Superparamagnetic Nanoparticles For Biomedical Applications · Magnetic silica microspheres are...

Superparamagnetic Nanoparticles For

Biomedical Applications

Suk Fun Chin (MSc)

This thesis is presented for the degree of Doctor of Philosophy at The

University of Western Australia

School of Biomedical, Biomolecular and Chemical Sciences

Discipline of Chemistry

2009

i

Abstract

In the past decade, the synthesis of superparamagnetic iron oxide nanoparticles (SPIONs)

has received considerable attention due to their potential applications in biomedical fields.

However, success in size and shape control of the SPIONs has been mostly achieved

through organic routes using large quantities of toxic or/and expensive precursors in

organic reaction medium at high reaction temperature. This has limited the biomedical

applications of SPIONs and therefore, development of a synthetic method under aqueous

condition that is reproducible, scalable, environmentally benign, and economically feasible

for industrial production is of paramount importance in order to fully realise their practical

applications. Spinning Disc Processing (SDP) has been used to synthesise

superparamagnetic magnetite (Fe3O4) nanoparticles at room temperature via a modified

chemical precipitation method under continuous flow condition and offer a potential

alternative to be applied to SPIONs production. SDP has extremely rapid mixing under

plug flow conditions, effective heat and mass transfer, allowing high throughput with low

wastage solvent efficiency. The synthesis process involves passing ammonia gas over a thin

aqueous film of Fe2+/3+ which is introduced through a jet feed close to the centre of a

rapidly rotating disc (500-2500 rpm). Synthetic parameters such as precursor

concentrations, temperature, flow rate, disc speed, and surface texture influence the particle

sizes. The size of the nanoparticles can be controlled within a narrow size distribution over

the range of 5-10 nm and the materials exhibit high saturated magnetisations, in the range

of 68 -78 emug-1.

Due to their superparamagnetic property, composite of SPIONs such as carbon nanotubes

(CNTs)/SPIONs have found many promising application such as in drug delivery and as

biosensors. The synthesis conditions associated with the typical bench-top chemistry

exhibit little size control and the resulting Fe3O4 nanoparticles attached to the CNTs are

very broad and the resulting composite Fe3O4/CNTs are ferromagnetic rather than

superparamagnetic. A novel, simple, rapid, cost effective and scalable method has been

developed to decorate single-walled carbon nanotubes (SWCNTs) with 2-3 nm Fe3O4 nano-

ii

particles using SDP. Ultra fine (2-3 nm) Fe3O4 nanoparticles are uniformly deposited on

SWCNTs, pre-functionalised with carboxylic acid groups using microwave radiation. The

deposition process involves a chemical precipitation approach associated with spinning disc

processing (SDP), under continous flow condition and is readily scalable for large scale

synthesis. The resulting decorated SWCNTs were superparamagnetic with specific

saturated magnetisation of 30 emug-1.

The use of Fe3O4 nanoparticles in biomedical applications also depends strongly on their

stability in solutions at physiological pHs and the functionalities of their surface. In the

absence of surface coating, Fe3O4 nanoparticles tend to aggregate due to the van der Waals

forces coupled with the magnetic dipole-dipole attractions between the particles. p-

Sulfonato-calix[n]arenes have been used to stabilise and functionalise Fe3O4 nanoparticles

to form stable ferrofluid. Sulfonato-calix[n]arenes are cyclic phenolic oligomers (where n

denotes the number if repeating units in the cycle) with a hydrophobic cavity, which can

form host–guest inclusion complexes. Such water soluble calixarenes have been shown to

complex with hydrophobic drug molecules and imparted solubility of the drug molecules in

aqueous solutions, and thus have potential as drug delivery carriers. Stable Fe3O4 ferrofluid

was formed by in situ co-precipitation of 1:2 molar ratio of Fe2+ and Fe3+ ions with

ammonia solution in the presence of the p-sulfonato-calixarenes and sulfonated p-

benzylcalixarenes. The samples prepared in the presence of p-sulfonato-calix[6]arene and

sulfonated p-benzyl-calix[4,5,6 and 8]arene have a narrow particle size distribution with

diameters ranging from 5-10 nm. Whereas, the Fe3O4 nanoparticles synthesised in the

presence of p-sulfonato-calix[8]arene have a broader size distribution with the particle size

ranging from 5-20 nm. Fe3O4 prepared in the presence of p-sulfonato- calix[4 and 5]arenes

did not form stable suspensions, in contrast to stable dispersed nanoparticles using p-

sulfonato-calix[6 and 8]arenes. The samples herein showed superparagmagnetic behaviour

with saturated magnetisation values ranging from 68-76 emug-1.

Gold (Au) and silver (Ag) are ideal coating for Fe3O4 nanoparticles due to their high

chemical stability, biocompatibility, and their affinity for binding to amine/thiol terminal

iii

groups of organic molecules. In addition these coatings also render the Fe3O4 nanoparticles

with plasmonic properties. The combination of magnetic and plasmonic propertis make these

composite nanoparticles very useful for diagnostics and therapeutic applications. However,

the current available synthesis methods for Fe3O4@Au and Fe3O4@Ag nanoparticles are

organic based and make them unsuitable for bio-applications. A novel, simple, aqueous

based method has been developed to synthesise Fe3O4@Au and Fe3O4@Ag nanoparticles at

room temperature. Fe3O4 nanoparticles are simultaneously stabilised and functionalised

with amine functional groups with dopamine as a surfactant. Nanoparticles of Au in the

range 2 – 3 nm are attached to amine functionalised Fe3O4 nanoparticles, acting as seed for

the growth of ultrathin Au or Ag shells. The monodispersed core-shell nanoparticles

Fe3O4@Au and Fe3O4@Ag, have a particle size range of 10-13 nm with a shell thickness of

approximately 2-3 nm. They are magnetically purified and are superparamagnetic at 300 K

with saturated magnetisation values of 41 and 35 emug-1, respectively.

Magnetic silica microspheres are receiving great attention for possible applications in

magnetic targeting drug delivery, bioseparation and enzyme isolation. However, the current

available methods for preparation suffer from the setback of low loading of Fe3O4

nanoparticles in the silica microsphere, which result in low magnetic moment, thereby

limiting their practical applications. Therefore it is of considerable importance to develop

new alternative synthetic methods for fabricating magnetic silica microspheres with high

magnetic nanoparticles loading. Superparamagentic Fe3O4 nanoparticles (8-10 nm

diameter) and curcumin have been encapsulated in mesoporous silica in a simple multiple-

step self assembly approach process with high Fe3O4 nanoparticles loading (37%). The

synthesis involves loading of curcumin in the Cetyltrimethylammonium bromide (CTAB)

micellar rod in the presence of superparamagnetic Fe3O4 nanoparticles via a parallel

synergistic approach. The synthesised magnetic mesoporous silica composite material is

stable, superparamagnetic with high saturation magnetisation before and after curcumin

leaching experiment. Under physiological pH in phosphate buffer, the curcumin is slowly

released over several days. These magnetic mesoporous silica are expected to have great

potential as targeted drug delivery systems.

iv

Table of Contents Abstract ................................................................................................................................... i

Table of Contents .................................................................................................................. iv

List of Figures ....................................................................................................................... vi

Abbreviations ....................................................................................................................... vii

Acknowledgements ............................................................................................................... ix

Details of Publications and Abstracts ..................................................................................... x

Statement of Candidate Contribution .................................................................................. xiv

1 General Introduction ....................................................................................................... 1

1.1 Overview ................................................................................................................ 1

1.2 Fundamentals of Magnetism .................................................................................. 1

1.2.1 Magnetism in Nanoparticles: Superparamagnetism........................................... 2

1.3 Superparamagnetic Iron Oxide Nanoparticles (SPIONs) ...................................... 5

1.4 Synthesis of Fe3O4 Nanoparticles .......................................................................... 7

1.4.1 Co-precipitation with Bases ............................................................................... 7

1.4.2 Microemulsion or Micelle Routes ...................................................................... 9

1.4.3 Thermal Decomposition ................................................................................... 10

1.4.4 Hydrothermal Synthesis ................................................................................... 11

1.5 Surface Properties of Fe3O4 Nanoparticles .......................................................... 12

1.5.1 Stabilisation and Surface Functionalisation of Fe3O4 Nanoparticles .............. 13

1.6 Biomedical Applications of SPIONs ................................................................... 23

1.6.1 Hyperthermia Treatment .................................................................................. 24

1.6.2 Magnetic Resonance Imaging (MRI) ............................................................... 25

1.6.3 Magnetic Targeted Drug Delivery ................................................................... 27

1.6.4 Magnetic Bioseparation ................................................................................... 29

1.7 Cytotoxicity of SPIONs ....................................................................................... 30

1.8 Challenges ............................................................................................................ 32

2 Introduction to Series of Papers .................................................................................... 33

2.1 Scalable Synthesis of SPIONs under Continuous Flow Conditions .................... 33

v

2.2 Developing the use of Calixarenes as Novel Surfactants to Stabilise and

Functionalise SPIONs ...................................................................................................... 36

2.3 Fabrication of SPIONs Hybrid Nanomaterials for Biomedical Applications ...... 38

2.4 Fabrication of Magnetic Silica Microspheres with High Magnetic Nanoparticle

Loading for Drug Delivery .............................................................................................. 40

3 Series of Papers ............................................................................................................. 42

4 Conclusions and Future Work ....................................................................................... 73

5 Appendices .................................................................................................................... 93

6 References ................................................................................................................... 108

vi

List of Figures

Figure 1.1 Magnetisation (M) versus magnetic field strength (H) where Ms is the

saturation magnetisation, Mr is the remanence magnetisation, and HC is the

coercivity.[2] ............................................................................................................ 2

Figure 1.2 (a) Multidomain materials and (b) single domain material. [6] ............................. 3

Figure 1.3 Schematic of changes in coercivity, Hc with particle diameter, D; Dc is the

single domain particle size.[7] ................................................................................. 4

Figure 1.4 Magnetisation curves of superparamagnetic nanoparticles. ................................. 5

Figure 1.5 Spin arrangement in a Fe3O4 molecule (net spins are in red). ............................. 6

Figure 1.6 Schematic representation of nanoparticle synthesis using reverse micelles.[30] .. 9

Figure 1.7 Thermal decomposition method for synthesis of SPIONs [39] ........................... 11

Figure 1.8 Surface chemistry of Fe3O4 as a function of pH.[42] ............................................ 13

Figure 1.9 Bidentate chelation of carboxylic acid on magnetite surface[43] ......................... 13

Figure 1.10 Schematic of particle stabilisation with (A) electric double layer and (B)

steric effect.[49] ..................................................................................................... 15

Figure 1.11 Schematic representation of a magnetic nanoparticle-based drug delivery

system.[3] .............................................................................................................. 28

Figure 2.1 Schematic representation of SDP ....................................................................... 35

Figure 2.2 Crystal structure of sulfonato-calix[n]arenes ..................................................... 37

Figure 2.3 p-Sulfonato-calix[n]arenes (1) and sulfonato p-benzyl calix[n]arenes (2)

showing a possible mode of interaction of 1, n = 6, at the surface of the

nanoparticles (NPs).[162] ........................................................................................ 37

Figure 2.4 Chemical structure of curcumin.......................................................................... 41

Figure 4.1 Schematic diagram of RTP. A, B and C are jet feeds for controlling the

processes using the RTP.[181] ................................................................................ 75

vii

Abbreviations

APTES (3-aminopropyl) triethoxysilane

CTAB Cetyltrimethylammonium bromide

DDP Dodecylphosphonic acid

DHDP Dihexadecyl phosphate

DMSA Dimercaptosuccinic acid

DNA Deoxyribonucleic acid

EMU Electromagnetic unit

EFTEM Energy-filtered transmission electron microscope

FDA Food and drug administration

FTIR Fourier transform infrared spectroscopy

HDP Hexadecylphosphonic acid

MPS Mononuclear phagocyte system

MPEG-COOH Monocarboxyl-terminated PEG

MRI Magnetic resonance imaging

NADBS Sodium Dodecylbenzensulfonate

NMR Nuclear magnetic resonance

PAA-PTTM Trithiol-terminated poly(acrylic acid)

PMMA-PTTM Trithiol-terminated poly(methacrylic acid)

PEG Poly(ethyleneglycol)

PEI Polyethylenimine

PLA Polylactide

PLGA Poly(lactic-co-glycolic acid)

PVA Poly(vinyl alcohol)

RES Reticuloendothelial system

SAXS Small angle x-ray scattering

SDS Sodium dodecylsulphate

SLN Solid lipid nanoparticles

SPIONs Superparamagnetic iron oxide nanoparticles

viii

SQUID Superconducting quantum interference device

TEM Transmission electron microscope

TEOS Tetraethoxysilane

TGA Thermogravimetric analysis

TMAOH Tetramethylammonium hydroxide

UV-Vis Ultraviolet-visible

XRD X-ray diffraction

ix

Acknowledgements

First and foremost I would like to thank my research advisors, Professor Colin L. Raston

and Dr. Iyer K. Swaminathan for their continuous support and constant guidance

throughout my PhD.

I am grateful to the staff from the Centre for Microscopy, Characterisation and Analysis for

training and help in microscopy analysis.

I would like to express my appreciation to Professor Tim St. Pierre and Dr. Robert

Woodward from the Physics Department for access to the SQUID Instrument and their

valuable discussion and suggestion.

I am grateful to all the members of Raston research group for their friendship and help.

Special thanks are due to Malaysia government and the University of Malaysia Sarawak for

the postgraduate scholarship.

Finally and certainly not least, my family members for their constant love and support.

x

Details of Publications and Abstracts

Publications

Refereed Journal Articles

1. S.F.Chin, K.S.Iyer and C.L.Raston. A Facile and Green Approach to Fabricate Gold

and Silver Coated Superparamagnetic Nanoparticles. Cryst. Growth Des., 2009, 9,

2685-2689.

Author Contributions: SFC, KSI and CLR designed the research; SFC performed

the research; and SFC, KSI and CLR finalised the manuscript.

(Overall contribution by SFC: 90%)

2. S.F.Chin, K.S.Iyer, M.Saunders, T.S.Pierre, C.Buckley, M.Paskevicius and

C.L.Raston. Encapsulation and sustained release of curcumin using superparamagnetic

silica reservoirs. Chem. Eur. J., 2009, 15, 5661-5665.

Author Contributions: SFC, KSI, and CLR designed the research; SFC performed

the research; SFC and M.Saunders performed EFTEM, TSP, MP and CB performed

the SAXS, SFC, CB and MP analyse the SAXS data and SFC,KSI and CLR finalised

the manuscript. (Overall contribution by SFC: 75 %)

3. S.F.Chin, K.S.Iyer, C.L.Raston, Fabrication of caron nanotubes decorated with ultra-

fine superparamagnetic nanoparticles under continuous flow conditions. Lab Chip.,

2008, 8, 439-442.

