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Martin Beres Dissertation.pdf (5.74 MB)

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Novel Analytical Methods for Improved Analysis of Biological Compounds

Beres, Martin Joseph
http://rave.ohiolink.edu/etdc/view?acc_num=osu1447408723

Abstract Details

2015, Doctor of Philosophy, Ohio State University, Chemistry.
The work contained within this dissertation focuses on innovative technologies in the field of analytical chemistry, particularly within high-performance liquid chromatography (HPLC) and mass spectrometry (MS). Enhanced-fluidity liquids (EFL), which have low viscosity and high diffusivity, were studied as alternative mobile phases in mixed-mode hydrophilic interaction strong ion-exchange chromatography (HILIC/SCX). Additionally, these mobile phases were evaluated as environmentally friendly alternatives to traditional HILIC solvents in gradient separations. Finally, electrospun nanofibrous materials with high surface area to volume ratios were assessed as substrates in surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS). The potential of enhanced-fluidity liquid chromatography (EFLC) HILIC/SCX was explored, using amino acids as analytes. EFL mobile phases were prepared by adding liquefied CO2 to methanol:water (MeOH:H2O) mixtures, which increases the diffusivity and decreases the viscosity of the mixture. The optimized chromatographic performance of these MeOH:H2O:CO2 EFL mixtures was compared to traditional acetonitrile:water (ACN:H2O) and MeOH:H2O liquid chromatography (LC) mobile phases. MeOH:H2O:CO2 mixtures offered higher efficiencies and resolution of the ten amino acids relative to the MeOH:H2O LC mobile phase, and decreased the required isocratic separation time by a factor of two relative to the ACN:H2O LC mobile phase. Large differences in selectivity were also observed between the EFLC and LC mobile phases. Retention mechanism studies revealed that the EFLC mobile phase separation was governed by a mixed-mode retention mechanism of HILIC/SCX. On the other hand, separations with ACN:H2O and MeOH:H2O LC mobile phases were strongly governed by only one retention mechanism, either HILIC or SCX, respectively. EFLC was then evaluated for “green” HILIC separations. The impact of CO2 addition to a MeOH:H2O mobile phase was studied as an alternative to traditional ACN:H2O HILIC mobile phases, while also optimizing buffer type, ionic strength, and pH. Using EFLC mixtures, a separation of 16 RNA nucleosides/nucleotides was achieved in 16 minutes with greater than 1.3 resolution for all analyte pairs. By using a reverse CO2 gradient, analysis time was reduced by over 100% in comparison to isocratic conditions. The optimal separation using MeOH:H2O:CO2 mobile phases was also compared to that using MeOH:H2O and ACN:H2O mobile phases. Based on the chromatographic performance parameters (efficiency, resolution, and speed of analysis) and the overall environmental impact of the mobile phase mixtures, MeOH:H2O:CO2 mixtures were preferred to ACN:H2O or MeOH:H2O mobile phases for the separation of mixtures of these RNA nucleosides and nucleotides. Finally, electrospun nanofibrous substrates were studied for the improvement of SALDI-MS analysis of large molecular weight proteins and polymers without the use of a chemical matrix. Various polymers (including polyacrylonitrile, polyvinyl alcohol, and SU-8 photoresist) and carbon substrates were examined. SALDI analysis using these substrates eliminated “sweet spot” formation typically seen in matrix-assisted laser desorption/ionization (MALDI), which lead to greater shot-to-shot reproducibility. The fiber diameter of these substrates played a significant role in the quality of the mass spectra generated, with smaller fiber diameter yielding higher signal to noise ratio (S/N). Additionally, the degree of pyrolysis also impacted the degree of fragmentation and overall S/N for the prepared carbon substrates.
Susan Olesik (Advisor)
Prabir Dutta (Committee Member)
Vicki Wysocki (Committee Member)
Pravin Kaumaya (Committee Member)
250 p.
Analytical Chemistry; Chemistry
High-performance liquid chromatography; Enhanced-fluidity liquid chromatography; Surface-assisted laser desorption ionization; Matrix-enhanced laser desorption ionization; Mass spectrometry

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Citations

  • Beres, M. J. (2015). Novel Analytical Methods for Improved Analysis of Biological Compounds [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1447408723

    APA Style (7th edition)

  • Beres, Martin. Novel Analytical Methods for Improved Analysis of Biological Compounds. 2015. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1447408723.

    MLA Style (8th edition)

  • Beres, Martin. "Novel Analytical Methods for Improved Analysis of Biological Compounds." Doctoral dissertation, Ohio State University, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=osu1447408723

    Chicago Manual of Style (17th edition)

osu1447408723
799
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This open access ETD is published by The Ohio State University and OhioLINK.