**3. Surface modification of hybrid nanostructures**

Functionalization of the hybrid nanostructures has received a great deal of interest because of their biomedical applications in targeted drug delivery, diagnosis and therapy. The synthesis of the hybrid nanomaterials has mostly been achieved in organic and aqueous media containing either hydrophobic or hydrophilic organic linkers. The hybrid nanostructures synthesized usually involves organic linkers binding to the surface of two or more components to stabilize the nuclei and larger nanoparticles against aggregation by repulsive force for controlled growth of the sized and shaped nanohybrids. As capped by the linkers, the resulting hybrid nanostructures become hydrophobic character and is insoluble in water, resulting in incompatible with biological systems. A surface modification of hydrophobic-surfaced nanohybrids to produce water-soluble functional nanohybrids are therefore a indispensable step prior to biomedical applications. These interactions with the environment ultimately affect the colloidal stability of the particles and may yield to the delivery of the appropriate functional nanoparticles to targeted species. There are common functionalization strategies of silica and amphiphilic polymer coatings. Since the coated hybrid nanostructures retained their original physical properties, the resulting nanocomposites showed the multifunctional properties. These routes can serve as a powerful paradigm for the further fabrication of antibody-conjugated nanohybrids for multifunctional theranostic applications.

#### **3.1. Polymer adsorption**

318 Practical Applications in Biomedical Engineering

seeds.[29]

**3. Surface modification of hybrid nanostructures** 

multifunctional theranostic applications.

nanohybrids with matchstick-like topology was predicted by crystal-oriented-attachment occurred the directional fusion of the generated Co nanocrystals to the nanorod tips. These Co-tipped CdSe/CdS heterostructures exhibited unusual room-temperature ferromagnetism and uorescent emission despite photoexcited charge transfer from the semiconductor to the metal domain. Talapin et al.[27] synthesized FePt/PbS and FePt/PbSe nanohybrids with magneto-transport properties by coupling ferromagnetic FePt particles with either PbS or PbSe in form of core-shells or dumbbells. The formation of the hybrid products by injecting bis(trimethylsilyl) sulfide to reaction mixture containing FePt nanoparticles, oleic acid, Pboleate complex dissolved in octadecene at 120-150 °C. The nanohybrid shape was controlled by capping of the ligand on the surface of the FePt seeds and the reaction temperature. These magnet-in-the-semiconductor hybrid nanostructures showed semiconductor-type transport properties with magnetoresistance characteristic of combining the advantages of both functional components. The maghemite-metal sulphide (ZnS, CdS, HgS) nanohybrids were synthesized by adding sulphur and appropriate metallorganic precursors to the Fe2O3 particles followed by heating treatment.[28] In other reports, the large lattice mismatch between Fe2O3 and metal sulphide resulted in the formation of non-centrosymmetric nanostructures. The formations of trimers and oligomers were observed for ZnS and dimers for CdS and HgS nanocomposites. Alloy-cadmium selenide core-shells including FePt/CdSe, NiPt/CdSe, FePt/CdSe, NiPt/CdSe were synthesized by hydrolysis of cadmium stearate in oleylamine, hexadecylamine/octyl ether, 1,2-hexadecandiol in the presence of alloy

Functionalization of the hybrid nanostructures has received a great deal of interest because of their biomedical applications in targeted drug delivery, diagnosis and therapy. The synthesis of the hybrid nanomaterials has mostly been achieved in organic and aqueous media containing either hydrophobic or hydrophilic organic linkers. The hybrid nanostructures synthesized usually involves organic linkers binding to the surface of two or more components to stabilize the nuclei and larger nanoparticles against aggregation by repulsive force for controlled growth of the sized and shaped nanohybrids. As capped by the linkers, the resulting hybrid nanostructures become hydrophobic character and is insoluble in water, resulting in incompatible with biological systems. A surface modification of hydrophobic-surfaced nanohybrids to produce water-soluble functional nanohybrids are therefore a indispensable step prior to biomedical applications. These interactions with the environment ultimately affect the colloidal stability of the particles and may yield to the delivery of the appropriate functional nanoparticles to targeted species. There are common functionalization strategies of silica and amphiphilic polymer coatings. Since the coated hybrid nanostructures retained their original physical properties, the resulting nanocomposites showed the multifunctional properties. These routes can serve as a powerful paradigm for the further fabrication of antibody-conjugated nanohybrids for The nanohybrid colloids could be soluble in water by adsorbing amphiphilic polymers onto hydrophobic molecules-capped nanohybrids. The amphiphilic polymer adsorption are carried out through hydrophobic interaction of hydrocarbon chains and van der Waals force between the molecules. The desorption of polymer molecules from the nanoparticles is usually prevented because numerous contact points formed from the interactions of organic linkers and polymer chains. Advantage of this approach is mainly not dependent on the types of the inorganic hybrid cores and the organic linkers, and the physiochemical surface properties of the coated particles are significantly unchanged. A popular example is the gold particles in aqueous media prepared by citrate reduction. Citrate ions adsorbed on the gold surface resulted in their negative charge and colloidal stability within several years by electrostatic repulsion. The citrate layer was replaced by stronger-binding ligands of mercaptocarboxylic acids. The surface modification of the particles with mercaptocarboxylic acids allowed for achieving concentrated particle solutions, that can precipitate out of particles by salt-induced aggregation and redissolved in low-salt buffers.[30] The surface functionalization of TOP/TOPO-capped CdSe/ZnS quantum dots was substituted phosphine-based hydrophobic ligands with hydrophilic mercaptocarboxylic acid molecules.[31] The quantum dots in aqueous solution stabilized with mercaptoacetic acid were modified by co-adsorption of polyethylene glycol and peptides.

