Novel Support Material with Polymeric Ligand for Ion Exchange Chromatography: Synthesis, Characterization and Chromatographic Performance

Year 2019, Volume: 3 Issue: 2, 173 - 183, 23.12.2019

Berna Saraçoğlu Kaya , Ali Tuncel

Abstract

Within the scope of the study, polymeric ligands
with weak cation exchange carboxyl (-COOH) groups formed on the surface,
designed for High Performance Liquid Chromatography (HPLC), monodisperse and
porous poly(2,3-dihydroxypropyl methacrylate-co-ethylene dimethacrylate),
poly(DHPM- co-EDM) particles were tested in Ion Exchange Chromatography (IEC)
mode for chromatographic separation of protein mixtures as a weak cation
exchange column-material. The synthesis of the
particles obtained for the design of the column support material, which is the
first step of the study carried out in three stages, takes place in four
different sections. Firstly, monodisperse
polystyrene microgels to form the skeletal structure were synthesized by “Dispersion
Polymerization”, then monodispersed-porous poly(glycidyl methacrylate-co-ethyleneglycol
dimethacrylate), poly(GMA-co-EDM) microspheres with epoxy groups were produced
by using “multi-stage microsuspension polymerization”. The particles were hydrolyzed in acidic medium
to form the surface in diol form and poly(2,3-dihydroxypropyl
methacrylate-co-ethylene dimethacrylate), poly(DHPM-co-EDM) particles were
obtained which were suitable for chromatographic use. The particles obtained in the last step were
used in the “Free Radical Grafting Polymerization” (FRGP) method for the
production of polymeric ligand containing carboxyl group. Poly (DHPM-co-EDM) particles with –OH groups on
the surface are silanated with 3-trimethoxysilylpropyl methacrylate (TMSPM) and
derivatized with methacrylic acid (MAA) to form poly(MAA) ligand on the surface
of poly(DHPM-c-EDM) particles were obtained for ion-exchange chromatography. The characterization and chromatographic
performance of the particles were investigated in the second and third stages
of the study. For particles
synthesized by the FRGP method, the average-pore size was 33 nm and the
specific-surface area was 21 m²/g. Theoretical plate height values ​​between 200-320 µm were obtained for weak
cation exchange column. The peak resolution values ​​in the chromatogram were
found to be above 1.4 for separation as short as 15 minutes. Protein recovery
values ​​were found to be between 90-100% for all proteins. For
the produced poly(MAA)-grafted-poly(DHPM-co-EDM) column, the BSS values ​​of
the analytes from analysis to analysis and from day to day reproducibility
tests were below 1%. These results showed that synthesized column packed
materials can be successfully used in ion exchange chromatography. 

Keywords

Free Radical Grafting Polymerization (FRGP), High Performance Liquid Chromatograpy (HPLC), Ion Exchange Chromatography (IEC), Monodisperse and porous polymeric particles

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