Author Contributions: SFC, KSI and CLR designed the research; SFC performed the

research; and SFC, KSI and CLR finalised the manuscript. (Overall contribution by

SFC: 90 %)

xi

4. S.F.Chin, K.S.Iyer, C.L.Raston, M.Saunders, Size selective synthesis of

superparamagnetic nanoparticles in thin fluids under continuous flow conditions. Adv.

Funct. Mater., 2008, 18, 922-927.

Author Contributions: SFC, KSI and CLR designed the research; SFC performed

the research; SFC and MS performed the TEM; and SFC, KSI and CLR finalised the

manuscript. (Overall contribution by SFC: 85%)

5. S.F.Chin, M.Makha, C.L.Raston, M.Saunders, Magnetite ferrofluids stabilised by

sulfonato-calixarenes. Chem. Commun., 2007, 1948-1950.

Author Contributions: SFC, MM and CLR designed the research; SFC performed

the research; SFC and MS performed the EFTEM; and S.F.Chin, M.Makha,

C.L.Raston finalised the manuscript. (Overall contribution by SFC: 80%)

Referred Conference Proceedings

1. S.F.Chin, K.S.Iyer, C.L.Raston and M.Saunders. Process intensification strategies for

the synthesis of superparamagnetic nanoparticles and fabrication of nano-hybrid.

Proceedings of The Nanotechnology Conference and Trade Show, NSTI Nanotech

2008, Boston, USA. 1-6 June, 2008. Vol.1, pp 782-785

2. S.F.Chin, M.Makha, C.L.Raston. Encapsulation of magnetic nanoparticles with

biopolymer for biomedical application. Proceedings of 2006 International Conference

on Nanoscience and Nanotechnology, IEEE, pp 374 -376

Conference Abstracts

1. S.F.Chin, K.S.Iyer and C.L.Raston. A Facile and Green Approach to Fabricate Gold

and Silver Coated Superparamagnetic Nanoparticles. Advanced Materials and

Nanotechnology Conference 4. University of Otago. Dunedin, New Zealand. 8-12th

February, 2009. (Oral presentation)

xii

2. S.F.Chin, K.S.Iyer and C.L.Raston. Encapsulation of Superparamagnetic Magnetite

Nanoparticles in Ultrathin Plasmonic Shells. Australian Research Network for

Advanced Materials Annual Workshop (ARNAM). Deakin University, Victoria,

Australia. 15-18th December, 2008. (Poster presentation)

3. S.F.Chin, K.S.Iyer and C.L.Raston. Encapsulation of Superparamagnetic Magnetite

Nanoparticles in Ultrathin Plasmonic Shells. The University of Western Australia

Postgraduate Student Research Forum. 7th November, 2008. (Poster presentation)

4. S.F.Chin, K.S.Iyer, C.L.Raston and M.Saunders, Process Intensification Strategies for

the Synthesis of Superparamagnetic Nanoparticles and Fabrication of Nano-Hybrid.

NSTI Nanotech 2008, Boston, USA. 1-6th June, 2008. (Poster presentation)

5. S.F.Chin, K.S.Iyer and C.L.Raston. Superparamagnetic Fluorescent Silica Capsules for

Biomedical Application. International Conference on Nanoscience and

Nanotechnology (ICONN). Melbourne, Australia. 24-29th February, 2008. (Oral

presentation)

6. S.F.Chin, K.S.Iyer, C.L.Raston and M.Saunders, Size Selective Synthesis of

Superparamagnetic Nanoparticles in Thin Fluids under Continuous Flow Conditions.

International Conference on Nanoscience and Nanotechnology (ICONN). Melbourne,

Australia. 24-29th February, 2008. (Poster presentation)

7. S.F.Chin, K.S.Iyer and C.L.Raston. Superparamagnetic Fluorescent Silica Capsules for

Biomedical Application. The University of Western Australia Postgraduate Student

Research Forum. 1st November, 2007. (Poster presentation)

8. S.F.Chin, M.Makha, C.L.Raston, M.Saunders. Magnetite Ferrofluids Stabilised by

Sulfonato-Calixarenes. Australian Research Network for Advanced Materials Annual

xiii

Workshop (ARNAM). Australia National University. Canberra, Australia. 3-7th July,

2007. (Poster presentation).

9. S.F.Chin, M.Makha, C.L.Raston, M.Saunders. Magnetite Ferrofluids Stabilised by

Sulfonato-Calixarenes. Conference of Royal Australian Chemical Institute IC07. Wrest

Point Convention Centre. Hobart, Tasmania, Australia. 4-8th February, 2007. (Poster

Presentation).

10. S.F.Chin, M.Makha, C.L.Raston. Encapsulation of Magnetic Nanoparticles with

Biopolymer for Biomedical Application. International Conference on Nanoscience and

Nanotechnology (ICONN). Brisbane Convention & Exhibition Centre, Queensland,

Australia. 3-7th July, 2006. (Poster Presentation).

11. S.F.Chin, M.Makha, C.L.Raston. Encapsulation of Magnetic Nanoparticles with

Biopolymer for Biomedical Application. Australian Research Network for Advanced

Materials Annual Workshop (ARNAM). University of Queensland, Brisbane,

Queensland, Australia. 28 – 30th June, 2006. (Poster Presentation).

xiv

Statement of Candidate Contribution

This thesis contains the results of work carried out by the author in the Discipline of

Chemistry, School of Biomedical, Biomolecular and Chemical Sciences at The University

of Western Australia during the period January 2006 to June 2009.

The work presented herein contains no materials which the author has submitted or

accepted for the award of another degree or diploma at any university and, to the best of the

author’s knowledge and belief, contains no material previously published or written by

another person, except where due reference is made in the text.

Suk Fun Chin

2009

Chapter 1: General Introduction

1

1 General Introduction 1.1 Overview

This thesis aims to develop a simple, environmentally benign, aqueous based

synthetic method for making monodispersed magnetite (Fe3O4) nanoparticles with

controllable size and narrow size distribution. The thesis also reports various ways to

modify the surface of Fe3O4 nanoparticles to render them with colloidal stability and

functionalities for potential biomedical applications.

1.2 Fundamentals of Magnetism

The magnetic properties of matter are fundamentally the result of the motion of

electrons in atoms. At the atomic level, there are two types of electron motion, spin and

orbital, and each has a magnetic moment associated with it. The response of a material

to a magnetic field (H) is characteristic of the magnetic induction or the flux density (B)

and the effect that a material has upon the magnetic induction in a magnetic field is

represented by the magnetisation (M).[1] A universal equation relating these three

magnetic quantities, magnetic field, magnetic induction and magnetisation is as follows

B = μ

0 (H + M)

(1.1) where μ

0 is a universal constant of permeability in a free space, and μ is the permeability

of the material. For equation (1.1), μ0H is the magnetic induction generated by the field

alone and μ0M is the additional magnetic induction contributed by a material.

Figure 1.1 schematically illustrates a hysteresis loop which is a plot of

magnetisation versus field/strength. The application of an external magnetic field

causes the spins within a material to align with the field, and the material is magnetised.


2

The more spins that are aligned with the field, the stronger the magnetic field. When the

external magnetic field is sufficiently strong enough to align all the spins, a maximum

value of the magnetisation called the saturation magnetisation, Ms is achieved. As the

magnitude of the magnetic field decreases, spins cease to be aligned with the field, and

the total magnetisation decreases. The value of the magnetisation at zero fields is called

the remanent magnetisation, Mr. The ratio of the remanent magnetisation to the

saturation magnetisation, Mr/Ms, is called the remanence ratio and varies from 0 to 1.

The coercive field Hc is the magnitude of the field that must be applied in the negative

direction to bring the magnetisation of the sample back to zero.

Figure 1.1 Magnetisation (M) versus magnetic field strength (H) where Ms is the

saturation magnetisation, Mr is the remanence magnetisation, and HC is the

coercivity.[2]

1.2.1 Magnetism in Nanoparticles: Superparamagnetism

Typically, macroscopic magnetic materials are separated into domains or

sections where magnetic spins are cooperatively oriented in the same direction. The

magnetisation inside each domain is uniform, but varies from domain to domain as they

are separated by an interface layer known as the domain wall. In the presence of an

external magnetic field, the domain spins will tend to align with that field, creating an


3

overall magnetic moment, Figure 1.2 a.[3] As the particle size decreases, the number of

magnetic domains per particle decreases down to the limit where it is energetically

unfavorable for a domain wall to exist, and the particle is said to contain a single

domain (SD), Figure 1.2b.[4] Consequently, the alignment of spins under an applied

field is no longer impeded by domain walls. A single-domain particle is uniformly

magnetised with all the spins aligned in the same direction. The magnetisation will be

reversed by spin rotation since there are no domain walls to move. Particles of Fe3O4 are

single domains when the diameter is 50 nm or less.[5] As the particle size continues to

decrease within the single domain range, the coercivity decreases until it reaches Hc = 0.

At this critical particle size (< 20 nm for iron based nanoparticles), the particles are

superparamagnetic. The change in coercivity with particle diameter is shown in Figure

1.3

Figure 1.2 (a) Multidomain materials and (b) single domain material. [6]

(a) (b)(a) (b)


4

Figure 1.3 Schematic of changes in coercivity, Hc with particle diameter, D; Dc is the

single domain particle size.[7]

Magnetic properties of a materials change drastically with their size.

Superparamagnetism is exhibited by very small (1–20 nm) nanoparticles, for which the

material is ferro/ferrimagnetic in the bulk form. The magnetisation relaxation depends

on KV/kT, in which K is the particle anisotropy constant, V is particle volume, k is

Boltzmann’s constant and T is temperature. At a certain reduced size (volume), KV

becomes comparable to the thermal energy kT, leading to the randomization of the

magnetic dipoles in a short period of time.[8] Such small particles do not have permanent

magnetic moments in the absence of an external field, but can respond to an external

magnetic field. The particles are then said to be superparamagnetic. They exhibit no

remanence or coercivity, and thus there is no hysteresis in the magnetisation curve,

Figure 1.4, are therefore often used in many magnetic systems in the biomedical field

because not only are they small, but they also do not retain any magnetic remanence.

The latter reason is important because it means that the particles will not aggregate due

to magnetic forces, however the trade-off is that the particles are paramagnetic in

behavior and therefore it is more difficult to achieve a high magnetisation.[9]


5

Figure 1.4 Magnetisation curves of superparamagnetic nanoparticles.

1.3 Superparamagnetic Iron Oxide Nanoparticles (SPIONs)

Superparamagnetic iron oxide nanoparticles (SPIONs) that display high

saturation magnetisation values and magnetic susceptibility have recently received a

surge of interest as their potential has been demonstrated in biomedical applications,

such as magnetic resonance imaging (MRI),[10, 11] drug delivery,[12] and hyperthermia

therapy.[13] The motivation of using iron based magnetic nanoparticles is due to their

superparamagnetic characteristic, high saturation magnetisation, and lower sensitivity to

oxidation than nanoparticles of transition metals. Furthermore, iron oxide particles are

known to be non-toxic, and are eventually broken down to form blood haemoglobin.

The most well-known synthetic magnetic iron oxides are magnetite (Fe3O4)

(saturation magnetisation (Ms) = 92 emug-1) and maghemite (γ-Fe2O3) (Ms = 74 emug-

1).[14] Fe3O4 is a mixed iron oxide (FeO·Fe2O3) which has the inverse spinel crystal

structure with a face-centered cubic (fcc) lattice and 8 formula units per unit cell. In the

inverse spinel structure, half of the Fe3+ ions are tetrahedrally coordinated and the other


6

half of the Fe3+ ions and all of the Fe2+

ions are octahedrally coordinated. Each

octahedral site has six nearest neighbour O2- ions arranged at the corners of an

octahedron, while each tetrahedral site has four nearest neighbor O2- atoms arranged at

the corners of a tetrahedron.[15] Magnetite is a ferrimagnetic material.[16] The spins of

cations in octahedral and tetrahedral sites oppose one another, but there is a still a net

magnetic moment. Figure 1.5 is a schematic of the spin configuration in magnetite.

Figure 1.5 Spin arrangement in a Fe3O4 molecule (net spins are in red).

Fe3O4 nanoparticles are not very stable under ambient conditions, and are easily

oxidized to maghemite (γ-Fe2O3) and dissolved in acidic medium. (Equation 1.2) γ-

Fe2O3, is a reddish brown black ferromagnetic material, and has a cubic crystal structure

similar to the inverse spinel crystal structure of Fe3O4. It is an important magnetic

pigment with properties similar to Fe3O4. Since γ-Fe2O3 is chemically stable in alkaline

and acidic medium, some researchers deliberately convert Fe3O4 nanoparticles into γ-

Fe2O3. The conversion can be achieved by dispersing them in acidic media, followed

by the addition of iron(III) nitrate or hydrogen peroxide as oxidizing agent.

Fe3O4(s) + 2H+(aq) → γ-Fe2O3(s) + Fe2+

(aq) + H2O(l)

(1.2)

Tetrahedral Fe3+ (3d5)

Octahedral Fe2+ (3d6)

Fe3+ (3d5)


7

1.4 Synthesis of Fe3O4 Nanoparticles

The physical and chemical properties of SPIONs are greatly affected by the

synthesis route, and for this reason various approaches have been developed to produce

SPIONs in order to obtain desired properties. Various synthesis methods for preparing

SPIONs have been developed which includes both aqueous and non-aqueous systems

involving co-precipitation,[17] thermal decomposition and/or reduction,[18]

microemulsion routes,[19] hydrothermal synthesis,[20] and electrospray techniques.[21]

These methods can produce a variety of sized SPIONs. In this thesis, chemical routes

are targeted and four typical approaches are discussed.