Polyethylene glycol (PEG) is a linear polymer consisted of ethylene oxide units and well soluble in water. PEG is high biocompatibility due to its inertness and non-toxic properties.[32] Apart from the post-modification approach by covalent chemistry, PEGmodified nanoparticles can be obtained by PEG-contained ligand molecules with functional group that can bind to the particle surface.[33] Owing to the solubility of PEG itself, the PEG-coated particles can also be dispersed in polar organic solvents.[34] **Figure 4** shows the modular design toward The synthesis of the amphiphilic polymer of poly(maleic anhydridealt-1-octadecene) (PMAO)-polyethylene glycol was carried out through reaction between maleic anhydride and primary amine-terminated polyethylene glycol methyl ethers and subsequent coating of PMAO-polyethylene glycol amphiphilic polymer with hydrophobic ligand-capped quantum dots.[35] The functionalized materials dissolved in water had the same optical spectra and quantum yield as those pre-synthesized quantum dots. The watersoluble encapsulated nanoparticles contain free carboxylic acid groups for conjugating anti-Her2 antibody to the polyethylene glycol-coated nanoparticles. The antibody-conjugated quantum dots were used as a probe to recognize human breast cancer cells with Her2 receptor with nonspecific binding of polyethylene glycol with cell receptors on the particle surface. Mattoussi et al.[33] achieved the capping of water-soluble ligand of mixed dihydrolipoic acid (DHLA) and poly(ethylene glycol) (PEG) on quantum dots. These ligands are consist of a poly(ethylene glycol) (PEG) segment attached anchoring DHLA group on one end to drive binding to the quantum dot. Cap exchange with these hydrophilic ligands made quantum dots to be stabilized in water over extended periods of time and over a broad pH range. The quantum dots capped with DHLA-PEG-biotin interacted with streptavidin coupled to proteins which were subsequently taken up by live cells. Yu et al.

Functional Inorganic Nanohybrids for Biomedical Diagnosis 321

Poly(acrylic acid)-based polymers with hydrophobic side chains were usually used for surface modification with aliphatic amine- or thiol-capped particles.[36] These polymers are soluble in organic solvent and can bind to the hydrophobic particle surface. After solvent evaporation, the particle solids can be dissolved in aqueous buffer, providing stable watersoluble particles. Poly(acrylic acid) was coated onto hydrophobic dodecylamine-capped CdTe/CdSe quantum dots to form amphiphilic double-layered nanohybrids soluble in either water or organic media.[37] The coating of these polymers can carry out in ethanol solvent, resulting of poly(acrylic acid) backbone linked with mixed octylamine and isopropylamine, giving numerous hydroxyl groups on the particle surface.[38, 39] The hydrophobic side chains of the polymers commonly cover or intercalate the hydrophobic ligand molecules, and the exposed hydrophilic backbone outwards to aqueous media. In addition, poly(maleic anhydride) copolymers prepared from copolymerization of maleic anhydride with olefin are used as alternating copolymers. In aqueous media, the maleic anhydride rings hydrolyze and open giving two carboxylic groups, which gives access to further functionalization. Each maleic anhydride ring yields a free carboxylic group, indicating that the surface of the polymer-coated particles could be covalently grafted to amino acids for biomolecule

Poly(vinyl pyrilidone) was also used to graft directly on the surface of the particles through one-pot process.[37] The further surface functionalization of the grafted products can be achieved by adding a next layer or exchanging original capping agents. Other polymers contained a mixture of aliphatic side chains and others with primary amines at their ends can bind to the nanoparticle surface through the amino groups.[40] Additionally, poly(acrylic acid) modified with free thiol and amino groups at the ends of the side chains was demonstrated as coating for quantum dots to form a thin shell with little effect on the quantum yield of the coated particles. The hydrophobic-hydrophilic block-copolymers formed in micellar structure dispersible in solvent were also used for the coating of the nanoparticles. The coating by block-copolymer micelles yields the particle aggregates instead of the monodisperse particles that could be suitable for further generation of the

Coating of a cross-linked silica shell to protect the organic agent-capped hybrid cores from external environment is carried out to produce the silica-coated hydrophobic nanohybrids. Coating with silica layer is one of the most widely used methods for surface modification of the inorganic nanoparticles, because the unique properties of the nanoparticles can be preserved by silica shells. After silica coating, the colloids stabilized in aqueous media and have low nonspecific interaction with biosystems and inert silica layer against degradation of optical properties. Silica can also be easily surface modified to link bioconjugators with interesting biofunctionalities. To this goal, water-to-oil microemulsion and Stober sol-gel

conjugation.

multifunctional porous materials.

have generally been achieved for silica coating.

