Browsing by Subject "Liquid chromatography"
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Item Bonded phase high performance liquid chromatography of spore mycotoxins(Texas Tech University, 2001-05) Qi, ZhuhuaMycotoxins produced by molds are common contaminants of many important crops, including wheat, com, rice and peanuts. Some mycotoxins are found in fruits and vegetables. These contaminants have a broad range of toxic effects, including carcinogenicity, neurotoxicity, and reproductive and developmental toxicity. The occurrence of mycotoxins in foods is an unavoidable worldwide problem. About 80 countries have imposed regulatory limits to minimize human and animal exposure to mycotoxins. Regulatory limits, including international standards, have tremendous economic impact and must be developed using scientific-based risk assessments. This thesis is focused on the high-performance liquid chromatography (HPLC) analysis of mycotoxins. Chapter I is a review of the application of HPLC on the determination of mycotoxins. Two major types of mycotoxins - aflatoxins and trichothecenes are discussed in details. Other mycotoxins are also surveyed. 113 references are cited. Chapter II describes a simple and sensitive method for simultaneous determination of verrucarin A, verrucarol and roridin A. This method was applied to the analysis of a spore mycotoxin sample form Stachybotrys atra. Greatly improved analytical separations resulted from the use a new phenyl-hexyl bonded phase column. Chromatographic and analytical figures of merit are presented. Additional work is also discussedItem Cyclodextrin stationary phases: synthesis, characterization and applications in liquid chromatography(Texas Tech University, 1986-08) Alak, Ala MSince the creation of the first octadecyl bonded phase for High Performance Liquid Chromatography (HPLC), liquid chromatograhers have investigated a wide variety of bonded phases in their ongoing search for increased selectivity. These studies have led to a number of useful phases which appear to separate by different mechanisms such as hydrophobicity, ionic properties, electron density, and so forth. A highly selective chiral bonded phase material for HPLC and Thin Layer Chromatography (TLC), was produced by bonding cyclodextrin to silica gel particles through a spacer "arm" via a reproducible process which yields a stable, non-hydrolytic, non-nitrogen containing bonds. The synthesis, characterization and applications of these new stationary phases are described. The basic property of cyclodextrin molecules, which allows them to effect numerous chemical separations, is their ability to form selective inclusion complexes with a wide variety of organic and inorganic molecules. The formation of these inclusion complexes may occur by a combination of factors, such as hydrophobic effects, dipole-dipole interaction, hydrogen bonding (between the cyclodextrins hydroxyl groups and guest molecules) or the release of high energy water or modifier during complex formation. In general, binding to the cyclodextrin is governed by the molecule's ability to closely fit inside the cavity of the cyclodextrin, although the polarity of the molecule also plays an important role. Besides their ability to separate different kinds of enantiomers by HPLC and TLC, cyclodextrin stationary phases also proved to be the media of choice in separation of dlastereomers, geometrical and structural isomers, and many nonisomeric compounds. The effects of temperature, mobile phase composition, and flow rate upon chromatographic selectivity and resolution are described. The results indicate that cyclodextrin stationary phases are versatile, flexible and effective compared to conventional normal or reversed phase stationary phases.Item Cyclodextrins and micelles in separations(Texas Tech University, 1987-05) Ward, Timothy JosephNot availableItem MS Based Approaches for the Analysis of Small Molecules such as Sugar Nucleotides and Gangliosides(2013-05) Garcia, Aldo; Mechref, Yehia; Thompson, Jonathan E.; Pare, PaulThe field of metabolomics has become a hot spot of analytical investigations since the discovery of the encompassing significance of the metabolome in complex eukaryotic systems. Most notably, emphasis in this field is currently being placed in the discovery of potential biomarkers through development of highly sensitive methods that are capable of identifying and quantifying small metabolites of interest in complex samples. In this thesis, a concise review is described in chapter 1 over the field of metabolomics as it pertains to the analytical developments of sugar nucleotides and ganglioside analysis. In addition, targeted mass spectrometric approaches are presented in which analytical methodologies are developed for sugar nucleotide and ganglioside profiling for the analysis of diseased biological samples (such as cancer) , with interest focused on the establishment of metabolic profiles of diseased states. Chapter 2 details the analytical method for targeted mass spectrometric analysis of sugar nucleotides. The method described permitted the establishment of a sugar nucleotide linear dynamic range of quantification expanding over almost three orders of magnitude with an extremely reproducible chromatographic separation in less than six minutes. Viability of this method was then tested by successfully quantifying the sugar nucleotide content of human cells from 3 different cancer cell lines. The developed short chromatographic method potentially permits for high throughputs analysis for its integration into population studies for the search for potential biomarkers. Chapter 3 describes the developed methodology for targeted ganglioside analysis. A chromatographic method was developed based on the use of a mixture of bovine brain gangliosides that allowed for complete separation in less than 7.5 minutes. Quantificationof gangliosides was also achieved by the construction