Adsorption of peptides and small proteins with control access polymer permeation to affinity binding sites. Part II

Polymer permeation-ion exchange separation adsorbents with polyethylene glycol and strong anion exchange groups

Omar González-Ortega, Jerker Porath, Roberto Z Guzman

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

In chromatographic separations, the most general problem in small biomolecule isolation and purification is that such biomolecules are usually found in extremely low concentrations together with high concentrations of large molecular weight proteins. In the first part of this work, adsorption and size exclusion chromatography (AdSEC) controlled access media, using polyethylene glycol (PEG) as a semi-permeable barrier on a polysaccharide Immobilized Metal Affinity Chromatography (IMAC) matrix was synthesized and used to develop chromatographic adsorbents that preferentially adsorb and separate low molecular weight biomolecules while rejecting large molecular weight proteins. In this second part, we expand the concept of controlled access polymer permeation adsorption (CAPPA) media by grafting polyethylene glycol (PEG) on a high capacity polysaccharide ion exchange (IEX) chromatographic resin where PEG acts as a semi-permeable barrier that preferentially allows the permeation of small molecules while rejecting large ones. The IEX resin bearing quaternary ammonium groups binds permeated biomolecules according to their ion exchange affinity while excluding large biomolecules by the PEG barrier and thus cannot compete for the binding sites. This new AdSEC media was used to study the retention of peptides and proteins covering a wide range of molecular weights from 1 to 150. kDa. The effect of protein molecular weight towards retention by ion exchange was performed using pure protein solutions. Recovery of insulin from insulin-spiked human serum and insulin-spiked human urine was evaluated under polymer controlled permeation conditions. The CAPPA media consisted of agarose beads modified with amino-PEG-methoxy and with trimethyl ammonium groups, having chloride capacities between 20 and 40. μeq/mL and were effective in rejecting high molecular weight proteins while allowing the preferential adsorption of small proteins and peptides.

Original languageEnglish (US)
Pages (from-to)126-137
Number of pages12
JournalJournal of Chromatography A
Volume1227
DOIs
StatePublished - Mar 2 2012

Fingerprint

Ion Exchange
Access control
Permeation
Adsorbents
Adsorption
Anions
Ion exchange
Polymers
Biomolecules
Binding Sites
Molecular Weight
Molecular weight
Peptides
Proteins
Ion Exchange Resins
Size exclusion chromatography
Insulin
Ammonium Compounds
Gel Chromatography
Polysaccharides

Keywords

  • Adsorption chromatography
  • Bioseparations
  • Ion exchange chromatography
  • Non-fouling surface
  • Polyethylene glycol
  • RAM

ASJC Scopus subject areas

  • Analytical Chemistry
  • Organic Chemistry
  • Biochemistry

Cite this

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abstract = "In chromatographic separations, the most general problem in small biomolecule isolation and purification is that such biomolecules are usually found in extremely low concentrations together with high concentrations of large molecular weight proteins. In the first part of this work, adsorption and size exclusion chromatography (AdSEC) controlled access media, using polyethylene glycol (PEG) as a semi-permeable barrier on a polysaccharide Immobilized Metal Affinity Chromatography (IMAC) matrix was synthesized and used to develop chromatographic adsorbents that preferentially adsorb and separate low molecular weight biomolecules while rejecting large molecular weight proteins. In this second part, we expand the concept of controlled access polymer permeation adsorption (CAPPA) media by grafting polyethylene glycol (PEG) on a high capacity polysaccharide ion exchange (IEX) chromatographic resin where PEG acts as a semi-permeable barrier that preferentially allows the permeation of small molecules while rejecting large ones. The IEX resin bearing quaternary ammonium groups binds permeated biomolecules according to their ion exchange affinity while excluding large biomolecules by the PEG barrier and thus cannot compete for the binding sites. This new AdSEC media was used to study the retention of peptides and proteins covering a wide range of molecular weights from 1 to 150. kDa. The effect of protein molecular weight towards retention by ion exchange was performed using pure protein solutions. Recovery of insulin from insulin-spiked human serum and insulin-spiked human urine was evaluated under polymer controlled permeation conditions. The CAPPA media consisted of agarose beads modified with amino-PEG-methoxy and with trimethyl ammonium groups, having chloride capacities between 20 and 40. μeq/mL and were effective in rejecting high molecular weight proteins while allowing the preferential adsorption of small proteins and peptides.",
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T1 - Adsorption of peptides and small proteins with control access polymer permeation to affinity binding sites. Part II

T2 - Polymer permeation-ion exchange separation adsorbents with polyethylene glycol and strong anion exchange groups

AU - González-Ortega, Omar

AU - Porath, Jerker

AU - Guzman, Roberto Z

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AB - In chromatographic separations, the most general problem in small biomolecule isolation and purification is that such biomolecules are usually found in extremely low concentrations together with high concentrations of large molecular weight proteins. In the first part of this work, adsorption and size exclusion chromatography (AdSEC) controlled access media, using polyethylene glycol (PEG) as a semi-permeable barrier on a polysaccharide Immobilized Metal Affinity Chromatography (IMAC) matrix was synthesized and used to develop chromatographic adsorbents that preferentially adsorb and separate low molecular weight biomolecules while rejecting large molecular weight proteins. In this second part, we expand the concept of controlled access polymer permeation adsorption (CAPPA) media by grafting polyethylene glycol (PEG) on a high capacity polysaccharide ion exchange (IEX) chromatographic resin where PEG acts as a semi-permeable barrier that preferentially allows the permeation of small molecules while rejecting large ones. The IEX resin bearing quaternary ammonium groups binds permeated biomolecules according to their ion exchange affinity while excluding large biomolecules by the PEG barrier and thus cannot compete for the binding sites. This new AdSEC media was used to study the retention of peptides and proteins covering a wide range of molecular weights from 1 to 150. kDa. The effect of protein molecular weight towards retention by ion exchange was performed using pure protein solutions. Recovery of insulin from insulin-spiked human serum and insulin-spiked human urine was evaluated under polymer controlled permeation conditions. The CAPPA media consisted of agarose beads modified with amino-PEG-methoxy and with trimethyl ammonium groups, having chloride capacities between 20 and 40. μeq/mL and were effective in rejecting high molecular weight proteins while allowing the preferential adsorption of small proteins and peptides.

KW - Adsorption chromatography

KW - Bioseparations

KW - Ion exchange chromatography

KW - Non-fouling surface

KW - Polyethylene glycol

KW - RAM

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DO - 10.1016/j.chroma.2011.12.092

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JO - Journal of Chromatography

JF - Journal of Chromatography

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