Gloria Braun

Thesis Title: Development of a ¹³C NMR Technique for the Analysis of the Water-Soluble Metabolic Products of Ethanol in the Gallus gallus Chicken Embryo.

Thesis Abstract: In order to study the effects of the water soluble metabolites of ethanol on chicken embryos via ¹³C NMR, a proper procedure for inducing and isolating these metabolites has been developed. Techniques previously used for Drosophila studies were used as a backbone but required modification for application to chicken embryos. A procedure for obtaining and interpreting the ¹³C NMR spectrum of the metabolites was also adapted from the Drospohila procedures.

Future Plans: Gloria is currently looking for employment in the job market.

Crystal Cornett

Thesis Title: Compositional Membrane Analysis of Cryopreserved Blue Fox (Alopex lagopus) and Silver Fox (Vulpes vulpes) Spermatozoa: Species Differences and Cryopreservation Diluent Effects.

Thesis Abstract: Cryogenic protocols have been successfully developed for storage of farmed silver fox (Vulpes vulpes) spermatozoa. However, these same protocols and modifications of these protocols have failed to satisfactorily preserve spermatozoa collected from farmed blue foxes (Alopex lagopus). Furthermore, morphological studies have failed to reveal any significant differences that could account for the observed fertility differences in the cryopreserved sperm of these two species. Thus, the plasma membrane lipid composition of farmed blue and silver fox spermatozoa was studied. Silver fox spermatozoal membranes contained higher levels of docosapentaenoic acid (DPA; 22:5 w-6), desmosterol, and cholesterol compared to blue fox spermatozoa. Species differences were also observed in somatic cell membrane fatty acid composition. Silver fox erythrocyte membranes had significantly lower levels of arachidonic acid (20:4 w-6) but significantly higher levels of DHA (22:6 w-3) and docosatetraenoic acid (DTA; 22:4 w-6) compared to silver fox erythrocytes. Thus, species differences were observed in both germ-line (spermatozoa) and somatic (erythrocyte) cell membranes.

Effects of cryopreservation diluent additives on spermatozoal membrane fatty acid composition were also studied. While extender-induced changes were observed for blue fox spermatozoal membranes, silver fox spermatozoal membranes were largely resistant to changes in fatty acid composition. Additionally, the effects of two cryoprotectants, egg yolk and catalase, were also studied. Egg yolk induced a decrease in palmitic acid (16:0) in blue fox spermatozoal membranes, but a decrease in DPA in silver fox spermatozoal membranes. The addition of catalase (200 units/mL), however, failed to induce any significant differences in the spermatozoal membrane fatty acid composition of either species.

Future Plans: Crystal will be attending the University of Utah in Salt Lake City, UT in the fall of 2004 as a graduate student in their Interdepartmental Neuroscience Ph.D. Program.

Chris Falkiewicz

Thesis Title: The Analysis and Reversible Binding of Molecular Oxygen in Cu₂B66.

Thesis Abstract: A previously synthesized HB66 ligand was purified via flash chromatography and evaluated using TLC plate analysis. The ligand was assayed employing ¹H NMR and the resulting data were compared to estimated values obtained by means of ChemDrawUltra^®. The binuclear Cu(II) complex [Cu₂(B66)(m-OH)](ClO₄)₂·3H₂O was synthesized, producing a Kelly green crystalline precipitate. A 1 mmol solution of the precipitate in a 5:2 ratio of DMSO:H₂O was assessed via UV-Vis instrumentation and gave a broad d-d band at 658 nm. A C, H, N elemental analysis of the [Cu₂(B66)(m-OH)](ClO₄)₂·3H₂O precipitate was performed which yielded percent differences between the actual and theoretical amounts of carbon, hydrogen, and nitrogen present in the sample less than 0.7%, 5.5%, and 3.0%, respectively. The Cu(I) salt [Cu(CH₃CN)₄]PF₆ was synthesized using standard Schlenk and vacuum line techniques, which produced a pearly-white precipitate that was reacted with the purified HB66 ligand in a Vacuum Atmospheres glovebox under N₂ atmosphere. The resulting Cu₂B66 complex yielded an absorbance peak at 630 nm with an absorbance of 0.098 Abs. In an attempt to reversibly bind molecular oxygen, the Cu₂B66 complex was reacted with O₂ and examined using UV-Vis instrumentation. Since the resulting spectrum only contained a d-d band at 667 nm with an absorbance of 0.312 Abs, and did not contain a Cu-O charge transfer band, it was deduced that oxygen did not bind to Cu₂B66 at room temperature.