1.4.1 Co-precipitation with Bases

Co-precipitation is a facile and convenient way to synthesise iron oxides (either

Fe3O4 or γ-Fe2O3).[22] This approach is attractive in terms of cost and including

sustainability metrics at the inception of the science. Typically, a strong base such as

NaOH, or NH4OH aqueous solution is added under stirring to an aqueous solution of

Fe(III) and Fe(II) salts at a molar ratio of 2:1, either at room or elevated temperatures. In

general, the synthesis can be expressed by the following equation:

Fe2+ + 2Fe3+ + 8OH- → Fe3O4 + 4H2O

(1.3)

According to the thermodynamics of this reaction, a complete precipitation of

Fe3O4 will only occur between pH 9 and 14, while maintaining a molar ratio of

Fe3+:Fe2+ at 2:1 under a non-oxidizing environment. Otherwise, Fe3O4 might also be

oxidized to non-magnetic ferric hydroxide according to the following:

Fe3O4 + 1/4O2 + 4/5H2O →3Fe(OH)3

(1.4)


8

Formation of the product and particle morphology is also affected by various

parameters of the coprecipitation process. A rapid pH increase in the range of 9–14 is

essential. Slow addition of a base may lead to the formation of a brown nonmagnetic

precipitate, probably hydroxides.[23] Ionic strength of the reaction solution also affects

the nanoparticle sizes. Nanoparticles of Fe3O4 formed in the presence of 1.0 M NaCl

aqueous solution results in nanoparticles approximately 1.5 nm smaller than those

formed in the absence of the salt. In addition, these smaller nanoparticles formed in the

higher ionic strength solutions have lower saturation magnetisation (63 emug-1) than

those prepared in solution devoid of NaCl free solutions (71 emug-1). The decrease in

saturation magnetisation could be attributed to the decrease in size of the nanoparticle

for the samples prepared in such media.[24] The effect of the nature of the cation of

hydroxide base used to form Fe3O4 nanoparticles has been investigated in terms of

magnetic properties and crystal structures. X-ray analysis shows nonmagnetic species

are formed when strong alkaline bases such as KOH and NaOH are used. This could be

due to the rapid shift in the pH of the mixture to pH ~14 which creates iron hydrate

complexes that are incapable of forming Fe3O4 nanoparticles. The same researchers also

report that Fe3O4 nanoparticles prepared using NH4OH in the pH range of 8.5 to 10 do

not produce any of the nonmagnetic iron oxide forms.[25] Reaction temperature is

another factor that effects the properties of the Fe3O4 nanoparticles, with increasing

reaction temperature decreasing the particle size.[26]

Although the co-precipitation method is simple, fast, and environmentally

benign, the nanoparticles prepared by this method unfortunately tend to be rather

polydispersed with particle size in the range from 3 to 50 nm. Laborious size selection

efforts are required to improve the polydispersity. The particle size distribution can be

controlled by carrying out the co-precipitation process in the presence of a surfactant.

However, this results in very small nanoparticles involving co-precipitating ferric and

ferrous chloride with base in the presence of surfactants. For example, 4.5 nm Fe3O4

nanoparticles were prepared by co-precipitation of Fe2+ and Fe3+ with ammonia solution

in the presence of trithiol-terminated poly(acrylic acid) (PAA-PTTM) and trithiol-


9

terminated poly(methacrylic acid) (PMAA-PTTM).[27] Co-precipitation involving FeCl3

and FeCl2 salts (2:1 molar ratio) with ammonia solution in the presence of hydrophilic

block copolymer (poly(ethylene oxide-b-methacrylic acid) generates Fe3O4

nanoparticles with broad particles size of 5 ± 4 nm.[28]

1.4.2 Microemulsion or Micelle Routes

Microemulsions are thermodynamically stable surfactant-water-oil mixtures,

which can be subdivided into different classes, notably water-in-oil (w/o) and oil-in-

water (o/w) microemulsions.[29] The synthesis of nanoparticles involving microemulsion

involves the preparation of two identical water-in-oil microemulsions (Figure 1.6),

which only differ by the nature of their aqueous phase. One contains the salts of Fe2+

and Fe3+ ions (reactant A) and the other one the precipitating agent (reactant B). Upon

mixing, collisions and droplet coalescence bring the reactants A and B into contact to

form the AB or Fe3O4 precipitate (Figure 1.6). The coprecipitation process occurs in

tiny droplets of water embedded within a surfactant. The surfactant-stabilised water

droplets act as nanoreactors to physically restrict the nanoparticles nucleation, growth,

and agglomeration in which the particle size is controlled by the size and shape of these

nanoreactors.[30]

Figure 1.6 Schematic representation of nanoparticle synthesis using reverse

micelles.[30]


10

Highly monodispersed Fe3O4 nanoparticles have been synthesised using aerosol

OT or AOT (sodium dioctylsulphosuccinate) as surfactant in (w/o) microemulsion.[31]

Other surfactants, such as sodium dodecylbenzensulfonate (NADBS)[32]

cetyltrimethylammonium bromide (CTAB),[33] sodium dodecylsulphate (SDS)[34] and

Triton X-100[35] have been used to synthesise Fe3O4 nanoparticles with very narrow size

distributions. Although this method is effective in controlling particle size and shape,

large amounts of solvents and surfactants are needed for the synthesis, which is

environmentally and economically undesirable. Moreover the yield of the particles is

lower as compared to other chemical methods, such as that involving co-precipitation

and thermal decomposition.

1.4.3 Thermal Decomposition

Highly crystalline monodisperse magnetic nanocrystals with smaller size can be

synthesised through the thermal decomposition of iron organic and organometallic

precursors, such as iron cupferronates, [Fe(Cup)3], iron pentacarbonyl, [Fe(CO)5] or

iron(III) acetylacetonate, [Fe(acac)3], in high-boiling organic solvents in the a presence

of surfactant (Figure 1.7). The size and morphology of the nanoparticles can be

controlled by adjusting reaction time, temperature, the concentration and or ratios of the

reactants, nature of the solvent, and precursors. Thermal decomposition of [Fe(CO)5]

has been used for the preparation of monodisperse γ-Fe2O3 nanoparticles with average

diameters from 4 to 16 nm as a result of a careful control of the molar ratio of metal

precursor to surfactant, [Fe(CO)5] and oleic acid respectively.[36]

Monodisperse Fe3O4 nanoparticles synthesised from a thermal decomposition of

[Fe(acac)3] at 265 oC has also been reported. Nanoparticles of Fe3O4 with 4 nm

diameters are available by refluxing a mixture of 2 mmol [Fe(acac)3], 20 mL of

diphenyl ether, 10 mmol of 1,2-hexadecanediol, 6 mmol of oleic acid, and 6 mmol of

oleyl amine for 30 min. Larger nanoparticles are generated through a seeded growth


11

process by adding more precursors to the 4 nm sized nanoparticles, then refluxing the

resulting mixture. By controlling the ratio of seed relative to the precursor, nanoparticles

up to 16 nm in diameter are formed.[37]

Nanoparticles of Fe3O4 synthesised by a thermal decomposition method can be

capped with an organic surfactant, but can only be dispersed in organic solvents, and

thus are unfavorable for biomedical applications. Some researchers have tried to

overcome this limitation by preparing water-dispersible Fe3O4 nanoparticles using

thermal decomposition methods. Water-dispersible Fe3O4 nanoparticles are formed by

refluxing a solution of [Fe(acac)3] or FeCl3 in 2-pyrrolidone. 2-Pyrrolidone is a high

boiling solvent and also serves as a stabiliser due to its coordination capacity with metal

ions. By controlling the reflux time, nanoparticles of 4, 12 and 60 nm in diameter can

be generated.[38]

Figure 1.7 Thermal decomposition method for synthesis of SPIONs [39]

1.4.4 Hydrothermal Synthesis

Hydrothermal synthesis of Fe3O4 nanoparticles has been reported in the

literature. The reactions are carried out in aqueous media in vessels or autoclaves where

the pressure can be higher than 2000 psi and the temperature can be above 200 oC. In


12

the hydrothermal process, the particle size is controlled mainly through the nucleation

and grain growth rate. The reaction conditions such as solvent, temperature, and time

have significant effects on the product. Nucleation is faster than grain growth at high

temperatures and results in decreased in particle size. On the other hand, a prolonged

reaction time and higher water content results in precipitation of larger Fe3O4

nanoparticles. Nanoparticles of Fe3O4 with diameter of 39 ± 5 nm have been prepared

through hydrothermal processes at 250 oC for 24 hours.[40] A generalised hydrothermal

method for synthesising a variety of different nanocrystals by a liquid–solid–solution

reaction has also been reported. The system consists of metal linoleate (solid), an

ethanol–linoleic acid liquid phase, and a water–ethanol solution at different reaction

temperatures under hydrothermal conditions. This method has been used to prepare

monodispersed 9 nm Fe3O4 nanoparticles.[41]

1.5 Surface Properties of Fe3O4 Nanoparticles

The surface iron atoms of Fe3O4 act as Lewis acids and coordinate with

molecules that donate lone-pairs of electrons. In aqueous solutions, the iron atoms

coordinate with water, which dissociate readily to leave surface hydroxyl

functionalities. In aqueous solution, the Fe3O4 surface will be either positive or negative,

depending on the pH of the solution. The isoelectric point of Fe3O4 is at a pH of 6.8[42]

where an equal number of positive and negative surface charges co-exist on the Fe3O4

surface; there is no electrostatic double layer, and thus no double layer repulsive forces,

and the particles accordingly aggregate. At pH values lower than the isoelectric point,

surface hydroxyl groups become protonated and the surface of Fe3O4 is positively

charged whereas at pH higher than the isoelectric point, the hydroxyl groups are

deprotonated and the Fe3O4 become negatively charged (Figure 8).


13

Fe-OH2+ ⇔ Fe-OH ⇔ Fe-O-

( pH< 6.8) (pH > 6.8)

Figure 1.8 Surface chemistry of Fe3O4 as a function of pH.[42]

It is well known that organic ligands (carboxylic acids) and inorganic ligands

(phosphates) form stable covalent bonds with the surface of iron oxides.[15] This has

been exploited for preparing stable dispersions of Fe3O4 nanoparticles. Oleic acid, a C18

fatty acid, is one example of a surfactant that covalently binds to the surface of iron

oxides and creates magnetic dispersion in nonpolar hydrocarbon solvents. The chemical

structure of the magnetite-carboxylate bond has been investigated by Massart and

coworkers using magnetite particles coated with oleic acid. Based on the FTIR data, the

carboxylic acids bound to the surface of Fe3O4 in a chelating bidentate configuration.[43]

Figure 1.9 Bidentate chelation of carboxylic acid on magnetite surface[43]

1.5.1 Stabilisation and Surface Functionalisation of Fe3O4

Nanoparticles

Even though the synthesis of Fe3O4 nanoparticles has been intensively pursued,

preparation of stable Fe3O4 nanoparticles dispersion in aqueous medium for biomedical

applications still remains as a challenge. Due to their large surface area to volume ratio,

in the absence of any surface coating, Fe3O4 nanoparticles tend to aggregate through van


14

der Waals force in order to reduce their inherently large surface energy (100 dyn/cm).[44]

In addition to this flocculation which is due to each particle in the magnetic field of a

neighbour getting further magnetised, which increases aggregation of the

nanoparticles.[45] The isoelectric point of Fe3O4 nanoparticles is at a pH 6.8,[46] where

there are equal numbers of positive and negative surface charges as discussed above.

The increasing tendency for the nanoparticles to aggregate has made the unmodified

bare Fe3O4 nanoparticles inherently unsuitable for biomedical applications.

Consequently, stabilisers such as surfactants or polymers are often employed to

coat the surface of the Fe3O4 nanoparticles during or after the synthesis to create a

repulsive force between the particles to prevent agglomeration and obtain a

thermodynamically stable colloidal suspension known as a ferrofluid. Two stabilisation

mechanisms are commonly employed to overcome van der Waals and magnetostatic

attractive forces in magnetic nanoparticles: (1) electrostatic stabilisation and (2) steric

stabilisation (Figure 1.10). Electrostatic stabilisation can be achieved by changing the

isoelectric point of the nanoparticles. When the surface of a particle is charged the

counter-ions are also present, which creates an electric double-layer. The potential

energy between two similarly charged particles increases upon their approach, leading

to repulsion. Electrostatic stabilisation is a pH and ionic strength sensitive method and

is of limited use, especially in biomedical applications.[47] Steric or entropic

stabilisation is the most common method used in ferrofluid preparation. Surfactants are

physically adsorbed or chemically grafted onto the surface of magnetic colloidal

particles. Surfactants containing functional groups, such as carboxylic acids,

phosphates, and sulfates, can bind to the surface of Fe3O4 nanoparticles.[15]

The range of biomedical applications of magnetic nanoparticles depends on their

stability in solutions at various physiological pH values, and the degree to which their

surfaces may be chemically functionalised.[48] Surface modifications of SPIONs with

different stabilisers including organic and inorganic surfactants affect their stability,

magnetisation and, therefore, their practical applications. Ideally surface modification of


15

SPIONs not only should stabilise the nanoparticles in solution, but they should also

protect them from degradation, endow biocompatibility and impart the desired

functionisation. For biomedical applications, the materials used for coating should be

biocompatible, non-toxic, hydrophilic and be able to stabilise the nanoparticles in

aqueous environment at physiological pH (7.6).

Figure 1.10 Schematic of particle stabilisation with (A) electric double layer and (B)

steric effect.[49]

1.5.1.1 Surface Modification with Ionic Surfactant

Tetramethylammonium hydroxide (TMAOH) a well known agent for

redispersing agglomerated iron oxide nanoparticles because it is a low polarizing cation

that favors stability of the particles in solution.[50] Flocculation is reversible, involving

substitution of the counterion in the precipitate For example, if sodium hydroxide is

used for precipitation of the magnetite, the Na+ cation can be exchanged with NH4

+

cation by stirring the precipitate with an ammonia-based buffer. Elimination of the

flocculating NH4 +

countercation through its conversion into neutral NH3 ammonia by

TMAOH then allows the peptization of the precipitate.[51] TMAOH is highly basic

(pH~14) significantly limiting its ability to serve as a stabilising agent for biological

applications.


16

1.5.1.2 Surface Modification with Polymer Stabilisers

Both synthetic and natural polymers have been used by researchers to coat Fe3O4

nanoparticles, either by in situ coating or post synthesis coating. Poly(ethylene glycol)

(PEG),[52-54] poly(lactic-co-glycolic acid) (PLGA),[55-59] and polyvinyl alcohol (PVA)[60,

61] are examples of synthetic polymers that have been used as a surfactant to coat and

stabilise Fe3O4 nanoparticles. Natural polymers or biopolymers are more favorable as

coating materials for Fe3O4 nanoparticles due to their biocompatibility and

bioavailability. Natural polymers such as dextran,[60, 62, 63] alginic acid[64-67] and

chitosan[68-71] have also been investigated as coating materials for Fe3O4 nanoparticles.

