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- All Subjects: NMR
- Creators: Department of Chemistry and Biochemistry
Description
Microwave hydrolysis of egg-white lysozyme was optimized using 1H liquid-state nuclear magnetic resonance (NMR) spectroscopy experiments for amino acid analysis (AAA). Time held under microwave hydrolysis was arrayed for 2, 4, 6, 8, 10, and 15 minutes. Correlations from gCOSY 2D NMR experiments combined with 1H assignments in the one-dimensional chemical shift spectra identified 18 of the 20 amino acids found in lysozyme. Comparison with Uniprot database amino acid composition values revealed the optimal microwave hydrolysis time lies between 4 and 6 minutes. Identification of lysozyme was confirmed with the ExPASy online database search tool AACompIdent. The microwave hydrolysis procedure presented is a simple analytical technique allowing quick and reliable sample preparation in less than one hour that requires no separation or derivation of amino acids residues prior to detection.
ContributorsEdwards, Maximillian Ashur (Author) / Yarger, Jeff (Thesis director) / Marzke, Robert (Committee member) / Barrett, The Honors College (Contributor) / School of Mathematical and Statistical Sciences (Contributor) / Department of Chemistry and Biochemistry (Contributor)
Created2015-05
Description
Amino acid analysis (AAA) of egg white lysozyme and bovine Achilles tendon collagen was performed using 1H solution-state nuclear magnetic resonance (NMR) spectroscopy. The proteins were hydrolyzed in 6M HCL with and without 0.02% phenol at 110\u00B0C for 24, 48, and 72 hours. For both proteins, 18 of 20 amino acids were characterized including hydroxyproline and hydroxylysine in collagen, using 1-dimensional (1D) and 2-dimensional (2D) NMR spectroscopy experiments. Errors ranging from <1% to 8% were seen in treatments with and without phenol. Both proteins could be correctly identified within their own species using the online database search AACompIdent. The proposed approach is a simple analytical technique that does not require the use of column separation or amino acid derivatization prior to compositional analysis.
ContributorsBaranowski, Michael Edward (Author) / Yarger, Jeffery (Thesis director) / Holland, Gregory (Committee member) / Barrett, The Honors College (Contributor) / Department of Chemistry and Biochemistry (Contributor) / Herberger Institute for Design and the Arts (Contributor)
Created2014-05
Description
The goal of the studies described in this thesis was to determine the changes in vascular density in the paraventricular nucleus (PVN) of the hypothalamus following prenatal exposure to the synthetic glucocorticoid hormone, dexamethasone (DEX). DEX is a synthetic glucocorticoid used clinically in women at risk for preterm delivery or in preterm infants to promote proper pulmonary development in high-risk neonates. Prenatal exposure to glucocorticoids such as DEX may change the development of important brain regulatory centers such as the PVN, resulting in increased risk for diseases in adulthood.
Previous studies have demonstrated that the hypothalamus regulates neuroendocrine and autonomic function and behavior. Within the hypothalamus, the paraventricular nucleus (PVN) is an integratory node that contains neurons associated with the control of neuroendocrine and autonomic responses. The PVN also has one of the highest density of blood vessels within the brain. Alterations of normal PVN angiogenesis by dexamethasone could potentially result in long-term modifications of brain and endocrine functions.
Timed-pregnant Sprague Dawley female rats received DEX on gestational days 18-21 and the resulting progeny were sacrificed at Postnatal Day (PND) 0, 4, 14, and 21. A tomato lectin, Lycopersicon Esculentum labeled with DyLight594 was used to stain blood vessels in the PVN and scanning confocal microscopy was used to analyze the experimental brains for PVN blood vessel density
Analysis of data using a 3-way analysis of variance (ANOVA) with age, sex and treatment as main factors, showed a significant age effect in vascular density. Analysis of female data by 2-way ANOVA demonstrated a significant effect of age, but no treatment or interaction effects. Post-hoc analysis shows significant differences at PND 2, 4, 14, and 21 compared to PND0. A Student‘s t-test of a planned comparison on PND2 showed a significant reduction by DEX treatment (p < 0.05). Analysis of data from females, using 2-way ANOVA demonstrated a significant effect of age, but no treatment or interaction effects. Post-hoc analysis shows significant differences at PND 2, 4, 14, and 21 compared to PND0. A planned comparison at PND 2 using Student’s t-test indicated a significant reduction by dex treatment.
The results of these studies demonstrate that there is significant postnatal angiogenic programming and that the vascular density of the PVN is altered by prenatal dexamethasone administration at PND2. The time-course shows developmental fluctuations in vessel density that may prove to be physiologically significant for normal brain function and developmental programming of brain and behavior.
Previous studies have demonstrated that the hypothalamus regulates neuroendocrine and autonomic function and behavior. Within the hypothalamus, the paraventricular nucleus (PVN) is an integratory node that contains neurons associated with the control of neuroendocrine and autonomic responses. The PVN also has one of the highest density of blood vessels within the brain. Alterations of normal PVN angiogenesis by dexamethasone could potentially result in long-term modifications of brain and endocrine functions.
Timed-pregnant Sprague Dawley female rats received DEX on gestational days 18-21 and the resulting progeny were sacrificed at Postnatal Day (PND) 0, 4, 14, and 21. A tomato lectin, Lycopersicon Esculentum labeled with DyLight594 was used to stain blood vessels in the PVN and scanning confocal microscopy was used to analyze the experimental brains for PVN blood vessel density
Analysis of data using a 3-way analysis of variance (ANOVA) with age, sex and treatment as main factors, showed a significant age effect in vascular density. Analysis of female data by 2-way ANOVA demonstrated a significant effect of age, but no treatment or interaction effects. Post-hoc analysis shows significant differences at PND 2, 4, 14, and 21 compared to PND0. A Student‘s t-test of a planned comparison on PND2 showed a significant reduction by DEX treatment (p < 0.05). Analysis of data from females, using 2-way ANOVA demonstrated a significant effect of age, but no treatment or interaction effects. Post-hoc analysis shows significant differences at PND 2, 4, 14, and 21 compared to PND0. A planned comparison at PND 2 using Student’s t-test indicated a significant reduction by dex treatment.
The results of these studies demonstrate that there is significant postnatal angiogenic programming and that the vascular density of the PVN is altered by prenatal dexamethasone administration at PND2. The time-course shows developmental fluctuations in vessel density that may prove to be physiologically significant for normal brain function and developmental programming of brain and behavior.
ContributorsWidener, Andrew John-Claude (Author) / Handa, Robert (Thesis director) / Orchinik, Miles (Committee member) / Mustard, Julie (Committee member) / Barrett, The Honors College (Contributor) / Department of Chemistry and Biochemistry (Contributor)
Created2014-05
Characterization of a Lipid Coating on the Surface of Silk Produced by the Embiid Antipaluria urichi
Description
Insects of the order Embiidina spin sheets of very thin silk fibers from their forelimbs to build silken shelters on bark and in leaf litter in tropical climates. Their shelters are very stiff and hydrophobic to keep out predators and rain. In this study, the existence of an outer lipid coating on silk produced by the embiid Antipaluria urichi is shown using scanning and transmission electron microscopy, FT-IR, and water drop contact angle analysis. Subsequently, the composition of the lipid layer is then characterized by 1H NMR and GC-MS.
ContributorsOsborn Popp, Thomas Michael (Author) / Yarger, Jeffery (Thesis director) / Holland, Gregory (Committee member) / Barrett, The Honors College (Contributor) / Department of Chemistry and Biochemistry (Contributor)
Created2014-05