**3.2. Silica coating** 

**Figure 4.** Forming biocompatible and nonaggregated nanocrystals in water using amphiphilic polymers. Top: one-step formation of poly(maleic anhydride-alt-1-octadecene) (PMAO)-polyethylene glycol (PEG) amphiphilic polymers through reaction between maleic anhydride and amino groups. Bottom: schematic structure of water-soluble quantum dots (F stands for a functional group instead of - OCH3, such as -OH, -COOH, -NH2). Quantum dots were encapsulated by PMAO-PEG amphiphilic polymer hydrophobic interaction. Reproduced with permission from ref. [35]. Copyright 2007, American Chemical Society.

[35] prepared amphiphilic polymer of PMAO-PEG through reaction between poly(maleic anhydride-alt-1-octadecene) (PMAO) and primary amine-terminated polyethylene glycol methyl ethers (PEG). The quantum dots were mixed with PMAO-PEG in chloroform. These PMAO-PEG-coated quantum dots were found to have the optical properties and recognized the cancer cells with Her2 receptor.

Poly(acrylic acid)-based polymers with hydrophobic side chains were usually used for surface modification with aliphatic amine- or thiol-capped particles.[36] These polymers are soluble in organic solvent and can bind to the hydrophobic particle surface. After solvent evaporation, the particle solids can be dissolved in aqueous buffer, providing stable watersoluble particles. Poly(acrylic acid) was coated onto hydrophobic dodecylamine-capped CdTe/CdSe quantum dots to form amphiphilic double-layered nanohybrids soluble in either water or organic media.[37] The coating of these polymers can carry out in ethanol solvent, resulting of poly(acrylic acid) backbone linked with mixed octylamine and isopropylamine, giving numerous hydroxyl groups on the particle surface.[38, 39] The hydrophobic side chains of the polymers commonly cover or intercalate the hydrophobic ligand molecules, and the exposed hydrophilic backbone outwards to aqueous media. In addition, poly(maleic anhydride) copolymers prepared from copolymerization of maleic anhydride with olefin are used as alternating copolymers. In aqueous media, the maleic anhydride rings hydrolyze and open giving two carboxylic groups, which gives access to further functionalization. Each maleic anhydride ring yields a free carboxylic group, indicating that the surface of the polymer-coated particles could be covalently grafted to amino acids for biomolecule conjugation.

Poly(vinyl pyrilidone) was also used to graft directly on the surface of the particles through one-pot process.[37] The further surface functionalization of the grafted products can be achieved by adding a next layer or exchanging original capping agents. Other polymers contained a mixture of aliphatic side chains and others with primary amines at their ends can bind to the nanoparticle surface through the amino groups.[40] Additionally, poly(acrylic acid) modified with free thiol and amino groups at the ends of the side chains was demonstrated as coating for quantum dots to form a thin shell with little effect on the quantum yield of the coated particles. The hydrophobic-hydrophilic block-copolymers formed in micellar structure dispersible in solvent were also used for the coating of the nanoparticles. The coating by block-copolymer micelles yields the particle aggregates instead of the monodisperse particles that could be suitable for further generation of the multifunctional porous materials.

#### **3.2. Silica coating**

**Figure 4.** Forming biocompatible and nonaggregated nanocrystals in water using amphiphilic polymers. Top: one-step formation of poly(maleic anhydride-alt-1-octadecene) (PMAO)-polyethylene glycol (PEG) amphiphilic polymers through reaction between maleic anhydride and amino groups. Bottom: schematic structure of water-soluble quantum dots (F stands for a functional group instead of - OCH3, such as -OH, -COOH, -NH2). Quantum dots were encapsulated by PMAO-PEG amphiphilic polymer hydrophobic interaction. Reproduced with permission from ref. [35]. Copyright 2007,

[35] prepared amphiphilic polymer of PMAO-PEG through reaction between poly(maleic anhydride-alt-1-octadecene) (PMAO) and primary amine-terminated polyethylene glycol methyl ethers (PEG). The quantum dots were mixed with PMAO-PEG in chloroform. These PMAO-PEG-coated quantum dots were found to have the optical properties and recognized

320 Practical Applications in Biomedical Engineering

American Chemical Society.

the cancer cells with Her2 receptor.

Coating of a cross-linked silica shell to protect the organic agent-capped hybrid cores from external environment is carried out to produce the silica-coated hydrophobic nanohybrids. Coating with silica layer is one of the most widely used methods for surface modification of the inorganic nanoparticles, because the unique properties of the nanoparticles can be preserved by silica shells. After silica coating, the colloids stabilized in aqueous media and have low nonspecific interaction with biosystems and inert silica layer against degradation of optical properties. Silica can also be easily surface modified to link bioconjugators with interesting biofunctionalities. To this goal, water-to-oil microemulsion and Stober sol-gel have generally been achieved for silica coating.

