Efficient Polymer Monolith for Strong Cation-Exchange Capillary Liquid Chromatography of Peptides

Abstract

A stable poly[2-acrylamido-2-methyl-1-propanesulfonic acid-co-poly(ethylene glycol) diacrylate] monolith was synthesized inside a 75-microm-i.d. capillary by photoinitiated copolymerization with water, methanol, and ethyl ether as porogens. The resulting monolith was evaluated for strong cation-exchange capillary liquid chromatography of both synthetic and natural peptides. Although the monolith possessed relatively strong hydrophobicity due to the use of 2-acrylamido-2-methyl-1-propanesulfonic acid as one monomer, the monolith had a high dynamic binding capacity of 157 microequiv of peptide/mL, or 332 mg of cytochrome c/mL. Exceptionally high resolution resulting from extremely narrow peaks was obtained, resulting in a peak capacity of 179 when using a shallow salt elution gradient. Although a second, naturally formed gradient might contribute to the sharp peaks obtained, high efficiency was mainly due to the use of poly(ethylene glycol) diacrylate as a biocompatible cross-linker.

Preparation of capillary hybrid monolithic column with sulfonate strong cation exchanger for proteome analysis

“…Monolithic columns are considered as the new generation of LC stationary phases, which are synthesized by in situ preparation of polymer-based organic and silica-based inorganic monoliths with various specific functionalities arising from the versatile in situ chemical reactions and/or post-modifications, and exhibit high porous structure and mass transfer rate [18,19]. Both organic and inorganic SCX monolithic columns were prepared for different purposes in LC separation [20,21]. Organic sulfonate SCX columns are more convenient to prepare than inorganic ones and the binding capacity can be increased by adjusting the amount of functional monomer in the prepolymerization mixture [22,23] or by increasing grafting time in the post-modification process [24].…”

“…Organic sulfonate SCX columns are more convenient to prepare than inorganic ones and the binding capacity can be increased by adjusting the amount of functional monomer in the prepolymerization mixture [22,23] or by increasing grafting time in the post-modification process [24]. However, the sulfonate organic monolith exhibited excessive shrinking and swelling in different types of separation buffer [22] and there is always some RP retention mechanism mixed into the sulfonate organic monolith [20].…”

Automation of nanoflow liquid chromatography‐tandem mass spectrometry for proteome and peptide profiling analysis by using a monolithic analytical capillary column

“…The viscosity data of the water was taken from ref. [16], then the permeabilities of monolithic column and particle-packed column were determined to be about 7.1610 214 and 1.2610 214 m 2 , respectively. It is obvious that the permeability of LMA-EDMA monolithic column is about six times higher than that of particle-packed columns under flowing with 0.1% formic acid in water.…”

“…On the other hand, different types of monolithic materials based on organic polymers or silica gel monolithic columns prove to be a potential alternative to particulate col-umns for efficient peptide/protein analysis [15][16][17][18][19][20][21]. Higher flow rates can be applied as the porous structure of the monolith significantly reduces the column backpressure and leads to sharp chromatographic peaks with relative high concentration of the eluting peptides [18,20].…”

Developments in ion chromatography using monolithic columns

“…One of the largest areas of use of latexcoated polymeric monoliths use has been in CEC. Capillary polymeric monoliths have also been prepared for biopolymer separation [100], tailored with anion exchangers for nucleotide and oligonucleotide separations [101], tailored with cation exchangers for peptide separation [102], used for hydrophobic interaction chromatography [103] and used or prepared for several other separations [104 -106]. The majority of CEC research has been focused on separating biological or macromolecules.…”