**2.2 Polymer coatings**

284 Biomedical Science, Engineering and Technology

Multifunctional MNPs (MFMNPs) are a major class of nanoscale materials with the potential to revolutionize clinical diagnostic and therapeutic techniques. Due to their unique magnetic properties and ability to function at the cellular and molecular levels of biological interactions, MFMNPs have been investigated as an attractive nanomedical platform. MFMNPs in the form of SPIOs have been actively investigated as contrast enhancement agents for magnetic resonance imaging (MRI) and hyperthermia in response to an external alternating magnetic field, due to their ability to enhance the proton relaxation of specific tissues. MFMNPs have been evaluated extensively as a nanomedical platform for the targeted delivery of pharmaceuticals through magnetic drug targeting (Neuberger et al., 2005) and through the attachment of high-affinity ligands (Zhang et al., 2002; Torchilin,

Iron oxides with a core/shell structure are widely used as sources of MFMNP platforms. Iron oxides have several crystalline polymorphs, but only γ-Fe2O3 (maghemite) or Fe3O4 (magnetite) can be used for biomedical applications. These particles, which range in diameter from about 5–20 nm, have unique advantages, including (1) superparamagnetic behavior, with no magnetism after removal of the magnetic field; (2) high saturation magnetization values and high magnetic susceptibility, for effective magnetic enrichment; (3) biocompatibility and rapid removal through extravasation and renal clearance; and (4)

Iron oxide nanoparticles have a significant tendency to agglomerate as a result of their high surface energy. Massart (1981) first prepared stable aqueous dispersions of Fe3O4 nanoparticles (ferrofluids) that were stabilized by electrical double layers. However, the colloidal electrostatic stabilization arising from surface charge repulsion on the nanoparticles typically is inadequate to prevent aggregation in biological solutions, due to the presence of salts or other electrolytes that can neutralize the charges. Furthermore, the iron oxide surfaces may be subjected to plasma protein adsorption or opsonization, leading to their rapid clearance by the reticuloendothelial system (RES) (Berry & Curtis, 2003). To solve the above problems, proper surface coatings have been exploited as an integral component of the MFMNP platform for biomedical applications. The iron oxide core can be coated by organic materials [e.g., polymers such as dextrant (Thorek et al., 2006) and polyethylene glycol (PEG) (Gref et al., 1994)], inorganic metallic materials [e.g., gold (Ji et al., 2007)], or oxides [e.g., silica or alumina (Bumb et al., 2008)]. Polymer coatings will be introduced in detail in the next section. Silica shells are attractive as protective coatings on the iron oxide core, due to their stability under aqueous conditions and ease of synthesis. Recently, Ma et al. (2006) described one such core–shell MFMNP, composed of an iron oxide core (approximately 10 nm diameter) surrounded by a SiO2 shell (10–15 nm thick). They doped an organic dye, tris(2,2′-bipyridine) ruthenium, inside a second silica shell to provide luminescence and prevent quenching by interaction with the magnetic core. As a core–shell structure exhibiting superparamagnetic and luminescent properties, this MFMNP platform

can be used as a multifunctional imaging agent for biomedical applications.

Gold offers several advantages as a coating material for iron oxide cores, due to its low chemical reactivity and unique ability to form self-assembled monolayers on the core surface using alkanethiols (Prime & Whitesides, 1991). A variety of methods (reversed

**2. Surface coatings and functionalization of MNPs** 

easily tailored surface chemistry and functionalization.

2006).

**2.1 Core–shell structure** 

Polymers comprise some of the most important materials used as shells. Polymer coatings not only provide a steric barrier to prevent nanoparticle agglomeration, but also allow MNPs to evade uptake by the RES and thereby to maintain a long plasma half-life. Polymer coatings provide a means to tailor the surface properties of MNPs, such as the surface charge and chemical functionality. An ideal polymer coating will have a high affinity for the iron oxide core, as well as nonimmunogenic and nonantigenic properties. It also will prevent opsonization by plasma proteins. Polymer materials comprised of lipids, proteins, dendrimers, gelatin, dextran, chitosan, pullulan, PEG, poly(ethylene-co-vinyl acetate), poly(vinylpyrrolidone), poly(vinyl alcohol) (PVA), or poly(glycerol monoacrylate) (PGA) are often chosen as the surface coatings for MNPs.