of a calibration curve from ganglioside standards. The established method was then successfully employed for the ganglioside profiling of 3 diseased samples of human blood serum along with one disease free control. As a result, ganglioside profiles were quantitatively established at high sensitivity for the different ganglioside samples.Item Novel uses of organized media in chemical analysis(Texas Tech University, 1987-08) Spino, Larry AngeloMicelles and cyclodextrins have been utilized for years in a variety of chemical analysis techniques. This work considers some modern applications of micelles and cyclodextrins in chemical analysis. Micellar and cyclodextrin mobile phases (pseudophases) were used in liquid chromatography (LC). Cyclodextrin mobile phases were used in microcolumn LC for separating optical isomers of racemic nicotine and eight other racemic analogues of nicotine. This was the first reported facile and direct separation of these racemates. Synchronous luminescence was used as the detection method for liquid chromatography to identify polynuclear aromatic components that co-elute in the course of a LC separation. As many as five compounds could be identified from a single chromatographic peak. Several scans can be made easily for every eluting peak, however, a single scan is sufficient to produce a complete synchronous luminescence spectra of a complex mixture. The micellar matrix served as the mobile phase and produced an enhancement effect on the luminescence signals. Equations were derived which allow one to determine alphacyclodextrin: substrate complex stoichiometries as well as primary and secondary binding constants by using LC retention values. An equation was also derived which describes the binding of a monoprotic species in which either its ionized or unionized form could bind to one or two cyclodextrin molecules. Becaused multiple binding constants are difficult to evaluate graphically, a non-linear least squares computer program was utilized. The approach works equally well forthe determination of binding cqnstants in micellar media. Resonance enhancement of Raman signals requires excitation on an absorption band of a molecule. This frequently produces background fluorescence from which it is difficult or impossible to extract a vibrational spectrum. Carrying out the resonance Raman analysis in certain dilute aqueous micellar solutions allows one to circumvent the luminescence problem in many cases. Several difÃerent micellar effects can be used simultaneously to enhance Raman signals relative to the background. Both the laser excitation line and the micellar system must be properly chosen so as to produce the best signal to noise ratio. The first examples of micelle mediated resonance Raman analysis of fluorescent compounds using UV and visible laser excitation was shown. Obtaining resonance Raman spectra from thin-layerchromatography plates was also demonstrated.Item Qualitative and Quantitative analysis of glycans and glycopeptides released from model glycoproteins and biological samples using MRM mode and ESI LC-MS/MS(2012-12) Pyreddy, Swetha; Mechref, Yehia; Pappas, Dimitri; Birney, David M.Glycosylation is one of the most important post translational modifications. Aberrant glycosylation has been correlated to a number of disease conditions. Changes in glycosylation can provide valuable information about the etiology of the disease state and also can be used as potential biomarkers for early prognosis and diagnosis. Currently, liquid chromatography interfaced to mass spectrometry is the most widely used techniques for the characterization of glycans and glycopeptides in order to detect changes in glycosylation. Development of reliable methods is required for correlation of these changes with the disease condition. The first part of the thesis describes method development and validation for the successful quantification of the glycopeptides using oxonium ions as transitions in multiple reaction monitoring (MRM) mode. The invariable presence of oxonium ions as a result of fragmentation of glycopeptides makes them an excellent choice as transitions for MRM of the glycopeptides. MRM on a triple quadrupole mass spectrometer enables very high sensitivity and is explored as a technique for quantification of glycopeptides. The various parameters influencing quantification of glycopeptides including MRM time segments, number of transitions and normalized collision energies were optimized. The results indicate that oxonium ions could be adopted for the characterization and quantification of glycopeptides in general, eliminating the need to select specific transitions for individual precursor ions. Also, enhanced sensitivity of analysis is attained when specific time segments are employed. This approach can also be applied to comparative and quantitative studies of glycopeptides in biological samples as illustrated for the case of depleted blood serum. The second part of the thesis describes characterization and quantification of glycans and glycopeptides of the aqueous humor samples collected from Fuchs endothelial corneal dystrophy (FECD) and normal subjects using LC-ESI-MS/MS. FECD is an eye disorder which results in progressive degeneration of the corneal epithelium causing impaired vision and in some cases total blindness. The exact biological events that cause this disorder are still unknown. In an attempt to establish a correlation between aberrant glycosylation and FCED, the glycome and the glycoproteome of aqueous humor was probed. The results suggest a significant difference between the disease and disease free states of one glycan. A general upregulation of glycan expression in the disease state was observed according to results of the glycomic study. These changes can be used to distinguish between the subjects with and without FECD. Also, these changes suggest that glycosylation is either an effect or cause of the disease condition.