Reverse microemulsion is a promising method for the synthesis of the monodisperse silicacoated nanoparticles.[41] Ying et al.[42] developed the reverse microemulsion-mediated route to encapsulate the hydrophobic trioctylphosphine oxide (TOPO)-capped quantum dots and magnetic particles within silica shells to form the silica-coated quantum dotmagnetic hybrid structures. **Figure 5** describes the water-in-oil (W/O) reverse microemulsion system for silica coating of the hydrophobic particles, where water droplets are stabilized by nonionic Igepal surfactant in a continuous oil phase (e.g., cyclohexane). After the addition of silane (TEOS), hydrolysis and condensation occur at W/O interface or in water phase to encapsulate the inorganic particles within a silica shell. The magnetic particles and quantum dots were confined in the silica layer to afford the hybrid structure. The silica-coated magnetic-quantum dot nanohybrids preserved the magnetic property of -Fe2O3 and optical property of CdSe quantum dots. The authors also used this route to synthesize silica-coated oleylamine-coated Au and Ag core-shells and subsequent conjugation with activated polymeric dextran.[43] The resulting materials were potentially used as glycobiological Functional Inorganic Nanohybrids for Biomedical Diagnosis 323

probes. Similarly, the SiO2-coated Fe2O3 rattle-type nanoball structures were also synthesized and used as support for decorating the Pd nanoclusters onto the support

The reverse microemulsion process was also used by other research groups. Meijerink et al. [45] elucidated the mechanism of incorporating hydrophobic quantum dots into monodisperse silica spheres. In water-in-oil reverse microemulsion system, the hydrolyzed TEOS had a high affinity for the quantum dot surface for replacement of hydrophobic amine ligands, which enabled the transfer of the quantum dots to the hydrophilic interior of the micelles where silica growth occurred. Serna et al.[46] achieved in-situ synthesis and further silica coating of the Fe nanoparticles in microemulsion system. The lamellar-like coated nanostructures were formed through the subtle interplay controlling the formation of nanospherical silica particles by the ammonia base-catalyzed hydrolysis of tetraethoxysilane (TEOS) in water-in-oil. Kang et al.[47] designed the reverse microemulsion based on the Igepal CO-520 surfactant to produce the silica-coated NiPt nanohybrids prepared from the reduction of nickel acetylacetionate and platinum acetylacetonate in oleic acid/oleylamine. Tsang et al.[48] reported that the templated sol-gel encapsulation of CTAB-stabilized micelles containing metal precursors with ultra-thin porous silica coating allows solvent extraction of organic based stabilizer from silica-coated Ag-Pt alloys. The water-in-oil microemulsion for silica coating on Y3Al5O12:Ce nanoparticles was presented by Chen et al. [49] through hydrolysis of tetraethyl orthosilicate. The silica shell thickness can be turned

The Stober method of base-catalyzed hydrolysis and condensation of tetraethyl orthosilicate (TEOS) to produce silica used to coat on the hybrid cores. [50] This reaction has several advantages such as mild conditions, low cost, without surfactant used. Earlier, Kotov et al.[51] coated hydrophilic CdTe quantum dots within 40-80 nm silica spheres using modified Stober method, which resulted in reduced emission intensity with broadening of the spectrum. The quantum dots acted as seeds for the silica growth in ethanol/water. This method yielded single or multiple quantum dot per silica sphere, but the size and dispersion of the silica-coated quantum dots were hard to control. Alivisatos et al.[52] achieved Stober silanization approach for the functionalization of mercapto-silane/(3-mercaptopropyl-trimethoxysilane) siloxane with thiol and/or amine groups to produce silica shell-coated hydrophobic CdSe/ZnS coreshells. Mercaptopropyltris(methyloxy)silane (MPS) was replaced TOPO molecules on the surface. The methoxysilane groups (Si-OCH3) of (MPS) hydrolyzed into silanol groups (Si-OH), and formed a primary polymerization layer. The silane precursors containing functional groups (F = -SH, -NH2) were then incorporated into the shell and may tailor the nanoparticle surface functionality. Adopting the Stober method, **Figure 6a** shows the seminal silica coating of citrate-reduced Au particles by Mulvaney et al. [53] involved the weak surface attachment with bifunctional (3-aminopropyl) trimethoxysilane in aqueous media. The -NH2 groups were bound to the gold surface and -Si(OEt)3 groups and facilitated for hydrolysis and condensation with sodium silicate to deposit a surface-coated silica layer. Later, the thicker silica shells can be grown on the surface-stabilized Au particles by further hydrolysis/condensation of

surface by mercapto- or amino-functionalized silica.[44]

from 8-16 nm by varying the ratio of NiPt particles to TEOS precursor.

tetraethyl orthosilicate (TEOS).

