Dual mode bioreactions on polymer nanoparticles covered with phosphorylcholine group
Introduction
Polymer nanoparticles are widely used in the life science fields for separation technologies, histological studies, clinical diagnostic systems and drug delivery [1], [2], [3]. We are continuously investigating the preparation of polymer nanoparticles covered with phosphorylcholine (PC) groups to obtain excellent bio/blood compatibility and stability in an aqueous medium including plasma [4]. To cover the surface of the polymer nanoparticles, we prepared a water-soluble amphiphilic phospholipid polymer, poly(2-methacryloyloxyethyl phosphorylcholine (MPC)-co-n-butyl methacrylate (BMA)) (PMB) [5]. Since the PMB formed a polymer aggregate in the aqueous medium, it functioned as a good solubilizer for hydrophobic compounds. Thus, we could prepare polymer nanoparticles by solvent evaporation and interfacial precipitation techniques from an organic solvent containing a core polymer in aqueous medium containing the PMB. Moreover, the introduction of active ester units to the PMB was made possible for reactions with biomolecules [6]. We conjugated biomolecules such as a protein enzyme or antibody on the polymer nanoparticles and revealed the good performance of these biomolecules even if they were located on the solid surface [7], [8].
Another viewpoint of biorecognition between the MPC polymer and living cells has been reported. On the cell membrane, carbohydrate and polysaccharide chains play an important role in molecular recognition from the outer medium and signal transport into the cells [9]. The incorporation of unnatural carbohydrates provides an opportunity to study the specific contributions of sialic acid and its N-acyl side chains to the sialic acid-dependent ligand–receptor interactions at a submolecular level. The MPC polymer surfaces with hydrazide groups, which can selectively react with unnatural ketone-containing carbohydrate as a cell surface tag, controlled the cell attachment [10].
From these fundamental research results, we propose novel diagnostic and medical treatment systems using polymer nanoparticles, that is, the polymer nanoparticles can selectively bind target cells, and enzymes conjugated on the polymer nanoparticles react with specific polysaccharide chains on the cell membrane. If the specific polysaccharide chains are digested by the enzymatic reaction, the cell cannot survive. In this study, the preparation of polymer nanoparticles covered with the PC groups and double bioconjugation with the antibody and enzyme on one polymer nanoparticle was carried out. Dual mode bioreactions, that is, aggregation of the polymer nanoparticles by the addition of an antigen and enzymatic reaction by the addition of an enzyme substrate were investigated.
Section snippets
Synthesis of the MPC polymer
MPC was synthesized by a previously reported method [11]. BMA was reagent grade and used after vacuum distillation (bp 68.5 °C/32 mmHg). p-Nitrophenyloxycarbonyl polyethyleneglycol methacrylate (MEONP) was synthesized by a previously reported method [6]. Poly(MPC-co-BMA-co-MEONP) (PMBN) was synthesized by a conventional radical polymerization technique using 2,2′-azobisisobutyronitrile (AIBN) as an initiator [12]. The polymerization was carried out at 60 °C for 5 h. The reaction mixture was then
Characterization of PMBN
The PMBN was synthesized using a conventional radical polymerization technique. The characterizations of PMBN are summarized in Table 1. The compositions of each monomer unit in the polymer were in good agreement with their compositions in the feed. The obtained PMBN was water-soluble, but had an amphiphilic nature because the hydrophobic BMA units were introduced and its composition was above 60 mol%. In our previous article, we reported other PMBNs with different compositions. Based on this
Conclusion
We prepared novel polymer nanoparticles which can conjugate protein including an antibody and enzyme. After conjugation, the proteins worked well and we observed dual mode bioreactions against the target molecules. Thus, we concluded that the polymer nanoparticles could be used to make a new diagnostic system and a medical treatment system.
Acknowledgements
The present research was supported in part by a Grant for the 21st Century COE Program “Human-Friendly Materials based on Chemistry” from the Ministry of Education, Culture, Sports, Science and Technology of Japan and a Grant-in-Aid for Scientific Research from the Japan Society for the Promotion of Science (16650098).
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