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DEVELOPMENT OF A UNIVERSAL POLYMERIC STATIONARY PHASE FOR SOLID PHASE EXTRACTION AND AN IONIC LIQUID MOBILE PHASE MODIFIER FOR SEPARATION OF NATIVE PROTEINS BY LIQUID CHROMATOGRAPHY

Abstract Details

2013, Doctor of Philosophy, Miami University, Chemistry and Biochemistry.
This dissertation consists of four research chapters, which address first the synthesis and characterization of a universal solid phase extraction (SPE) stationary phase and then the use of an alkylammonium formate ionic liquid as the reversed phase liquid chromatography (RPLC) mobile phase modifier for the separation of native proteins. The last chapter outlines a new chemical education experiment for teaching ion exchange. A universal SPE sorbent Strata-Kel-F-p-aminobenzoic acid (Strata-Kel-PABA) was synthesized by a two-step crosslinking reaction followed by an ester hydrolysis. The universal SPE sorbent combines fluorinated reversed phase, cation- and anion-exchange sites together. Compared with the commercially available cartridge Strata-X AW, Strata-Kel-PABA showed better cation exchange and reversed phase capacity and fair anion exchange capacity. The long term goal of this research is equipping this universal SPE cartridge with the autosampler of a RPLC instrument sample preparation. Proteins are hard to separate in their native conformation using RPLC because the tertiary and quaternary structures can be seriously affected by hydrophobic interactions with both stationary and mobile phases in RPLC. The ionic liquid, ispropylammonium formate (IPAF) has been investigated to improve protein stability as a novel RPLC mobile phase modifier. Promising results were shown for non-subunit and two-subunit proteins. The possible mechanism of the IPAF effect on protein stabilization can be explained by the micelle-like structure of the ionic liquid and the Hofmeister series. For labile four subunit proteins, such as lactate dehydrogenase (LDH), polyethylene glycol (PEG) has been combined with IPAF as the mobile phase additive to stabilize the protein. Remarkable enhancements in LDH peak intensity and LDH activity were observed by incorporating 6% PEG 8000 in the organic mobile phase that contained either acetonitrile (MeCN) or IPAF.
Neil Danielson (Advisor)
Andre Sommer (Committee Chair)
Shouzhong Zou (Committee Member)
Hong Wang (Committee Member)
Melany Fisk (Committee Member)
118 p.

Recommended Citations

Citations

  • Zhou, L. (2013). DEVELOPMENT OF A UNIVERSAL POLYMERIC STATIONARY PHASE FOR SOLID PHASE EXTRACTION AND AN IONIC LIQUID MOBILE PHASE MODIFIER FOR SEPARATION OF NATIVE PROTEINS BY LIQUID CHROMATOGRAPHY [Doctoral dissertation, Miami University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=miami1371503762

    APA Style (7th edition)

  • Zhou, Ling. DEVELOPMENT OF A UNIVERSAL POLYMERIC STATIONARY PHASE FOR SOLID PHASE EXTRACTION AND AN IONIC LIQUID MOBILE PHASE MODIFIER FOR SEPARATION OF NATIVE PROTEINS BY LIQUID CHROMATOGRAPHY . 2013. Miami University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=miami1371503762.

    MLA Style (8th edition)

  • Zhou, Ling. "DEVELOPMENT OF A UNIVERSAL POLYMERIC STATIONARY PHASE FOR SOLID PHASE EXTRACTION AND AN IONIC LIQUID MOBILE PHASE MODIFIER FOR SEPARATION OF NATIVE PROTEINS BY LIQUID CHROMATOGRAPHY ." Doctoral dissertation, Miami University, 2013. http://rave.ohiolink.edu/etdc/view?acc_num=miami1371503762

    Chicago Manual of Style (17th edition)