**Figure 5.** Silica-coated magnetic-quantum dot hybrid nanoparticles. (a) A scheme of the reverse microemulsion system for the synthesis of silica-coated magnetic-quantum dot nanohybrids; (b) TEM image of the silica-coated magnetic-quantum dot nanohybrids. Reproduced with permission from ref. [42]. Copyright 2005, American Chemical Society.

probes. Similarly, the SiO2-coated Fe2O3 rattle-type nanoball structures were also synthesized and used as support for decorating the Pd nanoclusters onto the support surface by mercapto- or amino-functionalized silica.[44]

322 Practical Applications in Biomedical Engineering

Reverse microemulsion is a promising method for the synthesis of the monodisperse silicacoated nanoparticles.[41] Ying et al.[42] developed the reverse microemulsion-mediated route to encapsulate the hydrophobic trioctylphosphine oxide (TOPO)-capped quantum dots and magnetic particles within silica shells to form the silica-coated quantum dotmagnetic hybrid structures. **Figure 5** describes the water-in-oil (W/O) reverse microemulsion system for silica coating of the hydrophobic particles, where water droplets are stabilized by nonionic Igepal surfactant in a continuous oil phase (e.g., cyclohexane). After the addition of silane (TEOS), hydrolysis and condensation occur at W/O interface or in water phase to encapsulate the inorganic particles within a silica shell. The magnetic particles and quantum dots were confined in the silica layer to afford the hybrid structure. The silica-coated magnetic-quantum dot nanohybrids preserved the magnetic property of -Fe2O3 and optical property of CdSe quantum dots. The authors also used this route to synthesize silica-coated oleylamine-coated Au and Ag core-shells and subsequent conjugation with activated polymeric dextran.[43] The resulting materials were potentially used as glycobiological

**Figure 5.** Silica-coated magnetic-quantum dot hybrid nanoparticles. (a) A scheme of the reverse microemulsion system for the synthesis of silica-coated magnetic-quantum dot nanohybrids; (b) TEM image of the silica-coated magnetic-quantum dot nanohybrids. Reproduced with permission from ref.

[42]. Copyright 2005, American Chemical Society.

The reverse microemulsion process was also used by other research groups. Meijerink et al. [45] elucidated the mechanism of incorporating hydrophobic quantum dots into monodisperse silica spheres. In water-in-oil reverse microemulsion system, the hydrolyzed TEOS had a high affinity for the quantum dot surface for replacement of hydrophobic amine ligands, which enabled the transfer of the quantum dots to the hydrophilic interior of the micelles where silica growth occurred. Serna et al.[46] achieved in-situ synthesis and further silica coating of the Fe nanoparticles in microemulsion system. The lamellar-like coated nanostructures were formed through the subtle interplay controlling the formation of nanospherical silica particles by the ammonia base-catalyzed hydrolysis of tetraethoxysilane (TEOS) in water-in-oil. Kang et al.[47] designed the reverse microemulsion based on the Igepal CO-520 surfactant to produce the silica-coated NiPt nanohybrids prepared from the reduction of nickel acetylacetionate and platinum acetylacetonate in oleic acid/oleylamine. Tsang et al.[48] reported that the templated sol-gel encapsulation of CTAB-stabilized micelles containing metal precursors with ultra-thin porous silica coating allows solvent extraction of organic based stabilizer from silica-coated Ag-Pt alloys. The water-in-oil microemulsion for silica coating on Y3Al5O12:Ce nanoparticles was presented by Chen et al. [49] through hydrolysis of tetraethyl orthosilicate. The silica shell thickness can be turned from 8-16 nm by varying the ratio of NiPt particles to TEOS precursor.

The Stober method of base-catalyzed hydrolysis and condensation of tetraethyl orthosilicate (TEOS) to produce silica used to coat on the hybrid cores. [50] This reaction has several advantages such as mild conditions, low cost, without surfactant used. Earlier, Kotov et al.[51] coated hydrophilic CdTe quantum dots within 40-80 nm silica spheres using modified Stober method, which resulted in reduced emission intensity with broadening of the spectrum. The quantum dots acted as seeds for the silica growth in ethanol/water. This method yielded single or multiple quantum dot per silica sphere, but the size and dispersion of the silica-coated quantum dots were hard to control. Alivisatos et al.[52] achieved Stober silanization approach for the functionalization of mercapto-silane/(3-mercaptopropyl-trimethoxysilane) siloxane with thiol and/or amine groups to produce silica shell-coated hydrophobic CdSe/ZnS coreshells. Mercaptopropyltris(methyloxy)silane (MPS) was replaced TOPO molecules on the surface. The methoxysilane groups (Si-OCH3) of (MPS) hydrolyzed into silanol groups (Si-OH), and formed a primary polymerization layer. The silane precursors containing functional groups (F = -SH, -NH2) were then incorporated into the shell and may tailor the nanoparticle surface functionality. Adopting the Stober method, **Figure 6a** shows the seminal silica coating of citrate-reduced Au particles by Mulvaney et al. [53] involved the weak surface attachment with bifunctional (3-aminopropyl) trimethoxysilane in aqueous media. The -NH2 groups were bound to the gold surface and -Si(OEt)3 groups and facilitated for hydrolysis and condensation with sodium silicate to deposit a surface-coated silica layer. Later, the thicker silica shells can be grown on the surface-stabilized Au particles by further hydrolysis/condensation of tetraethyl orthosilicate (TEOS).

