5 years ago

Poly (N-acryloxysuccinimide-co-ethylene glycol dimethacrylate) precursor monolith and its post polymerization modification with alkyl ligands, trypsin and lectins for reversed-phase chromatography, miniaturized enzyme reactors and lectin affinity chromatography, respectively

Ziad El Rassi, Murthy Jonnada
This investigation was aimed at introducing a monolithic precursor that can be conveniently grafted with the desired chromatographic ligand via the process of post polymerization modification (PPM). The precursor was obtained by the in-situ polymerization of N-acryloxysuccinimide (NAS) and ethylene glycol dimethacrylate (EDMA) in a narrow bore stainless steel column of 1 mm i.d. yielding a poly(NAS-co-EDMA) monolithic column designated as the poly(NAS-co-EDMA) monolith (NASM) column. In a first PPM, the NASM column was bonded with octadecyl (OD) ligands yielding a nonpolar NASM-OD column that proved useful for reversed phase chromatography (RPC) of proteins in gradient elution at increasing %ACN in the mobile phase. NASM-OD resulted from the reaction between the N-hydroxysuccinimide of NASM with octadecyl amine. In a second PPM, NASM was surface immobilized with trypsin generating a proteolytic narrow bore enzyme reactor called NASM-trypsin immobilized enzyme reactor (IMER) that permitted the online digestion of proteins in a 20-min single pass through the IMER incorporated in a setup equipped with a short RPC column to achieve simultaneously a peptide tryptic map. This constituted a rapid turnover whereby ∼95% of the protein was hydrolyzed by the immobilized trypsin. In a third PPM, the NASM column was surface immobilized with three different lectins (LCA, Con A and RCA) having complementary affinities toward serum glycoproteins thus permitting the capture of a wide range of glycoproteins/glycoforms. The three NASM-lectin columns when operated in a tandem format led to assessing the level of the various glycoforms in human serum via LC-MS/MS analysis of the captured protein fractions by each NASM-lectin column.

Publisher URL: http://onlinelibrary.wiley.com/resolve/doi

DOI: 10.1002/elps.201700221

You might also like
Discover & Discuss Important Research

Keeping up-to-date with research can feel impossible, with papers being published faster than you'll ever be able to read them. That's where Researcher comes in: we're simplifying discovery and making important discussions happen. With over 19,000 sources, including peer-reviewed journals, preprints, blogs, universities, podcasts and Live events across 10 research areas, you'll never miss what's important to you. It's like social media, but better. Oh, and we should mention - it's free.

  • Download from Google Play
  • Download from App Store
  • Download from AppInChina

Researcher displays publicly available abstracts and doesn’t host any full article content. If the content is open access, we will direct clicks from the abstracts to the publisher website and display the PDF copy on our platform. Clicks to view the full text will be directed to the publisher website, where only users with subscriptions or access through their institution are able to view the full article.