title: Nanostructured porous polymer monolithic columns for capillary liquid chromatography of peptides.
abstract: The macroporous structure of poly(styrene-co-divinylbenzene) monolithic capillary columns has been optimized for the gradient separation of peptides. To exploit monolithic supports with porosity exceeding 70%, the thermodynamic properties of the polymerization mixture were carefully tailored to yield homogeneous monolithic materials featuring macropore and polymer microglobule sizes in the range of 50–200 nm. The effects of (i) initiator content, (ii) composition of porogenic mixture, comprising tetrahydrofuran and 1-decanol, (iii) percentage of divinylbenzene crosslinker, and (iv) monomers to porogen ratio on the morphology was investigated. The resulting column structures were investigated using scanning electron microscopy and the prepared monolithic columns were tested for the separation of a tryptic digest of cytochrome c while applying a fixed flow rate and gradient time. To obtain a better understanding of the effects of macropore and microglobule size, and structure homogeneity on the separation performance in gradient elution, both in terms of peak capacity and gradient plate height, separations were also carried out at different flow rates while maintaining a constant gradient steepness. Furthermore, performance limits were determined applying ultra-high pressure conditions up to the maximum system pressure of 80 MPa. The potential of monolithic nanostructured columns is demonstrated for the separation of tryptic digests of cytochrome c and bovine serum albumin.
authors.affiliation: Vrije Universiteit Brussel, Department of Chemical Engineering, Brussels, Belgium.
authors.forename: Axel
authors.initials: A
authors.lastname: Vaast
authors.affiliation: Vrije Universiteit Brussel, Department of Materials and Chemistry, Brussels, Belgium.
authors.forename: Herman
authors.initials: H
authors.lastname: Terryn
authors.affiliation: The Molecular Foundry, E.O. Lawrence Berkeley National Laboratory, Berkeley, CA, USA.
authors.forename: Frantisek
authors.initials: F
authors.lastname: Svec
authors.affiliation: Vrije Universiteit Brussel, Department of Chemical Engineering, Brussels, Belgium. Electronic address: seeltink@vub.ac.be.
authors.forename: Sebastiaan
authors.initials: S
authors.lastname: Eeltink
mesh_terms: D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation* / Q000379:methods; D045304:Cytochromes c / Q000737:chemistry; D049329:Nanostructures / Q000737:chemistry*; D010455:Peptides / Q000302:isolation & purification*; D011137:Polystyrenes / Q000737:chemistry*; D016062:Porosity; D012710:Serum Albumin, Bovine / Q000737:chemistry
chemical_list: D010455:Peptides; D011137:Polystyrenes; C432006:divinylbenzene-polystyrene copolymer; D012710:Serum Albumin, Bovine; D045304:Cytochromes c