Poly(methyl vinyl ether-alt-maleic anhydride) (PMM(0)) was partially hydrolyzed in a 9/1 acetonitrile-d(3)/D(2)O mixture and then diluted with an aqueous buffer and coated onto poly-L-lysine (PLL)-coated calcium alginate capsules. The resulting 50% hydrolyzed polymer (PMM(50)) is bound to the surface-immobilized PLL through both electrostatic and covalent interactions, resulting in a shell-cross-linked hydrogel capsule that is resistant to chemical challenges. Further hydrolysis of PMM(50) in aqueous buffer was monitored by potentiometry and was found to proceed with a half-life time of about 2.5 min at 20 degrees C such that residual anhydride groups not consumed by cross-linking with PLL would be deactivated by hydrolysis within several minutes of shell formation, removing potential sites for undesired protein binding. Initial protein-binding tests involving incubation of the capsules in bovine serum albumin solutions for 24 h showed no indication of protein binding. The effects of coating temperature, PLL concentration and molecular weight, PMM(50) molecular weight, and multiple PLL-PMM(50) coatings on shell morphology and behavior were studied using confocal fluorescence microscopy as well as chemical challenges involving sodium citrate and sodium hydroxide. The resilience of the cross-linked shell improved with increasing concentrations of PLL and decreasing molecular weight of PMM(50), both of which resulted in more polyelectrolyte being bound to the capsule. The permeability of these covalently cross-linked capsules was studied using fluorescently labeled dextrans and was found to be comparable to standard calcium alginate-PLL-alginate (APA) capsules.

The formation of reactive polyanions by semibatch copolymerization of 4,4-dimethyl-2-vinyl-2-oxazoline-5one (VDMA) and methacrylic acid (MAA) by both free radical and photoinduced radical polymerization is described. The reactivity ratios of these two monomers were determined to be 1.36 and 0.41 for r 1 (VDMA) and r 2 (MAA), respectively, using 1 H NMR spectroscopy. During the free radical copolymerization of a 50:50 VDMA:MAA copolymer (PMV 50 ) in anhydrous DMSO or THF at 60 °C, up to 40% of the azlactone groups in the copolymer are hydrolyzed by water formed largely by conversion of methacrylic acid diads into cyclic anhydride. Storage in organic solutions leads to further transhydration, while solid PMV 50 is stable for at least 6 months at room temperature. Increasing the VDMA comonomer content reduces this transhydration, likely through decreasing the incidence of methacrylic acid diads in the backbone. Alternatively, conducting the copolymerization at 20 °C using photoinitiation is also effective at suppressing this transhydration. The resulting reactive polyanions bind under physiological conditions to poly-L-lysine-coated calcium alginate hydrogel beads and spontaneously cross-link with the polyamine to form covalent networks of interest for long-term therapeutic cell encapsulation. This represents the first such use of a VDMA-containing polyanion in aqueous environments. The copolymers were characterized by 1 H NMR, quantitative 13 C NMR, 13 C DEPT-135 NMR, and FT-IR spectroscopies as well as by elemental analysis.

Preparation and study of a series of copolymers incorporating 2-vinyl-4,4-dimethylazlactone (VDMA) is reported. The reactivity ratios for photo-initiated free radical copolymerization of VDMA with methacrylic acid (MAA), acrylic acid (AA), acrylamide (AAm), dimethylacrylamide (DMAA), hydroxyethyl methacrylate (HEMA), methoxy poly(ethylene glycol) methacrylate (MPEG 300 MA), and 2-methacryloyloxyethyl phosphorylcholine (MPC), were determined by fitting comonomer conversion data obtained by in situ 1 H NMR to a terminal copolymerization equation. Semi-batch photo-copolymerizations were then used to synthesize the corresponding VDMA copolymers with constant composition. Their solubility and dissolution behavior, as well as their hydrolysis half-lives under physiological conditions, were determined. P(VDMA-co-MAA) copolymers with 52 to 93 mol % VDMA showed decreasing initial solubility and increasing hydrolysis half-lives with increasing VDMA content. VDMA copolymers with nonionic monomers AAm and DMAA were water soluble only at VDMA contents of 41 and 22 mol % or less, respectively, and showed longer hydrolysis half-lives than comparable MAA copolymers. VDMA copolymers with HEMA and MPEG 300 MA were found to crosslink during storage, so their hydrolysis half-lives were not determined. VDMA copolymers with 18% zwitterionic MPC showed a much longer half-life and superior initial solubility compared to analogous p(VDMAco-MAA), identifying this copolymer as a promising candidate for macromolecular crosslinkers in, for example, aqueous layerby-layer co-depositions with polyamines. V C 2012 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 50: [4674][4675][4676][4677][4678][4679][4680][4681][4682][4683][4684][4685] 2012

Calcium alginate/poly-L-lysine beads were coated with either 50% hydrolyzed poly(methyl vinyl ether-alt-maleic anhydride) (PMM(50)), or with poly(vinyl dimethyl azlactone-co-methacrylic acid) (50:50, PMV(50)), to form covalently shell-crosslinked capsules, and compared with analogous capsules coated with sodium alginate. All capsule types were prepared with and without C2C12 murine myoblast cells, and implanted into mice for up to 6 weeks. Cell viability, capsule integrity, fibrotic overgrowth, and mechanical strength of the capsules were assessed, and correlated with inflammatory cytokine marker levels in tail vein blood samples taken at different time points. AP-PMM(50) capsules displayed the least amount of fibrotic overgrowth, were found to be the strongest, and showed the lowest levels of TNF-α in tail vein serum samples taken at 4 h, 24 h, 1 and 6 weeks post transplantation. The results for APA and AP-PMV(50) capsules were more variable and depended on the presence or absence of encapsulated cells.

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