title: Synthesis and application of novel phenylboronate affinity materials based on organic polymer particles for selective trapping of glycoproteins.
abstract: We report on synthesis concepts for the fabrication of various novel phenylboronate affinity materials based on polymethacrylate epoxy beads (Fractogel EMD Epoxy (M) 40-90 microm) and the testing of these functionalized polymer particles for selective trapping of a glycoprotein from a standard mixture containing a glycosylated and a nonglycosylated protein. Two inherently different approaches for the functionalization of the bare beads with boronate groups have been elucidated. In the first, the epoxy residues of the polymer particles were converted into reactive thiol groups which were subsequently used as anchor moieties for the immobilization of 4-vinylphenylboronic acid by radical addition or radical polymerization reaction. Three different ways for the generation of sulfhydryl groups have been examined leading to materials with distinct linker chemistries. In the second and more straightforward approach, the epoxy groups were reacted with 4-mercaptophenylboronic acid. The novel materials were thoroughly characterized by (i) quantitation of the sulfur content by elemental analysis, (ii) reactive sulfhydryls were determined in a photospectrometric assay, (iii) boron content was measured by inductively coupled plasma-atomic emission spectrometry, and (iv) the amount of reactive boronate groups was evaluated in a fast binding assay employing adenosine as test compound. A maximum concentration of 1.2 mmol boronate groups per gram dry beads could be achieved by the presented synthesis routes. Employing the novel phenylboronate affinity materials in capture and release experiments in the batch mode, a standard glycoprotein, viz. transferrin (Tf) from human serum was separated from a nonglycosylated protein, BSA. A commercial boronate affinity material based on 3-aminophenylboronic acid modified agarose gel was employed as reference material and was found to perform significantly worse compared to the herein presented novel polymethacrylate particles.
authors.affiliation: Christian Doppler Laboratory for Molecular Recognition Materials, Department of Analytical Chemistry and Food Chemistry, University of Vienna, Vienna, Austria.
authors.forename: Beatrix
authors.initials: B
authors.lastname: Preinerstorfer
authors.forename: Michael
authors.initials: M
authors.lastname: Lämmerhofer
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Lindner
mesh_terms: D000818:Animals; D001895:Boron / Q000032:analysis; D001896:Boron Compounds / Q000032:analysis; D001897:Boronic Acids / Q000138:chemical synthesis* / Q000737:chemistry*; D002417:Cattle; D005591:Chemical Fractionation / Q000379:methods*; D006023:Glycoproteins / Q000097:blood / Q000737:chemistry* / Q000302:isolation & purification*; D006801:Humans; D010316:Particle Size; D011108:Polymers / Q000138:chemical synthesis* / Q000737:chemistry*; D012710:Serum Albumin, Bovine / Q000737:chemistry; D013438:Sulfhydryl Compounds / Q000032:analysis; D013455:Sulfur / Q000032:analysis; D014168:Transferrin / Q000737:chemistry / Q000302:isolation & purification
chemical_list: D001896:Boron Compounds; D001897:Boronic Acids; D006023:Glycoproteins; D011108:Polymers; D013438:Sulfhydryl Compounds; D014168:Transferrin; D012710:Serum Albumin, Bovine; D013455:Sulfur; C010686:benzeneboronic acid; D001895:Boron