These polymer coated magnetic nanoparticles are intensively studied for magnetic-field-

directed drug targeting,[72] and as contrast agents for magnetic resonance imaging.[73]

Poly(ethylene glycol) (PEG)

PEG is widely used as coating materials for magnetic nanoparticles owing to its

hydrophilic, nonantigenic, nonimmunogenic and protein resistant polymers.[52] The

ability of PEG derivatives to improve uptake of nanoparticles has been reported and is

generally recognised as being the result of solubilisation the particles in the cell

membrane lipid bilayer mediated by PEG. Immobilization of PEG on the nanoparticles

increases the amount of nanoparticles uptake into cancer cells in comparison to the

unmodified nanoparticles.[53] Some studies have been carried out on the influence of

PEG coated SPIONs on human fibroflasts, with PEG coated SPIONs not affecting the

cell adhesion behavior, and their internalisation into endosomes offer the opportunity to

label a variety of cells with high efficiency.[74] Monocarboxyl-terminated PEG (MPEG-

COOH) modified SPIONs have been prepared using a one-pot route by thermal

decomposition of [Fe(acac)3] in 2 pyrolidone in the presence of MPEG-COOH. The

resultant nanoparticles exhibit excellent solubility in aqueous solution as well as in

physiological saline. MRI experiments on living rats demonstrate good biocompatibility


17

and a long blood circulation time, which makes them potentially useful as MRI contrast

agents.[75]

Poly(lactic-co-glycolic acid) (PLGA)

Poly(lactic-co-glycolic acid) (PLGA) has been widely studied for biomedical

applications such as in drug delivery and tissue engineering owing to its biocompatible,

biodegradable properties and, low toxicity, within its approval by the US FDA for

pharmaceutical applications.[55] SPIONs coated with oleic acid are encapsulated in

PLGA particles by emulsion evaporation,[56] oil-in-water-in-oil emulsion,[57] and a

emulsification–diffusion method[58] for drug delivery. The surface coverage of PLGA

on the SPIONs results in a decrease of the saturated magnetisation (Ms) to nearly half of

that of pure magnetite nanoparticles.[59]

Polyvinyl alcohol (PVA)

Polyvinyl alcohol (PVA) is a hydrophilic, synthetic biocompatible polymer

formed by the hydrolysis of poly(vinyl acetate) and often contains residual acetyl

groups. It has excellent film forming, emulsifying and adhesive properties. Coating of

PVA onto the surfaces of nanoparticles prevents their agglomeration. SPIONs

synthesised in the presence of aqueous PVA form necklace-like chains close to 100 nm

in length. The particle sizes of the PVA dispersed iron oxide particles are 5.78 ± 1.30

nm. The small sizes of the SPIONs is due to the presence of the polymer during the

synthesis. [60] Others have reported the same method to afford SPIONs 4-7 nm in

diameter which have saturation magnetisation values of ~55 emug-1. PVA binds to the

surface of the SPIONs irreversibly. The crystallinity of the SPIONs decrease with the

increase of PVA concentration used in the precipitation of process.[61]


18

Dextran

Dextran is a polysaccharide polymer composed exclusively of (-D-)

glucopyranosyl units with varying degrees of chain length and branching. Detailed

magnetic and structural properties on magnetite/maghemtite mixtures formed in the

presence of dextran (40k g/mol) have been studied.[60] These dextran coated

nanoparticles are 4.11 ± 0.85 nm in diameter. The formation of magnetite nanoparticles

in the presence of 40k g/mol dextran for immunospecific magnetic separation of cells

has also reported.[62] The dextran was functionalised with additional hydroxyl groups for

binding amino groups on proteins. The effect of reaction times, temperature, molecular

weight of dextran, amount of absorbed dextran and the stability of the dextran-

magnetite nanoparticles have been studied. Increase of absorption time, reaction

temperature, and the mass ratio of dextran to magnetite increase the absorption of

dextran on magnetite nanoparticles. Lower molecular weight dextran absorbs on the

nanoparticles better relative to the higher molecular weight analogue.[63] Dextran also

can be desorbed from the iron oxide surface by heating the particles at 120 oC or on

dilution. A crosslinker, epichlorhydrin can be added to dextran coated iron oxide

nanoparticles to avoid desorption of dextran. [76]

Alginic Acid

Alginic aicd or alginate is a natural-occurring polyelectrolytic polysaccharide

found in all species of brown algae and some species of bacteria. It is a linear polymer

composed of α-L-guluronate (G) and 1,4 linked β-D-mannuronate (M) units in varying

proportions and sequential arrangements. There are only three different block types that

make up the polymer: mannuronic acid blocks, guluronic acid blocks, and the

alternating mannuronic and guluronic acid blocks.[64] Alginic acid has been used

extensively in the food, cosmetic, pharmaceutical and biomedical industries for their gel

forming properties in the presence of multivalent cations.[65] Alginic acid has been


19

coated onto the preformed magnetite nanoparticles to form stable ferrofluids. The

stability is attributed to the strong interaction of the carboxylate groups in alginic acid

with iron ions. The reported hydrodynamic diameters for the alginic acid coated

magnetite nanoparticles is 30–150 nm.[66] The potential application of alginic acid

coated SPIONs as MRI contrast agent has been evaluated by some researchers,

indicating that the alginic acid coated SPIONs has high T2 relaxivity, and thus is

suitable to be used as negative MRI contrast agent.[77]

Chitosan

Chitosan is a cationic and hydrophilic polysaccharide derived by deacetylation

of chitin, a by-product of the seafood industry. It consists of repeating units of

glucosamine and N-acetyl-glucosamine, the proportions of which determines the degree

of deacetylation of the polymer.[68] Chitosan is receiving a great deal of interest in food

and pharmaceutical because of its biocompatibility and biodegradability, and is

nontoxic. [69] The bio-adhesiveness of chitosan makes it an ideal material for mucosal

drug delivery. The co-administration of chitosan with drugs has been shown to enhance

the transcellular and paracellular transport of drugs across mucosal epithelium.[70]

Nanoparticles of Fe3O4 have been encapsulated in chitosan microspheres using a

sonochemical method. Their potential as MRI contrast agents have been studied but, the

size of these magnetic chitosan microspheres are rather large, ranging from 100 to 150

μm in diameter.[71] Chitosan/magnetite nanocomposites have been synthesised in a

magnetic field via in situ hybridisation. Interestingly the Fe3O4 nanoparticles are

assembled to form chain-like structures under the influence of the external magnetic

field, which mimics the magnetite chains inside of magnetotatic bacteria.[78]


20

1.5.1.3 Surface Modification with Nonpolymeric Stabilisers

Fatty Acids

Papell was among the first to report the formation of colloid magnetite

nanoparticle dispersions with oleic acid in 1968.[79] Nanoparticles of Fe3O4 were

prepared by the ball milling method, and oleic acid was added to form nonpolar

hydrocarbon colloidal dispersions. Oleic acid, (CH3(CH2)7CH=CH(CH2)7CO2H), which

has a C18 (oleic) tail with a cis-double-bond in the middle, forming a kink. Stabilisation

occurs because the carboxylic acid group covalently links with the surface of the Fe3O4

nanoparticles with the aliphatic chain extending out into the nonpolar solvent,

preventing aggregation of the particles by a steric (entropic) mechanism. However,

stearic acid, (CH3(CH2)16CO2H), with no double bond in its C18 (stearic) tail, cannot

stabilise Fe3O4 nanoparticles in organic solvent.[80] The difference between the two

surfactants is that oleic acid contains a cis-double bond that forms kinks whereas stearic

acid contains no kinks. Researchers have concluded that the kink in oleic acid allows

the eighteen-carbon chain to be better solubilized by hydrocarbon solvents, resulting in

stable colloids.

Oleic acid has been coated to preformed Fe3O4 nanoparticles [81] and can be

added in situ in the reaction mixture during synthesis.[82] Dispersion of oleic acid coated

Fe3O4 nanoparticles in water is complicated by the nonpolar nature of the hydrocarbon

chain. Investigations on the effect of the chain length of the fatty acid on forming water

dispersible magnetite solutions have been carried out with fatty acids with C10 to C15 are

capable of forming stable water-based dispersions.[83]


21

Phosphate containing Surfactants

The possibility of using alkanephosphonic acid and alkanesulphonic as

surfactant for Fe3O4 nanoparticles in organic solvents has been investigated. Their zeta

potential and IR spectroscopy suggest that the phosphate ions form bidentate complexes

with adjacent sites on the iron oxide surface.[84] Some researchers have compared the

phosphonate and phosphate surfactants such as dodecylphosphonic acid (DDP),

hexadecylphosphonic acid (HDP) and dihexadecyl phosphate (DHDP) to fatty acids

such as oleic acid and lauric acid.[85] TEM on Fe3O4 nanoparticles coated with

phosphonate and phosphate show the particles are agglomerated whereas the fatty acid

coated magnetite particles appeared to be more discrete and separated. Even though

thermogravimetric analysis (TGA) suggests that alkyl phosphonates/phosphates

strongly bind to the surface of the iron oxide nanoparticles, there is reduced

dispersibility in apolar solvents due to the formation of quasi-bilayer structures.

Silica

SPIONs have been coated with silica shells by many researchers taking

advantages of the silanol groups on the silica shells reactivity with alcohols and silane

coupling agents to produce dispersions, which are stable in nonaqueous solvents. The

negative charge on the silica shells allows coated magnetic nanoparticles to be

redispersibled in water without the need of adding other surfactants. In addition, the

silanol groups are ideal for covalently binding specific ligands. For example, amine

groups have been introduced on the surface of the silica coated Fe3O4 nanoparticles by

hydrolysis and condensation of an organosilane, such as aminopropyltriethoxysilane.[86]

The main advantage of a silica shell is that it provides a biocompatible, nontoxic

surface coating with high hydrophilicity which would help to retard the rapid clearance

of the nanoparticles by the reticuloendothelial system (i.e. to liver, spleen, and bone


22

marrow).[87] Inorganic shells also protect the magnetic core from rapid biodegradation or

oxidation toward a less magnetic phase. Thus there is an advantage in coating magnetic

particles with silica. Many methods have been developed for this purpose, including the

Stöber method and sol–gel processes.[88, 89] The thickness of the coating can be tuned by

varying the concentration of ammonium and the ratio of tetraethoxysilane (TEOS) to

H2O. Since the iron oxide surface has a strong affinity towards silica, no primer is

required to promote the deposition and adhesion of silica. Uniformly sized silica-coated

Fe3O4 nanoparticles (Fe3O4@SiO2) synthesised in a simple one-pot process using

reverse micelles as nanoreactors have been reported.[90] The method affords control of

the core diameter of the Fe3O4 nanoparticles by adjusting the w value (w = [polar

solvent]/[surfactant] ) in the reverse-micelle solution, and the thickness of the silica

shell can be tuned by varying the amount of TEOS added after the synthesis of the

magnetite cores. The water in the oil microemulsion method has also been used for the

preparation of silica-coated iron oxide nanoparticles.[91] Three different non-ionic

surfactants (Triton X-100, Brij-97 and Igepal CO-520) feature in the preparation of

microemulsions. The effects of the surfactants on the particle size, crystallinity, and the

magnetic properties have been investigated. A layer of uniform silica coating as thin as

1 nm encapsulating the iron oxide nanoparticles is formed by base-catalysed hydrolysis

and polymerisation of TEOS in the microemulsion.

Noble Metals

Coating noble metals shell such as Au or Ag on magnetic nanoparticles not only

can improve chemical stability by protecting the core from oxidation, corrosion and

dispersibility of the naked core nanoparticles, but also modify or enhance their

properties. Elemental Au and Ag have been the preferred coating material because of

the well-known optical properties, good biocompatibility with the human body and their

ability to firmly combine with biomolecules such as polypeptides and DNA via the

thiols group (-SH).[92] Shells of Ag and Au also exhibit a surface plasmon resonance at


23

optical frequencies resulting in a strong absorption and scattering of light by the

particles at specific frequencies. Thus, the magnetic core/Au shell or Ag shell

nanoparticles combine the advantage of Au or Ag nanoparticles for convenient binding

and detection of biomolecules and magnetic particles for easy separation. However, the

direct coating of SPIONs with Au or Ag is very difficult, because of the dissimilar

nature of the two surfaces.[93]

Nevertheless, some progress has been made in synthesising core-shell

nanoparticles of Fe3O4@Au or Fe3O4@Ag. Au shells have been deposited on iron oxide

(Fe2O3 or partially oxidized Fe3O4) nanoparticles (~9 nm) in an aqueous solution via

hydroxylamine seeding to form 60 nm sized particles.[94] Core-shell nanoparticles of

Fe3O4@Au and Fe3O4@Ag from 18 to 30 nm in diameter have also been synthesised

using the reverse micelle method.[95] Recently, Fe3O4@Au and Fe3O4@Au@Ag core-

shell nanoparticles were synthesised by reducing HAuCl4 and AgNO3 in a chloroform

solution of oleylamine by using Fe3O4 as nucleation seeds. The resulting Fe3O4@Au

and Fe3O4@Au@Ag nanoparticles are capped with oleylamine, but the particles are not

dispersible in water, thus limiting their biomedical applications.[96] The synthesis of

Fe3O4@Au core/shell nanoparticles through a sequential growth mechanism has also

been reported. A mixture of gold acetate [Au(Ac)3] with surfactants is added to a

suspension of Fe3O4 and heated to 180–190°C for 1.5 h.[97] The method involves high

temperatures and the use of organic precursors and solvent which makes the product

unfavorable for biomedical applications.

1.6 Biomedical Applications of SPIONs

SPIONs (Fe3O4 and γ-Fe2O3) offer great potential in biomedical applications

such as magnetic guided targeting drug delivery, bioseparations, MRI contrast agent and

hyperthermia treatment owing to their unique superparamagnetic properties.

Superparamagnetism is important in these biomedical applications because once the

external magnetic field is removed, magnetisation disappears (negligible remanence and


24

coercivity), and thus agglomeration (and the possible embolisation of capillary vessels)

is avoided. SPIONs are considered to be biodegradable with iron being reused/recycled

by cells using normal biochemical pathways for iron metabolism.[98] The biomedical

applications of magnetic nanoparticles require that the nanoparticles are monodispersed

so that each individual nanoparticle has nearly identical physical and chemical

properties for controlled biodistribution, bioelimination and contrast effects. The

magnetic nanoparticles should also have a high magnetic moment, and readily modified

via surface reactions so that they are capable of binding specifically to the biomolecules

of interest, and impart stability under various physiological conditions.