Future Plans: Chris will attending the University of Michigan’s school of Pharmacy in the fall of 2004 to pursue a career in nuclear pharmacy.

Lucas Gates

Thesis Title: Computational Analysis of Active Sites in Copper Complexes Designed to Reversibly Bind Dioxygen.

Thesis Abstract: Four copper-based complexes similar to the active site found in hemocyanin were constructed using ab initio and semi-empirical methods. Resultant data were examined to determine which of the ligands were most likely to favor formation with molecular oxygen over formation with chlorine, carbon monoxide, hydroxide, and azide. This examination was based on heat of formation analysis and the SCF energies of reactants required to synthesize the copper ligands as compared to the bound copper ligand products. It was determined that [Cu₂(pzen)(O₂(H₂O)₂], or “copper ligand-B,” was the structure most favored for formation with molecular oxygen among the four structures analyzed. Finally, ab inito and semi-empirically optimized structures of [Cu₂(pzen)(N₃)(H₂O)₂] and [Cu₂(pzen)(O₂)(H₂O)₂] were compared to their X-ray crystal structures to ascertain the accuracy of computationally predicted bond lengths and angles. On average, ab initio optimized structures were in better agreements with X-ray crystal structures than were semi-empirically optimized structures by 1.338%.

Future Plans: For the last three years, Lucas has been writing a story about the impact Benjamin Franklin had on the course of the American Revolution. Following graduation, Lucas will be working full time in an effort toward publication of this work. Research for this novel has already taken him to England, France, Philadelphia and New York. With the bulk of research concluded and only editorial revisions remaining, Lucas hopes to see the first books on the shelves sometime in early 2005.

Carrie Johnston

Thesis Title: The Effect of pH on the Rate of Absorption of Dye by Chitin.

Thesis Abstract: The effect of pH on the absorption of a fiber reactive dye by chitin was studied. Dye solutions of 400, 800 and 1600 ppm were made at pHs of 2, 4, 6, 8 and 10.  The chitin was allowed to be in contact with the dye solution for a time period of three hours while aliquots were removed at various time intervals. The absorbances of aliquots were then analyzed at l_max for the given dye: 506 ± 2 nm, with a Cary 3 UV-Vis spectrometer. The absorbance after the chitin had adsorbed its maximum capacity of dye, A_¥, was taken for each solution and used in a mathematical “stripping” technique in order to separate the slower absorption rate from the faster adsorption rate.  However the results of the experiments were not consistent with results obtained in previous experiments.  In past investigations the chitin adsorbed dye faster at lower pH; however, in the latest experiments the rate of adsorption of dye at pH 10 was also rapid.  It also proved difficult to get R² values of 0.99.  These values ranged from 0.76 to 0.91, indicating that the data was not very good.  At pH 2 absorbances taken at later times of 90 or 120 minutes were sometimes greater than those taken at earlier times of 15 or 20 minutes.  This suggested that the chitin might be releasing dye back into solution after adsorbing it.  When it was discovered that the commercial dye being used was made of a combination of two individual dyes, it was suspected that perhaps the behavior of the two dyes changed with pH, so further analysis by HPLC was done to track the changes in individual dyes. The HPLC results indicated that one of the dyes was adsorbing faster than the other, but it was faster at all pHs, so this idea was discarded.

Future Plans: Carrie is currently looking for employment in the job market.

Sarah Pehlke

Thesis Title: Diffusion and Adsorption of Scarlet #10 Procion Dye by Chitin.

Thesis Abstract: Chitin was investigated for its affinity to adsorb Scarlet #10 procion dye. Three concentrations of dye solutions were exposed to chitin and the adsorption process of dye onto chitin was analyzed. The process proved to be biphasic, as the mass transfer of dye to chitin occurred quiet rapidly, and adsorption through intraparticle diffusion occurred at a much slower rate. The rate constants of intraparticle diffusion were 1.95 x 10^-4 s^-1 for 1600 ppm, 4.445 x 10^-5 s^-1 for 800 ppm, and 5.05 x 10^-5 s^-1 for 400 ppm. The rates were determined by measuring the decrease in absorbance of the dye solutions at l_max 506 nm. The Scarlet #10 procion dye was also investigated using High Performance Liquid Chromatography and was found to be composed of two distinct dyes. Studies suggest that any variation in the adsorption process of the individual dyes that make up Scarlet #10 onto chitin may impact the overall kinetics of the dye and chitin, resulting in slightly inconsistent results compared to previous research on student synthesized dyes.

Future Plans: Sarah will be working with a pharmacist next year and then she plans on attending pharmacy school in the fall of 2005.