In some cases prior to silica coating, the particle surface should be attached with hydrophilic molecules to create the surface-protected nanoparticles stabilized in aqueous media. This could facilitate hydrolysis/condensation of tetraethyl orthosilicate. For example, Han et al. [54] synthesized the monodisperse silica-coated gold particles derived from the citratestabilized gold particles. The prepared citrate-reduced gold particles are low stable for silica coating in alcoholic media. The colloidal stability needs to be increased by introducing a certain amount of sodium citrate into the synthetic solution to replace the surface charge of the gold particles. Chang et al.[55] presented the synthesis of the silica nanohybrids composed of the CuInS2/ZnS quantum dots and magnetite nanocrystals. The outside silica shell grafted with poly(ethyleneglycol) and amine groups to provide better biocompatibility and to allow further bioconjugation. These materials exhibited the exert excellent properties for drug delivery vehicles and magnetic resonance imaging. The conjugation of Pt(IV) anticancer drug onto the nanohybrids resulted in higher cytotoxicity than the free Pt(IV) anticancer drug, indicative of the multifunctional feature of the synthesized nanohybrids.

Functional Inorganic Nanohybrids for Biomedical Diagnosis 325

followed by a dense monolayer of CdSe quantum dots were formed via multistep procedure.[59] The formed products involved the synthesis of the gold particles, gold surface activation, silica-shell deposition, modification of the silica surfaces with -NH2 groups, and final self-assembly of the CdSe quantum dots onto the particle surfaces. In order to the surface activation of the gold particles, (3-mercaptopropyl)-trimethoxysilane was found to be better than (3-aminopropyl)-trimethoxysilane because of stronger binding of -SH groups to the gold surfaces. These hybrid structures were used to perform the accurate quantitative analysis of the effect of the metal on quantum dot photoluminescence intensity. Khlebtsov et al.[60] preformed the silica coating on Au-Ag nanocages through adding water-ammonia solution and TEOS to the reaction solution containing Au-Ag particles. Silica-coated Au-Ag nanocages were then functionalized with photodynamic sensitizer Yb-2,4-dimethoxyhematoporphyrin to form the nanocomposites potential in multifunctional capability of IR-luminescence detection, photosensitization, and

Cancer is the greatest challenge in human healthcare today. It is a result of unregulated cell division leading to the uncontrolled growth and spread of abnormal cells. This behaviour causes the formation of malignant tumors consisted of cancer cells plus some healthy cells (normal tissue) invade nearby parts of the body. At early growth stages, the cancer cells mostly do not look or act like the normal cells because they are readily disguised by the healthy cells on their surface. This mainly behaves an extremely danger of the cancer cells. Tumor cells have a strong tendency to displace healthy cells until the tumor reaches a diffusion-limited maximal size, frequently resulting of changes to the DNA (mutations),

Traditional cancer diagnosis and treatment modalities basically include post-surgical chemotherapy, radiotherapy, hormone therapy, and immunotherapy.[61] Each of these modalities has constantly limitations in treatment and also contribute to the rising costs of healthcare. Because of most human cancers relevant to solid tumors, so that the current cancer therapies are usually achieved some surgeries for removal of tumors, followed by chemotherapy and radiotherapy to kill the remaining tumor cells. However, the efficacy of the chemotherapy, serious side-effects on different healthy organs, the increased costs are a great obstruction by the fact that cancer stem cells be still survive and could continue to spread back. It is reasonably why cancer symptoms come back within relative short

Thermal therapies (hyperthermia) have often employed a variety of heat sources including laser light, focused ultrasound, microwaves to destroy the solid tumors.[62] The benefits of hyperthermia are minimally or non-invasive, relatively simple to perform in the absence of surgical resection. However, simple heating techniques have trouble discriminating between tumors and surrounding healthy tissues, and often heat intervening tissue between source and target site. To irradiating beams reached underlying tumors or dispersed into large

duration in patients who has passed through the post-surgical chemotherapy.

photothermolysis.

leading to deaths.