1.6.1 Hyperthermia Treatment

Hyperthermia is a medical treatment that depends on locally heating tissue over

42°C for a short period of time to destroy the tissue, especially tumors by utilizing the

magnetic properties of magnetic nanoparticles in an external alternating magnetic

field.[99] When magnetic particles are subjected to a variable magnetic field, some heat

is generated due to hysteresis loss in a process known as magnetic induction heating.[100]

The amount of heat generated depends on the nature of magnetic material and magnetic

field parameters. During the treatment, magnetic particles are concentrated around a

tumor site and placed within an oscillating magnetic field for heating. When the

temperature is above the therapeutic threshold of 42 oC for 30 min or more, the cancer

cells are destroyed as they are inefficient in dissipating the heat, whereas healthy cells

can survive at such temperatures.[101]

SPIONs have a much higher rate of specific absorption compared to larger

magnetic particles with several magnetic domains, and therefore are suitable for use in

hyperthermia treatment.[102] In many in vitro experiments with different types of tumor

cells and SPIONs, the optimal physical properties of the SPIONs,[103] the particle uptake

rate into tumor cells,[104] and duration and strength of the magnetic field[105] have been


25

investigated to improve the efficiency of the cancer treatment. Clinical trials in treating

human patients affected with prostate and brain tumors using local magnetic

hyperthermia in combination with radiotherapy have been undertaken.[99] When used

alone, hyperthermia is shown to be effective in shrinking tumors located close to the

skin. Clinical trials show significant tumor size reduction and destruction of tumor cells

in 8 of 10 breast cancer patients.[106] In conclusion the application of SPIONs for local

magnetic hyperthermia is a promising tool for future therapy due to the selective,

efficient and non-invasive surgical features of this treatment method.

1.6.2 Magnetic Resonance Imaging (MRI)

MRI is essentially proton NMR carried out on tissue. Protons are excited with

short pulses of radio frequency radiation; the free induction decay as they relax is

measured and deconvoluted by means of a Fourier transform, which provides an image

of the tissue that corresponds to proton density. Areas of high proton density, usually in

the form of water or lipid molecules, have a strong signal and appear bright. Areas of

bone or tendon, which have a low proton density because of the lack of water and lipids,

have a weak signal and appear dark. It is also dependent upon the rate of relaxation of

the spin of the protons T1 (the exponential decay constant in the longitudinal relaxation

of the spins) and T2 (the exponential relaxation constant in the dephasing (transversal)

of the spins).[107] Traditionally, a major limitation of MRI has been its inability to

distinguish differences in soft tissue types (e.g., healthy parts of the liver from diseased

lesions), as the relative proton densities can be very similar. Other regions, such as the

bowel, are hard to image because air pockets and fecal matter make the proton density

inconsistent.[108]

To obtain a better image with well-defined mapping, contrast agents are utilised

during the imaging procedure. MRI provides more legible images of both bones and soft

tissues of the human body and is a safer method of detection as compared to X-ray and

Computerized Tomography (CT) scanning.[109] The conventional, paramagnetic


26

gadolinium chelates typically behave as T1 contrast agents that cause positive contrast

enhancement and provide a bright state in the image where they are accumulated.[110]

There are some SPOIN types of contrast agents that are now commercially available

such as Feridex®, Endorem™, GastroMARK®, Lumirem®, Sinerem®, or Resovist®,

marketed by Advanced Magnetics Inc.[111] However, these conventional contrast agents

are not sensitive enough for scanning very tiny tumors or specific cancer cells in the

human body. Thus many researchers are focusing on developing new contrast agents for

MRI diagnosis in order to get a satisfactory level of contrast in imaging for all types of

tissue.

SPIONs were developed as a contrast agent in MRI as their nano-size allows

shortened relaxation times of protons along with improved sensitivity of the diagnosis

of MRI.[112] The size of SPIONs should be taken into consideration for development of

MRI contrast agents. When the diameter is larger than 30 nm as in the commercial

products Lumirem and Endorem, the particles are easily trapped in the liver and spleen;

thereby, their usage is mainly limited to detection of liver and spleen diseases.[113]

Smaller particles as in the commercial product Sinerem, have longer life times in the

bloodstream and can be used for a wider range of applications.

Appropriate surface modification of SPIONs with suitable biologically active

specific functional groups such as antibodies, hormones or proteins, can enhance the

sensitivity of the MRI diagnosis.[114] Dextran coated SPIONs are of particular interest

as they show an increase in the cell uptake (endocytosis) of nanoparticles, and

consequently enhances the cell imaging. However, they do not show sufficient cellular

uptake to enable cell tracking because of a relatively inefficient fluid phase endocytosis

pathway. There have been reports of cellular uptake of dextran-modified nanoparticles

resulting in cell damage and reduced cell proliferation.[115] The researchers found that a

large dextran coating on iron oxide leads to particle accumulation in the lymph

nodes.[116] As a result, other means of surface modification have been, and are currently

being explored, to increase the uptake efficiency of SPIONs.


27

Magnetoliposomes, which are prepared by derivatising magnetoliposomes with

poly(ethylene glycol) have been examined as MRI contrast agents. This was an attempt

to minimise the rapid uptake of the magnetoliposomes by cells of the reticulo-

endothelial system (liver, spleen, and lymph nodes). The iron oxide was stabilised with

lauric acid and the vesicle composition was a mixture of dipentadecanoyl-

phosphatidylglycerol and dimyristoyl-phophatidyl-ethanolamine-N poly(ethylene

oxide).[73] Other researchers have coated SPIONs with starch, albumins, silicone,

poly(ethylene glycol) with particle sizes ranged between 30-150 nm for MRI contrast

agent.[117] A remarkable result of the application of SPIONs in cancer diagnosis has

been recently reported, where the size of a prostate tumour as small as 2 nm can be

identified in a MRI scan in contrast to much larger size ( 30-50 nm ) for conventional

MRI.[118]

1.6.3 Magnetic Targeted Drug Delivery

Another possible and most promising application of the SPIONs is in targeted

drug delivery. Current problems associated with systemic drug administration include

even biodistribution of pharmaceuticals throughout the body, the lack of drug specificity

towards a pathological site, the necessity of a large dose to achieve high local

concentration, non-specific toxicity and other adverse side effects due to high drug

doses. Spatial and temporal control over release of a drug is thereby a key for increasing

drug efficacy and minimizing side effects. Amongst the current methods for drug

targeting, magnetic targeting of a drug immobilized on magnetic materials under the

action of an external magnetic field has been developed. Here a chemotherapeutic drug

is attached to a biocompatible magnetic nanoparticle and then injected into the patient

via a circulatory system; the particles enter the bloodstream, and external, high-gradient

magnetic fields are used to concentrate the complex at a specific target site. Once the

drug/carrier is concentrated at the targeted sites, the drug can be released either via

enzymatic activity or changes in physiological conditions such as pH, osmolality, or


28

temperature, and then taken up by the tumour cells (Figure 1.11).[119] A key motivation

for magnetic micro- and nanoparticle-based targeting lies in the potential to reduce or

eliminate the side effects of chemotherapy drugs by reducing their systemic distribution

as well as the possibility of administering lower but more accurately targeted doses of

the cytotoxic drugs in the targeted area. Besides chemotherapeutics, many other

biomolecules including peptides, proteins [120] and therapeutic radioisotopes,[121] can be

adsorbed or encapsulated into magnetic microspheres or nanospheres. Several properties

of the SPIONs such as the size (magnetic core, hydrodynamic volume, and size

distribution), charge and surface chemistry are particularly important and strongly affect

both the blood circulation time as well as bioavailability of the particles within the

body.[122]

Figure 1.11 Schematic representation of a magnetic nanoparticle-based drug delivery

system.[3]

One of the setbacks of the magnetic nanoparticles for drug delivery systems is

that they are rapidly cleared by macrophages or the reticuloendothelial system (RES)

before they are able to reach the site of the tumor cells. Depending on their size, surface

functionalisation, and hydrophilicity, a rapid uptake of uncoated nanoparticles by the

mononuclear phagocyte system (MPS) is likely after systemic administration, followed

by clearance to the liver, spleen, and bone marrow. To overcome these problems,

magnetic nanoparticles are coated with various types of polymers such as dextran or

PEG[123] to prevent detection and uptake of the nanoparticles by the macrophages of the

RES, and also increase their circulation in the blood stream. In addition, other

hydrophilic and biocompatible polymers such as PLGA,[124] or inorganic materials such


29

as silica,[125] have also been used to coated SPIONs. The coating prevents agglomeration

at various pH and salt concentrations in the body, and also allows binding of drug

molecules. However, the entrapment of magnetic nanoparticles into polymeric or

inorganic drug carrier systems has resulted in significant loss in magnetisation (~40-

50%) of the magnetic core.[126] The decrease in magnetisation negatively influences the

magnetic targeting ability of the carrier system in vivo. The above approach is further

limited by the amount of magnetic nanoparticles that can be incorporated into other

drug delivery systems; for example, only 6 wt % α-Fe can be incorporated in silica

nanospheres. The magnetic moment for such system may be too weak for effective drug

targeting.[127]

In addition to polymeric and inorganic materials, magnetoliposomes have been

synthesised for drug delivery.[128, 129] These nanoparticles have a typical core-shell

structure with a magnetic iron oxide core surrounded by an artificial liposome.

Improved drug delivery to cancer cells have been demonstrated using

magnetoliposomes that target epidermal growth factor receptors.[130] Although the

success in cytotoxic drug delivery and tumour remission has been achieved by several

groups using animals models including swine, [131, 132] rabbits [133] and rats, [123, 134] there

are still challenges to overcome which are associated with their applications in humans.

These limitations include: (i) difficulties in scaling up from animal models due to the

larger distances between the target site and the magnet, and (ii) there are risks for the

magnetic carriers to block the blood stream arising from accumulation in the target sites.

1.6.4 Magnetic Bioseparation

The basic concept in magnetic bioseparations is to selectively bind the

biomaterial of interest (e.g., a specific cell, protein, or DNA sequence) to a magnetic

particle and then separate them from their surrounding matrix using a magnetic field.

Magnetic separation using SPIONs is proven to be a highly sensitive technique for

separation of specific biological entities (e.g., DNAs, proteins, and cells) from their


30

native environment for analysis. Magnetic separation also offers benefits over

conventional methods due to quick processing time, reduced chemical need, easy

separation using a magnet and ease of automation. This is due to the on–off nature of

magnetisation with and without an external magnetic field, enabling the transportation

of biomaterials with a magnetic field.[135] Surface-derivatised SPIONs with appropriate

targeting ligands such as antibodies, hormone or folic acid are usually bound to the cells

and these SPION-bound cells are then magnetically separated by an external magnetic

field from the mixture.[136] Magnetic separation using antibodies has been applied to

many biomedical research and has proven to be highly accurate and efficient in

targeting because the antibodies can specifically bind to their matching antigens on the

surface of the targeted species. Successful use of immunospecific SPIONs for

separation of breast cancer cells, urological cancer cells,[137] lung cancer cells,[138] red

blood cells,[139] and bacteria[140] have been demonstrated. Polyethylenimine (PEI) coated

SPIONs have been employed to simplify the purification of plasmid DNA from

bacterial cells and to develop a rapid protocol for extracting and purifying transfection-

grade plasmid DNA from bacterial culture.[141]. Magnetic separations eliminate

pretreatment, such as centrifugation, or filtration. The separations are fast, gentle,

scalable, easily automated and can be used in situations, in which other techniques are

impossible or impractical to perform.

1.7 Cytotoxicity of SPIONs

Historically SPIONs have been regarded as clinically benign and well tolerated

in vivo, with LD50 in rats = 400 mg/kg.[142] However, it is vital to examine the potential

cytotoxicity arising from their size, shape and coating materials in order to ensure their

safe clinical uses. In vitro cytotoxicity test using 30 and 47 nm nanoparticles of Fe3O4 at

the highest dosage tested (250 μgmL-1) resulted in approximately 30% decrease in cell

viability, and the nanoparticles were judged as exhibiting little to no toxicity.[143] Higher

concentrations were tested by others at 23.05 mM of nanoparticles with no significant

difference between the exposed cells and the control was reported. However, this may


31

be due to the relatively short exposure time of four hours.[144] In other more extensive

studies, it was found that the cytotoxicity of SPIONs is dose dependent. At low dosage

of 250 μgmL-1, the SPIONs induce a 25–50% loss in fibroblast viability, whereas, with

the highest concentration tested (2.0 mg mL-1) resulting in about 60% loss of cell

viability.[145]

On the other hand, the choice of coating materials also has a significant impact

on the cytotocity. Coating of iron oxide using biocompatible coating such as poly(D,L

lactide) effectively lower the acute toxicity of the iron oxide nanoparticles in mice.[146]

In addition, in vitro cytotoxicity experiments have been undertaken in suspensions of

human granulocytes with Fe3O4 nanoparticles loaded polylactide (PLA),

polylactide/glycolide (PLA/GA) and solid lipid nanoparticles (SLN). The highest

toxicity was observed for the faster degrading polymers, low molecular weight PLA and

PLA/GA, while the Fe3O4 nanoparticles loaded SLN are the least cytotoxic. However,

the authors concluded that the cytotoxicity of the Fe3O4 nanoparticles loaded polylactide

(PLA), polylactide/glycolide (PLA/GA) are still relatively low which qualify them as

having potential for intravenous injection with regard to the toxicological

acceptance.[147] Cells exposed to PEG-coated SPIONs remained more than 99% viable

relative to control at an upper concentration of 1 mg mL-1.[148] Another test with

Pullulan (Pn)-coated SPIONs also showed no cytotoxic effects, with the cells

remaining more than 92% viable at 2.0 mg mL-1.[149] Core-shell nanoparticles of

Fe3O4@SiO2 with 50 nm thick silica shell have been administrated into mice in order to

evaluate the biodistribution and the toxicity. After a 4 weeks observation, the core-shell

nanoparticles were detected in the brain, indicating that such nanosized materials can

penetrate blood–brain barrier (BBB), with the presence of these nanaoparticles not

disturbing the function of the brain or producing apparent toxicity.[150] Recently a

quantifiable model cell system has been developed to quantify the effects of iron oxide

nanoparticles upon cell behavior and, in particular, the ability of cells to appropriately

respond to biological cues. The SPIONs used were coated with dimercaptosuccinic acid

(DMSA) in order to improve the cell uptake efficiency of the iron oxide nanoparticles.


32

Their results indicated that intracellular delivery of even moderate levels of DMSA

coated SPIONs may adversely affect the nerve cell function. [151]

1.8 Challenges In summary, even though numerous approaches of SPIONs synthesis have been

intensively pursued and some significant progress has been made over the past decade,

the synthesis of highly crystalline and monodispersed SPIONs with controllable size is

still a challenge. Clearly there is a compelling need to develop a synthetic protocol with

fine control over the nucleation and growth of SPIONs. Success in size and shape

control of the SPIONs has been mostly achieved through organic routes which often use

large quantities of toxic or/and expensive precursors and using high temperatures. Thus

there are advantages in developing synthetic methods that are aqueous based,

reproducible, scalable, environmentally benign and economically feasible for industrial

production in order to fully realise the practical applications of SPIONs. The successful

use of SPIONs in biomedical applications also requires SPIONs to be stable under

physiological conditions with specific targeting ligands on their surface. The major

challenge herein is overcoming the various chemistry issues to design multifunctional

systems with tailored properties and functionalities. These multifunctional systems

include SPIONs for magnetic manipulation, optical probes (e.g fluorescent dyes or

quantum dots) for tracking, functional groups for bioconjugation and selective targeting,

and encapsulated or immobilized drugs for therapy. Such multifunctional systems have

the potential to revolutionize both the diagnosis and treatment of diseases in the future.

