Synthesis of uniform, fluorescent poly(glycidyl methacrylate) based particles and their characterization by confocal laser scanning microscopy

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Abstract

Uniform poly(glycidyl methacrylate) (poly(GMA)) particles with different size and surface properties were obtained by dispersion polymerization formulations including different initiator and stabilizer types. The particle size was correlated with the half-life of the initiator. The use of poly(acrylic acid) (PAA) as a novel steric stabilizer in the dispersion polymerization of GMA resulted in the synthesis of smaller particles with higher surface charge density with respect to those in the literature. As a new initiator–stabilizer system for GMA, 4,4′-azobis(4-cyanovaleric acid)–poly(acrylic acid) provided the synthesis of bifunctional uniform latex particles carrying epoxide and carboxyl groups.

The reactive character of poly(GMA) particles allowed the preparation of their primary amine carrying form. The fluorescent form of the amine carrying poly(GMA) based latex particles was first obtained by labeling with FITC and characterized by confocal laser scanning microscopy (CLSM). A CLSM method was developed to determine the amine group distribution in the particles based on the fluorescent intensity profile. A core part with low primary amine content was observed in the particle interior while the amine group concentration was high on the shell part located on the particle surface. The photobleaching time of poly(GMA) based particles was found to be two-fold longer than the highest value reported for the uniform particles. Low photobleaching rate is extremely important for the CLSM applications of uniform particles particularly in microfluidics and biomedical area.

Introduction

In recent years, significant attention was paid on the production of uniform poly(glycidyl methacrylate), poly(GMA) microspheres by dispersion polymerization [1], [2], [3], [4], [5], [6], [7], [8], [9], [10], [11], [12], [13], [14], [15]. Zhao et al. proposed a dispersion polymerization process for the production of poly(GMA) microspheres in the size range of 1–8 μm [1]. An alternative dispersion polymerization process involving the use of polyvinylpyrrolidone, hydroxypropylcellulose or cellulose acetate butyrate as the stabilizer in dimethylformamide–alcohol mixture was developed [2]. Recently, two different methods were proposed for the dispersion polymerization of GMA in supercritical CO2 [3], [4].

Dispersion copolymerization is the another tool for the preparation of GMA functionalized uniform particles [5], [6], [7], [8], [9], [10]. The effects of various polymerization conditions on the particle size were investigated in the dispersion copolymerization of GMA with styrene or methylmethacrylate [5], [6]. Uniform polystyrene/GMA-divinylbenzene core/shell composite particles with epoxy groups in the shells were obtained by the seeded dispersion copolymerization in ethanol–water medium including polystyrene seed particles [7]. The dispersion copolymerization of various monomethacrylate monomers with GMA was achieved in a mixture of two organic solvents heptane and propanol including a reactive surfactant and a diacrylate crosslinker [8]. Recently, monodisperse poly(GMA) microspheres crosslinked with divinylbenzene were synthesized by a single-stage dispersion copolymerization [9]. In another study, the effects of reaction parameters on the particle size in the dispersion copolymerization of 2-hydroxyethyl methacrylate and GMA were investigated in the presence of a ferrofluid [10]. Epoxy functionalized polystyrene/silica core–shell composite nanoparticles were synthesized by post addition of GMA via emulsion polymerization [11].

Uniform poly(GMA) particles are suitable support materials in different chromatographic applications. Monodisperse-porous poly(GMA) beads suitable for gel permeation chromatography were obtained by a multistage polymerization [12]. Porous GMA based microspheres 3 μm in size prepared by single-stage dispersion polymerization exhibited a good separation performance in capillary electrochromatography [13], [14]. The nucleotide immobilized GMA–styrene copolymer particles prepared by dispersion polymerization were successfully used in the affinity-HPLC of DNA [15], [16]. A weak cation exchange stationary phase synthesized by starting from uniform-porous GMA particles was used as chromatographic support for affinity-HPLC and enantiomeric separation [17], [18], [19].

Although a considerable research effort was spent on the production of GMA carrying micron-size, monodisperse particles, there were still some gaps particularly regarding the synthesis of poly(GMA) particles with different surface characteristics. By considering the studies cited above, we selected three different initiators (i.e. 2,2′-azobis(isobutyronitrile), AIBN; 4,4′-azobis(4-cyanovaleric acid), ACVA; 2,2′-azobis(2-amidinopropane)dihyrochloride, AMPA) and two different steric stabilizers (i.e. polyvinylpyrrolidone, PVP and polyacrylic acid, PAA) for the preparation of uniform poly(GMA) particles with the size and surface properties different than those in the literature. Additionally, fluorescent derivatives of poly(GMA) based particles were first synthesized and characterized by confocal laser scanning microscopy (CLSM). A CLSM method was developed to determine the reactive group distribution on the particles.

Section snippets

Materials

The monomer, glycidyl methacrylate (GMA) was supplied from Aldrich Chem. Co., USA, and used without further purification. Technical ethanol (Birpa AS, Turkey, 96%) was used as continuous medium. The initiators, 2,2′-azobisizobutyronitrile (AIBN, BDH Chem. Ltd., Poole, England), 4,4′-azobis(4-cyanovaleric acid) (Aldrich Chem. Co.) 2,2′-azobis(2-methylpropionamidine) dihydrochloride (Aldrich Chem.) were used without further purification. One of the stabilizers, poly(acrylic acid) (PAA) was

Results and discussion

Poly(GMA) is a reactive polymer that can be converted easily into various types polymers carrying different functional groups. The polymeric particles in a wide polarity spectrum carrying hydroxyl, aryl groups or primary, secondary, tertiary and quaternary amine forms can be obtained by relatively simple derivatization protocols by starting from the poly(GMA) particles [2]. The types of functional groups on the particle surfaces and the surface charge density are the factors strongly affecting

Conclusion

The uniform poly(GMA) particles with different size and surface properties were synthesized by dispersion polymerization by using various initiator–stabilizer systems. The use of poly(acrylic acid) as a new stabilizer in the dispersion polymerization of GMA provided relatively smaller uniform latex particles carrying both epoxide and carboxyl groups. The primary amine functionalized form of the poly(GMA) particles was also synthesized and labeled with a fluorescent probe, FITC. The localization

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