**4. Nanotechnology in cancer treatments** 

**Figure 6.** Schemes of silica coatings of hydrophilic (a) and hydrophobic (b) nanoparticles.

**Figure 6b** shows a successful example involved the surface adsorption of methoxypoly(ethylene glycol) silane to replace oleylamine capped on silver nanoparticles by Yang et al.[56] The functionalized silver nanoparticles were then hydrolyzed and condensed further to form thin silica layer-stabilized silver nanoparticles followed by thick silica coating with the Stober process. Bifunctional Gd2O(CO3)2·H2O/silica/Au hybrid nanoparticles prepared by condensation of TEOS followed by conjugation with the gold shells were demonstrated potential as a MRI and therapeutic agent.[57] The hybrid particles showed the capability of absorbing NIR radiation for photothermal destruction of cancer tumors, in which the Au shell thickness strongly influenced the NIR optical absorption and photothermal effect. Pratsinis et al.[58] achieved coating of a thin silica shell on Ag/Fe2O3 Janus-shaped hybrids via one-step flame aerosol method. The silica coating still intacted their shape and plasmonic-magnetic properties but minimizes the release of toxic Ag+ ions from the Ag particle surface and their direct contact with live cells. Well-defined Au/SiO2/CdSe hybrid nanostructures constituted a gold core overcoated with a silica shell followed by a dense monolayer of CdSe quantum dots were formed via multistep procedure.[59] The formed products involved the synthesis of the gold particles, gold surface activation, silica-shell deposition, modification of the silica surfaces with -NH2 groups, and final self-assembly of the CdSe quantum dots onto the particle surfaces. In order to the surface activation of the gold particles, (3-mercaptopropyl)-trimethoxysilane was found to be better than (3-aminopropyl)-trimethoxysilane because of stronger binding of -SH groups to the gold surfaces. These hybrid structures were used to perform the accurate quantitative analysis of the effect of the metal on quantum dot photoluminescence intensity. Khlebtsov et al.[60] preformed the silica coating on Au-Ag nanocages through adding water-ammonia solution and TEOS to the reaction solution containing Au-Ag particles. Silica-coated Au-Ag nanocages were then functionalized with photodynamic sensitizer Yb-2,4-dimethoxyhematoporphyrin to form the nanocomposites potential in multifunctional capability of IR-luminescence detection, photosensitization, and photothermolysis.

### **4. Nanotechnology in cancer treatments**

324 Practical Applications in Biomedical Engineering

In some cases prior to silica coating, the particle surface should be attached with hydrophilic molecules to create the surface-protected nanoparticles stabilized in aqueous media. This could facilitate hydrolysis/condensation of tetraethyl orthosilicate. For example, Han et al. [54] synthesized the monodisperse silica-coated gold particles derived from the citratestabilized gold particles. The prepared citrate-reduced gold particles are low stable for silica coating in alcoholic media. The colloidal stability needs to be increased by introducing a certain amount of sodium citrate into the synthetic solution to replace the surface charge of the gold particles. Chang et al.[55] presented the synthesis of the silica nanohybrids composed of the CuInS2/ZnS quantum dots and magnetite nanocrystals. The outside silica shell grafted with poly(ethyleneglycol) and amine groups to provide better biocompatibility and to allow further bioconjugation. These materials exhibited the exert excellent properties for drug delivery vehicles and magnetic resonance imaging. The conjugation of Pt(IV) anticancer drug onto the nanohybrids resulted in higher cytotoxicity than the free Pt(IV) anticancer drug, indicative of the multifunctional feature of the synthesized nanohybrids.

**Figure 6.** Schemes of silica coatings of hydrophilic (a) and hydrophobic (b) nanoparticles.

**Figure 6b** shows a successful example involved the surface adsorption of methoxypoly(ethylene glycol) silane to replace oleylamine capped on silver nanoparticles by Yang et al.[56] The functionalized silver nanoparticles were then hydrolyzed and condensed further to form thin silica layer-stabilized silver nanoparticles followed by thick silica coating with the Stober process. Bifunctional Gd2O(CO3)2·H2O/silica/Au hybrid nanoparticles prepared by condensation of TEOS followed by conjugation with the gold shells were demonstrated potential as a MRI and therapeutic agent.[57] The hybrid particles showed the capability of absorbing NIR radiation for photothermal destruction of cancer tumors, in which the Au shell thickness strongly influenced the NIR optical absorption and photothermal effect. Pratsinis et al.[58] achieved coating of a thin silica shell on Ag/Fe2O3 Janus-shaped hybrids via one-step flame aerosol method. The silica coating still intacted their shape and plasmonic-magnetic properties but minimizes the release of toxic Ag+ ions from the Ag particle surface and their direct contact with live cells. Well-defined Au/SiO2/CdSe hybrid nanostructures constituted a gold core overcoated with a silica shell Cancer is the greatest challenge in human healthcare today. It is a result of unregulated cell division leading to the uncontrolled growth and spread of abnormal cells. This behaviour causes the formation of malignant tumors consisted of cancer cells plus some healthy cells (normal tissue) invade nearby parts of the body. At early growth stages, the cancer cells mostly do not look or act like the normal cells because they are readily disguised by the healthy cells on their surface. This mainly behaves an extremely danger of the cancer cells. Tumor cells have a strong tendency to displace healthy cells until the tumor reaches a diffusion-limited maximal size, frequently resulting of changes to the DNA (mutations), leading to deaths.

Traditional cancer diagnosis and treatment modalities basically include post-surgical chemotherapy, radiotherapy, hormone therapy, and immunotherapy.[61] Each of these modalities has constantly limitations in treatment and also contribute to the rising costs of healthcare. Because of most human cancers relevant to solid tumors, so that the current cancer therapies are usually achieved some surgeries for removal of tumors, followed by chemotherapy and radiotherapy to kill the remaining tumor cells. However, the efficacy of the chemotherapy, serious side-effects on different healthy organs, the increased costs are a great obstruction by the fact that cancer stem cells be still survive and could continue to spread back. It is reasonably why cancer symptoms come back within relative short duration in patients who has passed through the post-surgical chemotherapy.