The thesis herein reports on the synthesis of SPIONs as novel hybrid systems which

address the aforementioned issues.

Chapter 2: Introduction to Series of Papers

33

2 Introduction to Series of Papers

It is evident from the literature review (Chapter 1: Introduction) that SPIONs are

an important class of materials for biomedical applications such as drug delivery,

hyperthermia, MRI, and magnetic separation. In order to realise the medical potential of

SPIONs, the following issues need to be addressed: (i) a better synthesis route that is

scalable, reproducible and cost effective; (ii) enhanced colloidal stability at

physiological conditions using novel surfactants; (iii) effective strategies to overcome

the difference in surface energy to fabricate hybrid materials (silver coated SPIONs,

gold coated SPIONs, SPIONs coated carbon nanotubes); (iv) methods to assemble high

loading of SPIONs in mesoporous carriers for drug delivery applications. A series of

experiments were undertaken to address the above-mentioned issues, which forms the

focus of the research for the PhD thesis as summarised below:

2.1 Scalable Synthesis of SPIONs under Continuous Flow

Conditions

The current method of choice for the production of SPIONs with very narrow

size distribution involves high temperature thermal decomposition of an organometallic

precursor in the presence of surfactants. This method is currently the best in terms of

size and shape control. However, thermal decomposition requires high reaction

temperature (100–320 oC), long reaction time (from hours to days), and the use of toxic

and/or expensive organic solvents, organic surfactants and precursors which make

process scalability problematic.[152] As an alternative, microemulsion methods can also

be used to synthesise monodispersed SPIONs with controlled size and shape. However,

only a very low yield and poorly crystalline nanoparticles are usually obtained from this

method.[153] Moreover, SPIONs obtained from both thermal decomposition and

microemulsion methods are capped with organic bio-incompatible surfactants and this

limits their biomedical applications. Co-precipitation with bases is the preferred route


34

for preparation of SPIONs, as it is simple, fast, and more environmentally benign since

the reaction medium is water.[154] Nonetheless, this method gives very little control on

the size of the particles and their shape, and requires extensive size selection processes

after the synthesis. The laborious size selection process limits both the throughput and

the yield of the nanoparticles. Therefore, the search for simple synthetic pathways for

water-dispersible SPIONs with controlled size and shape is an important challenge, and

clearly, a simple, environmental benign, scalable and economically feasible synthetic

method is highly desirable for the production of magnetic nanoparticles.

Conventional bench wet chemistry synthetic routes for the production of

nanoparticles have inherent limitations, such as poor particle size distribution and

reproducibility, and difficulties in scalability for commercial production. Process

intensification strategy offers alternative routes, alleviating the obstacles of the relaxed

fluid dynamic regime associated with conventional batch processes. Process

intensification attempts to exploit fluid flow characteristics within a range of reactor

geometries, using mechanical effects (mixing, turbulence and shear forces) to provide

extremely high heat transfer (192 kwm-2s-1), mass transfer (10-4ms-1), and mixing

rates.[155] Spinning disc processing (SDP) is a form of process intensification for

application in both chemical transformations of molecules and in the formation of

monodispersed nanoparticles with a very narrow particle size distribution.[156] SDP

allows control over all the operating parameters involved in particle formation, enabling

the simultaneous and individual optimization of many interdependent operating

mechanisms, with the ultimate goal of achieving very narrow particle size

distributions.[157]

An important feature of SDP is that it is under continuous flow affording

scalability for mass production. Varying the synthesis parameters for SDP, such as flow

rates, rotation speed and concentration can be used to control the formation of

spheroidal nanoparticles of β-carotene with a diameter as low as 35 nm and a narrow

size distribution.[158] In addition, SDP has also been used to prepare nanoparticles of


35

silver down to 5 nm, with control over particle size, morphology, agglomeration, phases

and defects, by varying the operating conditions and the choice of surfactant.[159]

The capability of SDP in gain access to superparamagnetic Fe3O4 nanoparticles

via modified co-precipitation method has been explored. The key components of SDP

include a rotating disc with controllable speed, and feed jets located slightly off-center

from the disc. A dynamics thin fluid film (1-200 μm) is generated on a rapidly rotating

disc surface (30-3000 rpm) within which nanoparticles form, Figure 2.1. Fe3O4

nanoparticles were synthesised by introducing ammonia gas over a thin film of Fe2+/3+

on a disc rotating between 500 and 2500 rpm with the aqueous solution being

introduced by a jet feed. High shear forces generated in the thin films produce waves

that intensify the absorption of gas into the liquid. High mass transfer of the gas to the

liquid is thereby effective in controlling the amount of base in the liquid and in

controlling the build up of the particles, which is under plug flow conditions. All the

reactants are subjected to the same conditions; thereby producing nanoparticles of

narrow size distribution with all the size of the Fe3O4 nanoparticles controlled, over the

range of 5–10 nm by judicious choice of synthetic conditions and operating parameters

for the SDP. The resulting particles have remarkably high saturation magnetisation

values (68-78 emug-1). This approach is cost effective, fast and can be readily scaled up

for large-scale production of Fe3O4 nanoparticles. Indeed the SDP can serve both as the

research reactor as well as the production reactor, pending on the quantities and how

long the system is operated.

Figure 2.1 Schematic representation of SDP


36

Results published in: S.F.Chin, K.S.Iyer, C.L.Raston, M.Saunders, Size selective

synthesis of superparamagnetic nanoparticles in thin fluids under continous flow

conditions. Adv. Funct. Mater., 2008, 18, 922-927.

2.2 Developing the use of Calixarenes as Novel Surfactants to

Stabilise and Functionalise SPIONs The range of biomedical applications of magnetic nanoparticles depends on their

stability in solutions at physiological pH and the degree to which their surfaces may be

chemically functionalised. In the absence of surface coating, Fe3O4 nanoparticles tend to

aggregate due to the van der Waals forces coupled with the magnetic dipole-dipole

attractions between the particles. In the current study, the utilization of sulfonato[6,8]-

calixarene and sulfonated p-benzylcalix[4,5,6 and 8]arenes as surfactants for stabilisation

of Fe3O4 nanoparticles have been investigated. Calix[n]arenes (n = 4–8) are a family of

cyclic host molecules comprising [n] phenolic units, having hydrophobic cavities, which

can result in inclusion complexes with a wide variety of guest molecules. para-

Sulfonato-calix[n]arenes, represents a class of water soluble cavitands which have been

widely studied for their biomedical applications. They have shown to complex with

amino acid and Bovine Serum Albumin (BSA)[160] and solubilize a variety of otherwise

non-soluble drugs.[161] In addition, sulfonato-calix[4,5,6,8]arenes can also act as

surfactants in stabilising trans-β-carotene nanoparticles.[158] Stable ferrofluids are

formed by a rapid and simple in situ co-precipitation from a solution of Fe(II) and Fe(III)

chloride in the appropriate ratio, with aqueous ammonia, in the presence of the p-

sulfonato-calix[n]arenes, 1 and sulfonated p-benzylcalix[n]arenes, 2 (Figure 2.3). The

deprotonated phenolic OH groups at the lower rim of the calixarene bind to the surface

iron center on the Fe3O4 nanoparticles and leave the SO3- groups of the calixarene facing

outward, thereby electrostatically repelling other nanoparticles, and circumventing

aggregation. The calixarenes not only serve as surfactants to stabilise the Fe3O4

nanoparticles, they also functionalise the Fe3O4 nanoparticles for potential biomedical

applications. The calixarene stabilised Fe3O4 nanoparticles are colloidally stable at


37

physiological pH, and exhibited superparamagnetic behaviour with high saturation

magnetic moment at room temperature.

Figure 2.2 Crystal structure of sulfonato-calix[n]arenes

SO3-

OHn

OHn

SO3-

1 2 n = 4,5,6,8

FeOOFe

O

NPs

O

SOO

SOOO

OO

SOOO

O

O

SOO

O

SOOO

OS

OOO

O

FeOO

Figure 2.3 p-Sulfonato-calix[n]arenes (1) and sulfonato p-benzyl calix[n]arenes (2)

showing a possible mode of interaction of 1, n = 6, at the surface of the nanoparticles

(NPs).[162]

Results published in: S.F.Chin, M.Makha, C.L.Raston, M.Saunders, Magnetite

ferrofluids stabilised by sulfonato-calixarenes. Chem. Commun., 2007, 1948-1950.

Sulfonato-calix[4]arene Sulfonato-calix[6]arene Sulfonato-calix[8]arene


38

2.3 Fabrication of SPIONs Hybrid Nanomaterials for

Biomedical Applications

Functionalisation of carbon nanotubes (CNTs) with SPIONs has received much

interests owing to their promising applications in drug delivery[163] and as chemical

sensors.[164] In situ chemical precipitation from solutions of Fe2+/3+ using aqueous

NH4OH or NaOH to directly coat CNTs with Fe3O4 nanoparticles using conventional

bench top batch chemistry results in a broad distribution of Fe3O4 nanoparticles

attached to the CNTs, ranging from 25–80 nm, with low coating density.[165] This is

due to the lack of control of the synthetic conditions associated with typical bench-top

batch chemistry and is further complicated due to the tendency of CNTs to clump

together.[166] Moreover, the resulting composite Fe3O4/CNT material is ferromagnetic

rather than superparamagnetic. Therefore a more efficient way for coating the CNTs is

highly desirable. The potential capability of SDP for coating single-wall carbon

nanotubes (SWCNTs) with narrow size distribution of Fe3O4 nanoparticles was

explored by in situ coating of SWCNTs with Fe3O4 nanoparticles via a chemical

precipitation method using SDP in aqueous media at room temperature under

continuous flow conditions. The use of SDP to coat SWCNTs led to ultrafine (2-3 nm)

Fe3O4 nanoparticles with narrow size distribution, uniformly distributed on the surface

of the SWCNTs. The SWCNTs were pre-functionalised with carboxylic acid using

microwave radiation. The strong shearing force on the rotating disc creates waves and

ripples, and ensures efficient mixing throughout the entire fluid layer on the disc. This

resulted in the nanoparticles having similar growth conditions after nucleation, and led

to uniform size distribution of Fe3O4

Noble metal (Au and Ag) coating of Fe3O4 nanoparticles not only enhance the

colloidal stability of the Fe3O4 nanoparticles, but they can protect the magnetic core from

oxidation, at the same time imparting plasmonic properties of the metal particles in the

nanocomposite. The combination of magnetic and plasmonic properties makes

Fe3O4@Au and Fe3O4@Ag core-shell nanoparticles are candidates for bio-diagnostic


39

and therapeutic applications. Furthermore, Au and Ag coatings are also inert,

biocompatible, and readily bind to amine/thiol terminate groups of organic molecules,

such as enzymes and antibodies. Synthesis methods for coating Fe3O4 with Au and Ag

shells reported in the literature involve organic routes which produced Fe3O4@Au and

Fe3O4@Ag nanoparticles that are capped with organic surfactants and are only

dispersible in organic solvents.[95, 167] An aqueous based synthetic method is preferable

in order to synthesise water dispersible nanoparticles that can be readily used in bio-

applications. In accepting this challenge, a simple aqueous based synthetic approach has

been developed to coat Fe3O4 nanoparticles with a thin layer of Au or Ag shell. The

strategy involves a “seed-mediated growth” method, where preformed 2-3 nm Au

nanoparticles were attached to the –NH2 functionalised Fe3O4 nanoparticles, with Au3+

or Ag+ reduced onto the surface of the composite nanoparticles for complete

encapsulation in ultrathin shells. Fe3O4 nanoparticles were stabilised and pre-

functionalised with amine (-NH2) groups using dopamine as a surfactant. Dopamine

serves as linker to bind the Fe3O4 nanoparticles and small Au seeds. The two adjacent

OH group on the aromatic ring of the dopamine bind covalently to the iron atoms while

the terminal NH2 groups promote binding of Au seeds. The resulting Fe3O4@Au and

Fe3O4@Ag core-shell nanoparticles exhibit superparamagnetic behaviour with saturated

magnetisation values of 41 and 35 emug-1, respectively.

Results published in:

(i) S.F.Chin, K.S.Iyer, C.L.Raston, Fabrication of carbon nanotubes

decorated with ultra-fine superparamagnetic nanoparticles under

continuous flow conditions. Lab Chip. 2008, 8, 439-442.

(ii) S.F.Chin, K.S.Iyer, C.L.Raston. A facile and green approach to fabricate

gold and silver coated superparamagnetic nanoparticles. Cryst. Growth

Des. 2009, 6, 2685-2689


40

2.4 Fabrication of Magnetic Silica Microspheres with High

Magnetic Nanoparticle Loading for Drug Delivery

Magnetic silica microspheres are receiving great attention for possible

applications in magnetic targeting drug delivery, bioseparation, enzyme isolation, and

wastewater treatment. Silica microspheres are biocompatible, hydrophilic, stable and a

highly porous material. The silica surface is covered with silanol groups (Si-OH), which

facilitates its chemical modification by organofunctional silane coupling tethered to the

surface to covalently immobilize specific ligands.[168] The magnetic properties of the

silica matrix allow not only the targeting or accumulation of drugs in a desired place of

the body, but also the possibility of using hyperthermia combined with drugs for the

treatment of cancerous diseases. Consequently, many techniques have been developed

for fabrication of magnetic silica microsphere, such as a sol-gel method,[169] aerosol

pyrolysis,[170] microemulsion[171] and backfilling.[172] All these methods suffer from the

setback of low loading of Fe3O4 nanoparticles in the silica microsphere, which results in

a low magnetic moment, thereby limiting their practical applications. Therefore it is of

considerable importance to develop new alternative synthesis methods for fabricating

magnetic silica microspheres with high magnetic nanoparticles loading and this is

addressed in the research herein.

Curcumin (Figure 2.4) is a natural polyphenolic compound originally isolated

from the rhizomes of Curcuma longa, and has been used in Indian and Chinese

traditional medicine for hundreds of years.[173, 174] Curcumin has been shown to act as a

potent anti-inflammatory and antioxidant agent.[175, 176] Recently it has generated great

attention as a possible novel anticancer agent because of its encouraging antitumor

activity and negligible toxicity in animal models.[177, 178] However, the limited

bioavailability and poor aqueous solubility of this compound needs to be overcome to

harness the full potential of curcumin in such a way that it is absorbed by the body in a

controlled way.


