPs) of target genes such as DNA gyrase (gyrA) and topoisomerase (parC), together with mutations in the promoter regions of the efflux pumps norM and mtr can confer resistance to the macrolides, penicillins and fluoroquinolones. These SNPs were analyzed using the SNaPshot method to allow for rapid detection of resistant isolates. Oligonucleotides were developed in the 5’ to the 3’ direction, ending one nucleotide adjacent to the specific SNP of interest. Single base extension reactions were performed and were detected using capillary electrophoresis. The SNaPshot procedure from Applied Biosystems employed in this study adds a single fluorescently-labelled nucleotide complementary to this SNP at the 3’ end by a primer extension polymerase reaction. Then using capillary electrophoresis, the labelled nucleotide is detected, enabling differentiation between A, C, T, or G. SNP results obtained were verified using DNA sequencing and both single and multiplexed reactions were carried out to increase the efficiency of the procedure. Spiked urine samples were also observed to determine if SNPs could be detected clinically. Single reactions enabled the characterization of all confirmed and relevant SNPs. With multiplex primer extension, multiple peaks were observed, each corresponding to one of the SNPs in the gene. This technique was explored for its applicability to detect SNPs of gyrA and parC mutations. Observable SNP detection limits were seen in spiked urine samples at 108 cells/mL in as early as 4 hours. DNA sequencing results confirmed the SNPs identity in each case. Thus, capillary electrophoresis using the SNaPshot protocol is another way to rapidly identify clinically resistant strains of Neisseria gonorrhoeae. This technique has also been shown to reduce analysis time compared to DNA sequencing and produces the same results. / February 2005

Prostate cancer is one of the most common malignant tumors in solid organs of old men. Prostate-specific antigen (PSA) is a valuable prostate cancer biomarker that is now wildly used for population screening, diagnosis, and monitoring of patients with prostate cancer. Howere it is reported that PSA is not feasible to discriminate the progress of prostate cancer, so many investigators still works on developing new biomarker of prostate carcinoma. Here, we propose the study of differentially expressed prostate proteins in blood of patients. With the aid of two-dimensional polyacrylamide gel electrophoresis (2-DE) and matrix-assisted laser desorption-induced time of flight (MALDI-TOF) mass spectrometry, comparison of normal men and prostate cancer patients serum proteins and analysis protein variation of different stages of prostate cancer serum. We find 31 and 17 protein spots overexpression in cancer development and after treatment, respectively. At present, with the aid of SWISS-PORT database and MALDI-TOF mass spectrometry, we had identified fibrinogen gamma chain, fibrinogen alpha/alpha-E chain, major histocompatibility complex (MHC), class I, C and Mayven were overexpressed in prostate cancer development, where Mayven is fist reported by us.

Photo-polymerized cross-linked polyacrylamide gels are becoming increasingly important for use in micro-fabricated DNA electrophoresis systems because they allow a concentrated sieving matrix to be precisely positioned at any location within a complex micro-channel network. The rate of photo-initiation in the free radical gel polymerization reaction, however, can exert a strong influence on the resulting gel structure. Experimental data on separation resolution of single stranded DNA (ssDNA) in photoinitiated polyacrylamide gels is very sparse. In this study, we investigate the performance of ssDNA electrophoresis in an ALF Express automated DNA sequencer using various photo-initiation chemistries. Cross-linked polyacrylamide gels with concentrations ranging from 6 to 12 %T were prepared using riboflavin, methylene blue, irgacure 651 and ReproGel (AP Biotech) photo-initiators. Separation resolution is compared with that attained in cross-linked polyacrylamide gels prepared using conventional chemical initiators (e.g. ammonium persulfate/ Tetramethylethylenediamine) in order to determine the polymerization conditions necessary for optimum performance.

The synthesis of the sodium salt of heptakis (2, 3-di-O-acetyl-6-O-sulfo)-β-cyclodextrin was modified to increase the isomeric purity to 98.5%. This salt was used to obtain the organic-solvent-soluble, single-isomer, charged tetrabutylammonium salt of heptakis (2, 3-di-O-acetyl-6-O-sulfo)-β-cyclodextrin. Its isomeric purity was higher than 99%, as determined by CE, and its structure was confirmed by NMR and ESI-MS analysis. The hydrophobic single-isomer cyclodextrin was utilized to separate the enantiomers of weak base analytes in aprotic media by capillary electrophoresis. The effective mobilities and separation selectivities follow trends observed with negatively charged cyclodextrins in amphiprotic solvents. The properties of the dissolved cyclodextrin are altered by its counter ion, thereby affecting the separations of enantiomers. The aprotic media allow the modification of the separation selectivity, since the binding strength of the enantiomers to the cyclodextrin is intermediate between that reported in aqueous and methanolic buffers.

The chicken has long been a model used for the study of plasma lipoproteins due to the ability to increase VLDL production by administration of estrogen. In this study we were able to demonstrate successful isolation of VLDL assembly intermediates from the livers of hens, roosters, and estrogen treated rosters. Particle diameter of first step particles, as determined by dynamic laser light scattering, was decreased from an average diameter of 31.5 nm in untreated birds, to 16.1 nm 12 hours after estrogen treatment. Effects of estrogen waned after 24 hours and particle diameter of first step particles increased to an average of 23.9 nm. These assembly intermediates, as well as plasma VLDL and VLDLy, were successfully studied using capillary electrophoresis (CE). Effective mobilities of intact plasma VLDL and first step particles decreased after estrogen administration. Hen VLDL showed a single uniform peak whereas rooster VLDL separated into distinct “subclasses”. Delipidated VLDL, VLDLy and first step assembly intermediates were also successfully separated using CE. This thesis is dedicated to my family who always encouraged me through this process.

Thesis (Ph. D.)--University of Texas at Austin, 2000. / Vita. Includes bibliographical references. Available also in a digital version from Dissertation Abstracts.

Thesis (Ph.D.)--Drexel University, 2009. / Includes abstract and vita. Includes bibliographical references (leaves 117-118).

Thesis (Ph.D.)--Drexel University, 2009. / Includes abstract and vita. Includes bibliographical references (leaves 180-183).