PEG is the most widely used polymer for nanoparticle coating in biomedical applications. PEG provides a very attractive combination of properties: excellent solubility in aqueous solutions; high flexibility of its polymer chain; very low toxicity, immunogenicity, and antigenicity; lack of accumulation in the RES cells; and minimal influence on the specific biological properties of modified pharmaceuticals (Yamaoka et al., 1994). As a so-called "stealth" surface, PEG prevents the nanomedical platform from being recognized by RES, and thereby extends its blood circulation time in vivo. On the biological level, coating nanoparticles with PEG sterically hinders the interaction of blood components with the nanoparticle surface and reduces the binding of plasma proteins. Mechanisms of preventing opsonization by PEG include the shielding of the surface charges, increased surface hydrophilicity (Gabizon & Papahadjopoulos, 1992), and enhanced repulsive interaction between polymer-coated nanoparticles and blood components (Needham et al., 1992). Various methods have been utilized to attach PEG to the MNP surface, including silane grafting to the oxide surface (Butterworth et al., 2001), alkaline coprecipitation of ferric and ferrous ions in the presence of PEG-containing block copolymers (Wan et al., 2005), direct attachment of PEG-containing block copolymers (Guo et al., 2010), polymerization at the MNP surface (Flesch et al., 2005), and modification through sol-gel approaches (Y. Lu et al., 2002).

Polysaccharide dextran is another polymer coating that has been used widely and successfully in vivo. Dextran-coated iron oxide nanoparticles have become an important part of clinical cancer imaging, and have been shown to increase the accuracy of cancer nodal staging (Harisinghani & Weissleder, 2004; Ferrari, 2005). Because the dextran coating is not strongly associated with the iron oxide core, the polymer is susceptible to detachment. Accordingly, cross-linked iron oxide nanoparticles have been developed by cross-linking the dextran shell with epichlorohydrin (Josephson et al., 1999). The resulting particle offers superb stability under harsh conditions, without causing any change in size or blood halflife or loss of the dextran coat. Chemical functionality can be established by treating crosslinked iron oxide nanoparticles with ammonia to provide primary amino groups for the attachment of biomolecules such as proteins or peptides (Wunderbaldinger et al., 2002; Schellenberger et al., 2002). These formulations of dextran-coated iron oxide nanoparticles have been evaluated extensively for a variety of MRI applications (Josephson et al., 1999).

In addition to the traditional polymer coatings, a new kind of biocompatible polymer material has been reported by Wan et al. (2005): namely, homopolymers of glycerol monoacrylate or glycerol monomethacrylate, or their block copolymers. Highly stable aqueous magnetic fluids were prepared by coating Fe3O4 nanoparticles with poly(glycerol monoacrylate) (PGA), poly(glycerol monomethacrylate) (PGMA), or diblock copolymers with PGA or PGMA segments. As shown in Fig. 1, the proposed mechanism of stabilization was the multidentate interactions of 1,2-diols on the polymer chain with iron atoms at the surface of the iron oxide nanoparticles (Wan et al., 2005). This process was a good choice for the preparation of stable magnetic fluids with tailored surfaces; PGA or PGMA binds very tightly to the iron oxide surface, is highly hydrophilic, and does not introduce charges on the surface. Moreover, various block copolymers containing PGA or PGMA can be used to modify the iron oxides and to introduce tailored functional groups for further functionalization.

Fig. 1. Proposed structure in the interaction between the iron oxide surface and PGA (Wan et al., 2005).