Thermal therapies (hyperthermia) have often employed a variety of heat sources including laser light, focused ultrasound, microwaves to destroy the solid tumors.[62] The benefits of hyperthermia are minimally or non-invasive, relatively simple to perform in the absence of surgical resection. However, simple heating techniques have trouble discriminating between tumors and surrounding healthy tissues, and often heat intervening tissue between source and target site. To irradiating beams reached underlying tumors or dispersed into large tumors, high activating energy source must achieve at long duration of time, leading to sufficiently penetrate and damage healthy tissues.

Functional Inorganic Nanohybrids for Biomedical Diagnosis 327

therapy through the selective localized photothermal heating of the cancer tumors.[65] To treat a tumor, the gold particles conjugated with biomolecules can be selectively targeted to cancer cells without significant binding to healthy cells. The nanoparticles in the bloodstream generally have to firstly move across the tumor blood vessels. The tumors are then exposed to an excitation source, such as NIR laser light, radiowave, or an alternating magnetic field. When the gold nanoparticles are exposed to the light radiation at their resonance wavelength, the electric field of light causes the collective oscillation of the conduction-band electrons at the particle surface. The coherent oscillation of the metal free electrons in resonance with the electro-magnetic field is called the surface plasmon resonance (SPR). The excitation of the maximum SPR absorption results in enhancement of the photophysical properties of gold particles.[66] The gold nanoparticles absorb the incident energy and convert them to heat, which raises the temperature (~42°C) of the tissue and ablates the cancerous cells by disrupting the cell membrane. The photoradiations do not often kill healthy cells because the laser power requires to heat/destroy the cancer cells much low than the healthy cells to which nanoparticles do not bind specifically. The physical heating mechanism of ablative therapies would provide an advantage against chemotherapy-resistant cancers, as well as improved tumor response when combined with

Key features to consider when selecting a compatible particle for hyperthermia are the wavelength of maximal absorption, absorption cross-section, and shape/size of the particle. NIR laser light is ideal for in-vivo hyperthermia applications because of its low absorption by tissue chromophores (hemoglobin and water), which prevents them from damaging healthy tissue. The absorption coefficient of these tissue chromophores is as much as two orders of magnitude greater in the visible region (400-600 nm) as compared to the NIR region (650-900 nm).[66] Gold-nanoparticle-mediated photothermal therapy is predominantly designed to operate in this window of wavelengths ("NIR window") to minimize energy interaction of light-tissue, preventing damaging heating of healthy tissue. Upon tumor laser irradiation, NIR light is absorbed by the nanoparticles and heat dissipation is generated as a consequence of electron-phonon interactions. For successful cancer ablation, the tissue must be heated to a minimum temperature for a minimum

The plasmon absorbance of the gold particles can be easily tuned from the visible region into the NIR by simple manipulation of their aspect ratio (from sphere to rod).[66] For the gold nanospheres, this resonance occurs in the visible spectral region at about 520 nm, originating from the brilliant colour of the gold particle solution. Owing to their distinctive rod shape, the gold nanorods have two absorption peaks attributed to the free electron oscillation along the longitudinal and transverse axis, resulting in a stronger resonance band in the NIR region and a weaker band in the visible region (~520 nm for gold nanospheres). The synthesis of the colloidal gold nanorods would therefore prove effectively for photothermal therapy because

they can absorb low-energy NIR light and convert it to heat in the usual way.

chemotherapy and photoradiation.

duration of time to induce tumor cell death.

**Figure 7.** A scheme of thiolated DNA conjugation onto the silica-coated nanoparticles.

To the goal of the cost and performance, the development of new efficient approaches based on an advanced combination between "smart drug delivery" chemotherapy and photoradiation companied by "near-infrared (NIR) laser-adsorbing nanomaterials" to create the most effective results have been interested in medicine technology.[63] Lack of target specificity is one of the major disadvantages of many drugs. When drugs administered into human body are distributed to all organs through bloodstream, rather than to specific target organ that needs the pharmacological treatment. Biochemical and physiological barriers of certain organs also limit drug delivery to the desired organ. Chemotherapeutic drugs may destroy the cancer cells along with destroy the healthy tissue and cytotoxic effect of the drugs. To overcome these disadvantages, newer and effective methods should be developed to safely shepherd a pharmacological agent to avoid specific organs, where healthy tissue might be adversely affected.

The nanoparticles with the size smaller 100-10,000 times than the cells can be conjugated with various complementary biomolecules including DNA strands and antigens, as shown in **Figure 7**. The conjugated nanoparticles can easily pass through the cell membrane and accumulate into target sites by manipulation, which is advantageous in targeted imaging, diagnosis, and delivery.[64] The functionalization of the silica-coated nanohybrids is usually achieved by adsorption or chemical conjugation of the biomolecules to the particle surface. The silica-coated gold-based nanohybrid colloids can be surface-functionalized with mercapto-, amino-, carboxy-terminated silanes for biomolecule conjugation. Homogeneously water-dissolved biomolecules-conjugated silica-coated nanoparticles could bind to the surface of the cancer cells with greater affinity than to the noncancerous cells.