41

A simple strategic route has been developed to encapsulate a high loading of

Fe3O4 nanoparticles (37 % wt) and curcumin (30 % wt) into a porous silica matrix in a

single process at room temperature under mild condition. These magnetic silica capsules

are superparamagnetic with high saturation magnetisation. The rate of curcumin release

from the magnetic silica capsules was investigated under physiological pH, to determine

the potential applications of these silica capsules as delivery carriers for delivering

biologically active molecules to the body.

Figure 2.4 Chemical structure of curcumin

Results published in: S.F.Chin, K.S.Iyer, M.Saunders, T.S.Pierre, C.Buckley,

M.Paskevicius and C.L.Raston. Encapsulation and sustained release of curcumin using

superparamagnetic silica reservoirs. Chem. Eur. J. 2009, 15, 5661-5665

Chapter 3: Series of Papers

42

3 Series of Papers

1. S.F.Chin, M.Makha, C.L.Raston, M.Saunders, Magnetite ferrofluids stabilised by

sulfonato-calixarenes. Chem. Commun., 2007, 1948-1950.

2. S.F.Chin, K.S.Iyer, C.L.Raston, M.Saunders, Size selective synthesis of

superparamagnetic nanoparticles in thin fluids under continous flow conditions. Adv.

Funct. Mater., 2008, 18, 922-927.

3. S.F.Chin, K.S.Iyer, C.L.Raston, Fabrication of carbon nanotubes decorated with ultra-

fine superparamagnetic nanoparticles under continuous flow conditions. Lab Chip.

2008, 8, 439-422.

4. S.F.Chin, K.S.Iyer, C.L.Raston, A facile and green approach to fabricate gold and silver

coated superparamagnetic nanoparticles. Cryst. Growth Des. 2009, 6, 2685-2689.

5. S.F.Chin, K.S.Iyer, M.Saunders, T.S.Pierre, C.Buckley, M.Paskevicius and C.L.Raston,

Encapsulation and sustained release of curcumin using superparamagnetic silica

reservoirs. Chem. Eur. J., 2009, 15, 5661-5665.

6. S.F.Chin, M.Makha, C.L.Raston. Encapsulation of magnetic nanoparticles with

biopolymer for biomedical application. Proceedings of International Conference on

Nanoscience and Nanotechnology, IEEE, 2006, 374-376

7. S.F.Chin, K.S. Iyer C.L.Raston and M.Saunders. Process Intensification Strategies for

the Synthesis of Superparamagnetic Nanoparticles and Fabrication of Nano-Hybrid.

Proceedings of The Nanotechnology Conference and Trade Show 2008. Boston, USA,

2008. 1, 782-785.

Magnetite ferrofluids stabilized by sulfonato-calixarenes

Suk Fun Chin,a Mohamed Makha,*a Colin L. Raston*a and Martin Saundersb

Received (in CAMBS) 20th December 2006, Accepted 19th January 2007

First published as an Advance Article on the web 16th February 2007

DOI: 10.1039/b618596g

Magnetite (Fe3O4) nanoparticles stabilised by sulfonato-

calixarene macrocycles are readily accessible by a rapid in situ

co-precipitation, and exhibit ferro-fluidic and superparamag-

netic behaviour.

Superparamagnetic magnetite (Fe3O4) nanoparticles have been

widely studied for various scientific and technological applications

such as magnetic storage media,1 contrast agents for magnetic

resonance imaging (MRI),2 biolabelling and separation of

biomolecules,3 and magnetic targeted drug delivery.4 All these

applications require the magnetic nanoparticles to be chemically

stable, have particle size ,20 nm with a narrow size distribution,

and to be well dispersed in aqueous medium.5 Preparation of

stable magnetic nanoparticles is a challenge as the particles have

large surface area to volume ratios and thus they tend to aggregate

to reduce their surface energy. In addition, there are strong

magnetic dipole–dipole attractions between the particles that also

cause the particles to aggregate.

Tetramethylammonium hydroxide (TMAOH) is well known as

a surfactant to stabilize such magnetite nanoparticles.6 However,

TMAOH is highly basic and is not biocompatible. Stabilization of

magnetic nanoparticles can also be achieved by coating the particle

surfaces with organic surfactants or polymers. Oleic acid7 and

lauric acid8 are commonly used for this purpose. Khalafalla and

Reimers9 and Wooding et al.10 have stabilised suspensions of

magnetite using various saturated and unsaturated fatty acids as

primary and secondary surfactants. Synthetic polymers and

biopolymers have also been used to coat and stabilize magnetic

nanoparticles. Examples of the former include poly(vinyl

alcohol),11 poly(acrylic acid),12 and triblock co-polymers (PEO–

COOH–PEO).13 Among the biopolymers used to stabilize

magnetite nanoparticles are dextran14 and alginic acid.15 Most of

this work involves adding the surfactant to the preformed

magnetite nanoparticles.

One of the drawbacks of the above approach is that the particle

size distribution is difficult to control, and the process is time-

consuming requiring further work up. On the other hand, the

synthesis of magnetite nanoparticles in the presence of a surfactant

can circumvent aggregation and control the size of the nano-

particles. Yaacob et al. have prepared magnetite nanoparticles at

room temperature ,15 nm in diameter with a narrow size

distribution of the particles which are inside vesicles based on a

variety surfactants, namely cetyltrimethylammonium bromide

(CTAB) and dedecylbenzenesulfonic acid (DBSA).16

p-Sulfonato-calix[n]arenes are cyclic phenolic oligomers with a

hydrophobic cavity, which can form host–guest inclusion com-

plexes in a similar way to cyclodextrins. Such water soluble

calixarenes display interesting biological properties such as anti-

viral and anti-bacterial activity,17 and form inclusion complexes

with a variety of small molecules.18 Complexes with hydrophobic

drugs impart increased solubility of the drug molecules in aqueous

medium.19 Complexation of Bovine Serum Albumin (BSA), an

arginine- and lysine-rich protein, with sulfonato calixarenes has

been demonstrated by Memmi et al.20 Furthermore, both in vitro

and in vivo toxicity studies show that sulfonato-calixarenes have

low toxicity.21 Overall, p-sulfonato-calixarenes have potential for

biomedical applications, and in this context we note that

sulfonato-calix[4,5,6,8]arenes act as surfactants in stabilizing

trans-b-carotene nanoparticles.22

In this study we report the stabilization of superparamagnetic

magnetite nanoparticles by coating them with p-sulfonato-calix[6

and 8]arenes, 1, n = 6,8, and sulfonated p-benzylcalix[4,5,6 and

8]arenes, 2, n = 4,5,6,8, Fig. 1. Remarkably, stable ferrofluids are

formed by a rapid and simple in situ co-precipitation from a

solution of Fe(II) and Fe(III) chloride in the appropriate ratio, with

aqueous ammonia, in the presence of the p-sulfonato-calixarenes

and sulfonated p-benzylcalixarenes. The calixarenes not only serve

as surfactants to stabilize the magnetite nanoparticles, they also

functionalize the magnetite nanoparticles for potential biomedical

applications, Fig. 1. To the best of our knowledge, this is the first

report in the literature of coating magnetite nanoparticles with

calix[n]arenes.

The formation of superparamagnetic magnetite nanoparticles

was confirmed by SQUID measurements and TEM, while the

aCentre for Strategic Nano-Fabrication, School of Biomedical,Biomolecular and Chemical Sciences, The University of WesternAustralia, 35 Stirling Highway, Crawley, WA 6009, Australia.E-mail: [email protected]; [email protected];Fax: (618) 64881005; Tel: (618) 64881572bCentre for Microscopy Characterisation and Analysis, The Universityof Western Australia, 35 Stirling Highway, Crawley, WA 6009,Australia Electronic supplementary information (ESI) available: synthesis andexperimental details TEM, Diffraction patterns, DLS, zeta potential andFTIR for coated magnetite with calix[n]arene sulfonates. See DOI:10.1039/b618596g

Fig. 1 p-Sulfonato-calix[n]arenes and sulfonato p-benzylcalix[n]arenes

showing a possible mode of interaction of 1, n = 6, at the surface of the

nanoparticles (NPs).

COMMUNICATION www.rsc.org/chemcomm | ChemComm

1948 | Chem. Commun., 2007, 1948–1950 This journal is The Royal Society of Chemistry 2007

interaction of the magnetite with the calixarenes surfactants was

investigated using FTIR, and DLS (dynamic light scattering)

studies in solution. TEM images indicate that the samples prepared

in the presence of p-sulfonato-calix[6]arene and sulfonated

p-benzylcalix[4,5,6 and 8]arene have a narrow particle size

distribution with diameters ranging from 5 to 10 nm. Whereas,

the magnetite nanoparticles synthesized in the presence of

p-sulfonato-calix[8]arene have a broader size distribution with

the particle size ranging from 5 to 20 nm, Fig. 2. The d spacing

values calculated from selected area diffraction patterns obtained

from each of the samples are in good agreement with those for

bulk magnetite (Joint Committee on Powder Diffraction

Standards, JCPDS, card 19-0629, see supporting information).

The particles are roughly spherical in shape and high resolution

TEM imaging of the sample prepared in the presence of

p-sulfonato-calix[6]arene show an amorphous material surround-

ing the iron oxide nanocrystals, Fig. 3. DLS measurements show

the particles are approximately 30% larger, which is consistent with

the assembly of calixarenes on the surface, and associated aquated

environment.23

Elemental maps obtained by energy-filtered TEM showed that

the iron oxide nanoparticles are surrounded by a carbon-rich shell,

and thus the calixarenes are coating the surface of the nano-

particles. The thickness of the coating is around 1.2 to 1.6 nm, as

measured from high resolution TEM images and elemental maps,

which is consistent with the thickness of a monolayer of

calix[n]arene sulfonates and associated aquated sodium ions, Fig. 4.

Calixarenes form complexes with Fe(II and III), with the iron

centres bound to deprotonated phenolic OH groups.24 This is also

likely in the present study for iron centres on the surface of the

nanoparticles, noting that the phenolic groups are deprotonated

under basic conditions and unless the generated phenolate groups

form one calixarene are associated with the same nanoparticle,

there would be spontaneous aggregation. Thus complexation

necessitates the –SO32 groups of the calixarenes to be facing

outward form the surface of the magentite nanoparticles, thereby

electrostatically repelling other nanoparticles, Fig. 1. At the same

time, the presence of calix[n]arenes during the formation of the

nanoparticles may also limit the rapid growth of the crystals and

control the particle size.

Magnetite prepared in the presence of p-sulfonato-calix[4 and

5]arenes did not form stable suspensions, in contrast to stable

dispersed nanoparticles using p-sulfonato-calix[6 and 8]arenes.

This could be due to the cone shaped p-sulfonato-calix[4 and

5]arenes tending to form the well known bilayer arrangement of

these calixarenes with their cavities alternating up and down,

which is associated through hydrophobic interplay.25 Such an

arrangement would effectively have adjacent nanoparticles locked

together through sharing a common bilayer. p-Sulfonato-calix[6

and 8]arene are more fluxional and binding of metal centres to the

calixarenes is less likely to result in bilayer formation between

surface bound calixarenes from adjacant nanoparticle, and thus

favour dispersion of the nanoparticles. Sulfonated p-benzylcalix[4,

5, 6 and 8]arene do not from crystalline bilayer arrangements, and

the dangling benzyl moieties are more likely to act as surfactants

for the attached nanoparticle rather than intertwine in the form of

a bilayer or some other arrangement between adjacent particles,

thereby stabilizing the nanoparticles.

FTIR spectra of the nano-particles are dominated by water

absorption bands which is expected from the aquated sodium ions.

This aside, there is a broad absorption band at ca 580 cm21 (see

supporting information) which corresponds to nFe–O in the crystal

lattice of Fe3O4.26 Peaks in the region 1036 to 1164 cm21

correspond to S–O–C stretching.27 Importantly, the usual broad

peak nC–O at 1450–1460 cm21 for free calixarene27 is shifted to

1400 cm21, with the intensity increased. This is consistent with the

p-sulfonato-calix[6 and 8]arenes and sulfonated p-benzylcalix[4,5,6

and 8]arenes on the surface of the magnetite essentially in the

deprotonated form, with the iron centres attached to the phenolic

O-centres, Fig. 1.

Fig. 2 TEM micrographs of p-sulfonato-calix[n]arene and sulfonated

p-benzylcalix[n]arene coated magnetite: (A) 1, n = 6; (B) 1, n = 8; (C) 2, n =

4; (D) 2, n = 5; (E) 2, n = 6; (F) 2, n = 8.

Fig. 3 TEM micrograph show the coating of p-sulfonato-calix[6]arene

on the surface of the magnetite nanoparticles.

Fig. 4 Carbon and iron elemental maps (left and right respectively) of

magnetite nanoparticles coated with p-sulfonato-calix[6]arene.

This journal is The Royal Society of Chemistry 2007 Chem. Commun., 2007, 1948–1950 | 1949

SQUID measurements all show superparamagnetism. The

hysteresis loops of all the samples measured at room temperature

are presented in Fig. 5. The superparamagnetic behaviour is

evidenced by zero coercivity, zero remnance and the absence of

hysteresis loops. The specific saturation magnetization of the

samples ranged from 68–76 emu g21. The theoretical specific

saturation magnetization of bulk magnetite is reported to be

92 emu g21.28 Some studies suggested that the lower specific

saturation magnetization of nanoparticles as compared to the bulk

materials are due to the reduction of crystalline magnetic

anisotropy constant K for the material.29 Nevertheless, some

studies also suggest that the effects of particle size in the

nanolength scales are complex and alter relaxation processes and

inter-particle interactions.30 Therefore, values of 60–70 emu g21 of

Fe3O4 may be approaching the limit in the specific magnetization

for magnetite nanoparticles with diameters less than 20 nm.

In summary, we have simultaneously stabilized and modified

the surface of magnetite nanoparticles using p-sulfonato-calix[6

and 8]arene and sulfonated p-benzylcalix[4, 5, 6 and 8]arenes, in a

convenient in situ process. The nanoparticles have good colloidal

stability at physiological pH and exhibited superparamagnetic

behaviour with high saturation magnetic moment at room

temperature. These findings along with the ferrofluidic behaviour

have implications in materials, contrast agents and drug delivery,

and more.

The authors graciously acknowledge support of this work by the

Australian Research Council, The University of Western

Australia, Centre for Microscopy and Microanalysis and The

University of Malaysia Sarawak.