### **2.3 Functional ligands**

As discussed in the above sections, the core–shell structure of MFMNPs provides a means to tailor the nanoparticle surface properties, such as surface charge and chemical functionality. Various functional ligands, including targeting agents, permeation enhancers, optical dyes, and therapeutic agents, can be conjugated on the surface or incorporated within the nanostructure. The modification of the nanoparticle surface with targeting ligands was described recently as a promising biotargeting strategy. To generate target-specific nanoparticles, various biological molecules, such as antibodies, proteins, small molecular targeting agents, etc., can be bound to the coating surfaces of the MFMNPs by chemical coupling. Tumor cells are rapidly proliferating and overexpress certain receptors that lead to the enhanced uptake of nutrients, including folic acid, vitamins, sugars, and proteins. MFMNPs conjugated with these molecules can be targeted to tumor cells that overexpress the corresponding receptors. Table 1 summarizes a number of different ligands and their corresponding functions that have been investigated for the in vivo targeting of MFMNPs.

attachment of biomolecules such as proteins or peptides (Wunderbaldinger et al., 2002; Schellenberger et al., 2002). These formulations of dextran-coated iron oxide nanoparticles have been evaluated extensively for a variety of MRI applications (Josephson et al., 1999). In addition to the traditional polymer coatings, a new kind of biocompatible polymer material has been reported by Wan et al. (2005): namely, homopolymers of glycerol monoacrylate or glycerol monomethacrylate, or their block copolymers. Highly stable aqueous magnetic fluids were prepared by coating Fe3O4 nanoparticles with poly(glycerol monoacrylate) (PGA), poly(glycerol monomethacrylate) (PGMA), or diblock copolymers with PGA or PGMA segments. As shown in Fig. 1, the proposed mechanism of stabilization was the multidentate interactions of 1,2-diols on the polymer chain with iron atoms at the surface of the iron oxide nanoparticles (Wan et al., 2005). This process was a good choice for the preparation of stable magnetic fluids with tailored surfaces; PGA or PGMA binds very tightly to the iron oxide surface, is highly hydrophilic, and does not introduce charges on the surface. Moreover, various block copolymers containing PGA or PGMA can be used to modify the iron oxides and to introduce tailored functional groups for further

Fig. 1. Proposed structure in the interaction between the iron oxide surface and PGA (Wan

As discussed in the above sections, the core–shell structure of MFMNPs provides a means to tailor the nanoparticle surface properties, such as surface charge and chemical functionality. Various functional ligands, including targeting agents, permeation enhancers, optical dyes, and therapeutic agents, can be conjugated on the surface or incorporated within the nanostructure. The modification of the nanoparticle surface with targeting ligands was described recently as a promising biotargeting strategy. To generate target-specific nanoparticles, various biological molecules, such as antibodies, proteins, small molecular targeting agents, etc., can be bound to the coating surfaces of the MFMNPs by chemical coupling. Tumor cells are rapidly proliferating and overexpress certain receptors that lead to the enhanced uptake of nutrients, including folic acid, vitamins, sugars, and proteins. MFMNPs conjugated with these molecules can be targeted to tumor cells that overexpress the corresponding receptors. Table 1 summarizes a number of different ligands and their corresponding functions that have been investigated for the in vivo targeting of MFMNPs.

functionalization.

et al., 2005).

**2.3 Functional ligands** 


Table 1. Selected functional ligands used for MFMNPs in biomedical applications

Organic dyes or fluorophores have been loaded on MNPs as optical imaging agents to allow detection by multiple imaging modalities. In addition to their use as contrast enhancement agents, FITC- (Zhang et al., 2002), rhodamine- (Bertorelle et al., 2006), or other fluorophorelabeled MNPs can be used for the in vitro fluorescent imaging of cells. Since both MRI and optical signals come from the same nanoparticles, the MR image can serve as a roadmap to the fluorescently labeled tumor cells. The conjugation of near-infrared fluorescent (NIRF) dyes to MNPs has received recent attention due to the deep penetration of NIRF light in the tissues (Weissleder & Ntziachristos, 2003). The integration of NIRF detectability allows for these nanoparticles to be used for presurgical planning by MRI and intraoperative resection of malignant tissues by optical imaging.