Amy Peterson

Thesis Title: Applications of Chemiluminescent Nitrogen Detection to Liquid Chromatographic Separations of Underivatized Amino Acids and Peptide Degradation Products.

Thesis Abstract: Chitin was investigated for its affinity to adsorb Scarlet #10 procion dye. Three concentrations of dye solutions were exposed to chitin and the adsorption process of dye onto chitin was analyzed. The process proved to be biphasic, as the mass transfer of dye to chitin occurred quiet rapidly, and adsorption through intraparticle diffusion occurred at a much slower rate. The rate constants of intraparticle diffusion were 1.95 x 10^-4 s^-1 for 1600 ppm, 4.445 x 10^-5 s^-1 for 800 ppm, and 5.05 x 10^-5 s^-1 for 400 ppm. The rates were determined by measuring the decrease in absorbance of the dye solutions at l_max 506 nm. The Scarlet #10 procion dye was also investigated using High Performance Liquid Chromatography and was found to be composed of two distinct dyes. Studies suggest that any variation in the adsorption process of the individual dyes that make up Scarlet #10 onto chitin may impact the overall kinetics of the dye and chitin, resulting in slightly inconsistent results compared to previous research on student synthesized dyes.

Future Plans: Amy will be attending the University of Michigan’s School of Dentistry in the fall of 2004.

Michelle Phillips

Thesis Title: Separation Conditions for Underivatized Amino Acids with High Performance Liquid Chromatography and Chemiluminescent Nitrogen-Specific Detection.

Thesis Abstract: This research was done in order to find the most effective separation conditions for amino acids produced by hydrolysis of the degradation products of Vasopressin and Oxytocin using Reverse Phase-High Performance Liquid Chromatography and Chemiluminescent Nitrogen-Specific Detection. While there is room for further development great progress was made in developing conditions that would lead to a timely process, providing well-defined peaks for each amino acid in the sample. Multiple mobile phases were investigated and led to the discovery of a 25 minute overall run time. The rather short re-equilibration time used in this run was established using 10 mM NFPA with 5 mM TSA in 75/25 MeOH/Milli Q H₂O. The best run was on a 30 minute gradient. Isocratic runs were also used but didn’t provide results as nice as the gradient. The hydrolysis of peptides was also experimented with, trying a process that would take less than 24 hours in an attempt to be time efficient. However, the amino acids can be destroyed in some forms of hydrolysis and caution was necessary. The shorter hydrolysis process used in this research was not found to be satisfactory.

Future Plans: Michelle is currently looking for employment in the job market.

Amanda Proctor

Thesis Title: The Effects of Ethanol on the Brain Tissue of the Gallus gallus Chick Embryo as Determined by Nuclear Magnetic Resonance.

Thesis Abstract: The effects of ethanol on the membrane composition of the brain tissue in developing Gallus gallus chick embryos were examined by ¹³C NMR. Experimental procedures were developed to isolate and then analyze the fat-soluble components in chick brain samples using ¹³C NMR. It was determined that the effects of ethanol on brain tissue in developing chicks includes a decrease in polyunsaturated fatty acids and a subsequent increase in saturated, short chain fatty acids. Through ¹³C NMR studies, the incorporation of ethyl-2-¹³C alcohol into biomembranes through ethanol metabolism has been observed and may be responsible for the changes seen in membrane composition.

Future Plans: Amanda will be attending medical school at the Medical College of Ohio in the fall of 2004.

Ethan Rouse

Thesis Title: Chitosan Membranes for Reduction of Cu2+ Ions in Industrial Waste Waters.

Thesis Abstract: A macroporus chitosan membrane was prepared from chitosan. The prepared chitosan membrane was tested as a binding agent for Cu²⁺ ions that are commonly found in industrial waste water. The chitosan was placed into a glass funnel through which sample solutions with concentrations of 1000, 500, 250, 100, 50, 25 ppm Cu²⁺ were passed. Removal of Cu²⁺ from solutions was an average of 93.11%. UV-Visible spectroscopy was used to determine Cu²⁺ concentrations in the samples before and after passing through membrane. It was found that the multiple factors, including surface area, the amount of chitosan present, molecular weight, and mechanical design, affect chitosan membrane’s ability to absorb Cu²⁺ from aqueous solutions. Chitosan absorption of Cu²⁺ ions is possible, but with the experimental setup used, it was not possible to obtain concentrations of Cu²⁺ below 25 ppm using chitosan membranes.

Future Plans: Ethan is currently looking for employment in the job market.