Notes and references

Sodium p-sulfonato-calix[n]arenes and sulfonated p-benzylcalix[4, 5, 6and 8]arenes were prepared using literature methods31 while iron chlorideswere purchased from Fluka. Stable suspension ferrofluid of calix[n]arenesulfonates coated magnetite were prepared by coprecipitation of Fe(II) andFe(III) chloride (1:2 molar ratio) with aqueous ammonia in the presencethe calix[n]arene. The solutions were stirred vigorously at roomtemperature under N2 gas until stable black magnetite suspensions areformed. The magnetite suspensions were then centrifuged and thesupernatant decanted. The collected solids were re-dispersed in deoxyge-nated ultra pure Mili-Q water. This process was repeated a few times toremove the excess of surfactants and ammonia.

1 S. Sun, C. B. Murray, D. Weller, L. Folks and A. Moser, Science, 2000,287, 1989.

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3 P. S. Doyle, J. Bubette, A. Bancaud and J. L. Viovy, Science, 2002, 295,2237.

4 C. Alexiou, W. Arnold, P. Hulin, R. J. Klein, H. Renz, F. G. Parak andC. Bergemann, J. Magn. Magn. Mater., 2001, 225, 187.

5 A. K. Gupta and M. Gupta, Biomaterials, 2005, 26, 3995–4021.6 R. J. Hunter. Foundations of Colloid Science, Clarendon, Oxford, 1987;

R. C. Plaza, J. Gomez-Lopera and A. V. Delgado, J. Colloid InterfaceSci., 2001, 242, 306.

7 N. Q. Wu, L. Fu, M. Su, M. Aslam, K. C. Wong and V. P. Dravid,Nano Lett., 2001, 4, 383; G. Kataby, M. Cojomu, R. Prozorov andA. Gedanken, Langmuir, 1999, 15, 1703.

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9 S. E. Khalafalla and G. W. Reimers, IEEE Trans. Magn., 1980, 16(2),178.

10 A. Wooding, M. Kilner and D. B. Lambrick, J. Colloid Interface Sci.,1991, 144, 236.

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12 C. L. Lin, C. F. Lee and W. Y. Chiu, J. Colloid Interface Sci., 2005, 291,411.

13 L. A. Harris, J. D. Harburn, T. G. St. Pierre and M. Saunders, Chem.Mater., 2003, 15, 1367.

14 X. Q. Xu, H. Shen, J. R. Xu, J. Xu, X. J. Li and X. M. Xiong, Appl.Surf. Sci., 2005, 202, 494.

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16 I. I. Yaacob, A. C. Nunes, A. Bose and D. O. Shah, J. Colloid InterfaceSci., 1994, 168, 289; I. I. Yaacob, A. C. Nunes, A. Bose and D. O. Shah,J. Colloid Interface Sci., 1995, 171, 73.

17 F. Perret, A. N. Lazar and A. W. Coleman, Chem. Commun., 2006,2425.

18 T. E. Clark, M. Makha, C. L. Raston and A. N. Sobolev, Cryst. GrowthDes., 2006, 6(12), 2783–2787; W. Yang and M. M. de Villiers, Eur. J.Pharm. Biopharm., 2004, 58, 629–636; S. Shinkai, K. Araki, T. Matsuda,N. Nishiyama, H. Ikeda, I. Takasu and M. Iwamoto, J. Am. Chem.Soc., 1990, 112, 9053–9058.

19 W. Yang and M. M. de Villiers, AAPS J., 2005, 7E241; W. Yang andM. M. de Villiers, Eur. J. Pharm. Biopharm., 2004, 58, 629; W. Yangand M. M. de Villiers, J. Pharm. Pharmacol., 2004, 56, 703.

20 L. Memmi, A. Lazar, A. Brioude, V. Ball and A. W. Coleman, Chem.Commun., 2001, 2474.

21 E. Da Silva, P. Shahgaldian and A. W. Coleman, Int. J. Pharm., 2004,273, 57.

22 N. Anantachoke, M. Makha, C. L. Raston, V. Reutrakul, N. C. Smithand M. Saunders, J. Am. Chem. Soc., 2006, 128, 13847.

23 S. Takata, M. Shibayama, R. Sasabe and H. Kawaguchi, Polymer,2003, 44, 495–501; W. Xu, L. Li, W.-L. Yang, J.-H. Hu, C.-C. Wangand S.-k. Fu, J. Macromol. Sci., Pure Appl. Chem., 2003, 40, 511–523.

24 W. Sliwa, J. Inclusion Phenom. Macrocyclic Chem., 2005, 52, 13; J. Zeller,S. Koenig and U. Radius, Inorg. Chim. Acta, 2004, 357, 1813.

25 J. L. Atwood, L. J. Barbour, M. J. Hardie and C. L. Raston, Coord.Chem. Rev., 2001, 222, 3–32; M. Makha, C. L. Raston, A. N. Sobolevand A. H. White, Chem. Commun., 2004, 1066–1067.

26 A. V. Ramesh Kumar and R. Balasubramaniam, Corros. Sci., 1998,40(7), 1169; Z. X. Sun, F. W. Su, W. Forsling and P. Samskorg,J. Colloid Interface Sci., 1998, 197, 151; U. Schwertmann andR. M. Cornell, Iron Oxides in the Laboratory: Preparation andCharacterization, 2nd edn., Wiley, New York, 2002.

27 J. M. Davey, C. O. Too, S. F. Ralph, L. A. P. Kane-Maguire,G. G. Wallace and A. C. Partridge, Macromolecules, 2000, 33, 7044.

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Fig. 5 Hysteresis loops for calix[n]arenes 1 and 2 coated magnetite

nanoparticles at 300 K.

1950 | Chem. Commun., 2007, 1948–1950 This journal is The Royal Society of Chemistry 2007

DOI: 10.1002/adfm.200701101

Size Selective Synthesis of Superparamagnetic Nanoparticles inThin Fluids under Continuous Flow Conditions**

By Suk Fun Chin, K. Swaminathan Iyer, Colin L. Raston,* and Martin Saunders

1. Introduction

Superparamagnetic magnetite (Fe3O4) nanoparticles are

important for a diverse range of applications such as, magnetic

resonance imaging, targeted drug delivery and magnetic

separation.[1] These applications require the nanoparticles to

be superparamagnetic with sizes smaller than 20 nm, and with

samples having a narrow size distribution to ensure the

particles have uniform physical and chemical properties.[2] The

synthetic protocols for gaining access to such particles range

from thermal decomposition of organometallic compounds in

high-boiling organic solvents in the presence of surfactants[3] to

synthesis involving the use of reverse micelles, where

surfactant-stabilized water-in-oil-emulsions control the shape

and size of the Fe3O4 nanoparticles.[4] However, these methods

cannot be applied to large-scale and economic production as

they require expensive or toxic organic reagents, and sequ-

ential and lengthy processing steps. A synthesis route for

ultra-large-scale production of monodispersed Fe3O4 nano-

crystals was recently reported.[5] However, the particles were

synthesized in the presence of oleic acid, which makes the

particles only dispersible in organic solvents and this limits

their bioavailability and hence their medical applications.

The most common cost effective and convenient way to

synthesize Fe3O4 nanoparticles is by co-precipitating ferrous

and ferric salt solutions with a base, such as aqueous NaOH

or NH4OH.[6] However, the size distribution of the Fe3O4

nanoparticles produced using this method is normally very

broad. Consequently, the downstream purification and isola-

tion process is more expensive and is time and energy intensive.

Furthermore, scale-up of this method using conventional

reactors can be problematic given the inhomogeneous

agitation and areas of localized pH variations, resulting in

the precipitation of non-magnetic iron oxides.[7]

Accordingly, there is a growing demand in the nanotechnol-

ogy industries for processes that promise to make dramatic

improvements in the design and performance of the manu-

facturing equipment involved. The concept of ‘‘Process

Intensification’’ offers alternative routes alleviating the

obstacles of the relaxed fluid dynamic regime associated with

conventional batch processes. Herein we demonstrate the

successful synthesis of Fe3O4 nanoparticles via co-precipitation

using NH3 gas as a base source using spinning disc processing

(SDP) under scalable and continuous flow conditions. To our

knowledge, this is the first use of NH3 gas as a precipitating

agent to make Fe3O4 nanoparticles in a thin fluid film. The

technology offers a realistic route towards large scale synthesis

of Fe3O4 nanoparticles with precise control within the 10 nm

size range.

The hydrodynamics of film flow over a spinning disc is

important in controlling the reactions. The demand for

intensified processing for which SDP is a subset has led to

the design and development of a range of reactors that offer

operating conditions with rapid heat and mass transfer under

continuous flow conditions with residence times reduced to

seconds rather than minutes or hours. SDP offers a novel

avenue for intensified nanotechnology via exploitation of high

centrifugal acceleration to generate thin films providing rapid

heat and mass transfers.[8] SDP is a form of process intensi-

fication where all reacting components are exposed to the same

conditions, in contrast to traditional batch technology where

conditions can vary across the dimensions of the vessel.[9] The

geometry and key elements of a SDP are illustrated in

FULLPAPER

Continuous flow spinning disc processing (SDP), which has extremely rapid mixing under plug flow conditions, effective heat

and mass transfer, allowing high throughput with low wastage solvent efficiency, is effective in gaining access to

superparamagnetic Fe3O4 nanoparticles at room temperature. These are formed by passing ammonia gas over a thin

aqueous film of Fe2þ/3þ which is introduced through a jet feed close to the centre of a rapidly rotating disc (500 to 2500 rpm),

the particle size being controlled with a narrow size distribution over the range 5 nm to 10 nm, and the material having very

high saturation magnetizations, in the range 68–78 emu g1.

[*] Prof. C. L. Raston, S. F. Chin, Dr. K. S. IyerCenter for Strategic Nano-FabricationSchool of Biomedical, Biomolecular and Chemical SciencesThe University of Western AustraliaCrawley, W.A. 6009 (Australia)E-mail: [email protected]

Dr. M. SaundersCenter for Microscopy, Characterization and Analysis,The University of Western AustraliaCrawley, W.A. 6009 (Australia)

[**] The authors are grateful for the financial support for this work by theAustralian Research Council, The University of Western Australia, andThe University of Malaysia Sarawak. The microscopy analysis wascarried out using facilities at the Centre for Microscopy, Characteriz-ation and Analysis, The University of Western Australia, which aresupported by University, State and Federal Government funding.Supporting Information is available online from Wiley InterScienceor from the authors.

922 2008 WILEY-VCH Verlag GmbH & Co. KGaA,Weinheim Adv. Funct. Mater. 2008, 18, 922–927

FULLPAPER

Figure 1a. The key components of SDP include: (i) a 100 mm

rotating disc with controllable speed (up to 3000 rpm), and (ii)

feed jets located at a radial distance of 5 mm from the center of

the disc. SDP generates a very thin fluid film (1 to 200 mm) on a

rapidly rotating disc surface, within which nano-particle

formation occurs.

The behavior of the thin film on a rapidly rotating disc can be

classified into two zones, Figure 1b. One corresponds to the

injection zone where the reagents hit the spinning disc to form

a pool at the centre of the disc. The pool is often referred to as

the spin-up zone and is related to viscous drag of the liquid film

associated with centrifugal forces.[10] The other zone cone

corresponds to the acceleration and synchronized flow zone

where the fluid film initially experiences an increase in radial

flow velocity and then the liquid is rotating close to the disc

velocity. The flow here becomes similar to the Nusselt

model.[11] The shear forces and viscous drag between the

moving fluid layer and the disc surface create turbulence and

ripples thus giving rise to highly efficient turbulent mixing

within the thin fluid layer.[12] The turbulent waves thus

generated can be a combination of circum-

ferential waves moving from the disc centre

to the disc periphery and helical waves,

depending on the operating parameters.

Controlling the levels of mixing associated

with the waves therefore offers the possibi-

lity of controlling the transport rates within

the thin film and hence controlling the

nanoparticle size and size distribution. The

thinness of the film contributes to many

influential chemical processing characteris-

tics, one being a very high surface area to

volume ratio, resulting in more favorable

interactions between the film and its sur-

roundings. These waves and ripples add to

the vigor of mixing, enabling very high heat

and mass transfer rates in the film. This, in

turn, ensures extremely short reaction

residence times enabling impulse heating

and immediate subsequent cooling, along

with plug flow conditions ensuring even

micro-mixing and transfer through the

entire film.[13]

2. Results and Discussion

The use of NH4OH aqueous solution as a

basic precipitation agent to synthesize

Fe3O4 nanoparticles on the spinning disc

was initially investigated (Two feed jets

operating at the feed rates of 0.5 ml s1þ0.5 ml s1). TEM analysis of the samples

revealed that the operating parameters,

including disc spinning speed had very little

effect on the particle size. However, it is

noteworthy that the size distribution of our samples was very

narrow, with the particle size ranging from 8–10 nm, Figure 2.

The co-precipitation of Fe3O4 is an instantaneous reaction[14]

and the lack of control over particle size suggests that the

reaction is complete within the initial injection or mix up zone

(zone 1, Fig. 1b) where the SDP operating parameters,

S. F. Chin et al. / Size Selective Synthesis of Superparamagnetic Nanoparticles

Figure 1. a) Schematic representation of a SDP, b) Hydrodynamics of the fluid flow over aspinning surface.

Figure 2. Typical TEM image of Fe3O4 nanoparticles a) (10 mM Fe2þ)synthesized using NH4OH solution at 2500 rpm and b) 50 mM Fe2þ at500 rpm.

Adv. Funct. Mater. 2008, 18, 922–927 2008 WILEY-VCH Verlag GmbH & Co. KGaA,Weinheim www.afm-journal.de 923

FULLPAPER

especially disc speed, have minimal effect on the outcome of

the reaction. The ability to control the size and size distribution

of the nanoparticles crucially relies on separating the

nucleation and growth processes. The rapid co-precipitation

of Fe3O4 in the presence of NH4OH aqueous solution often

inhibits such control to be realized in conventional processes.

The wavy thin films generated on a rotating disc surface[15] in

the presence of a gas, notably ammonia, offers the ability to

control the size of the ensuing particles by controlling the

delivery of base to the thin film. A recent report highlights that

waves generated in the fluid film over a moderately spinning

disc clearly enhance the gas adsorption into the liquid.[16] The

flow is accompanied by non-linear waves, which strongly

influence the diffusion boundary that develops beneath the

surface of the film. The progressive waves are generated in the

region of active micromixing (zone 2, Fig. 1b) and hence offer

the proposed control necessary to synthesize magnetic

nanoparticles with narrow size distributions.

Indeed, the operating parameters of the SDP were effective

in controlling the size, size distribution and shape of the Fe3O4

nanoparticles in the presence of NH3 gas feed as the base

rather than NH4OH aqueous solution. In a typical synthesis,

the samples were synthesized using 10 mM of Fe2þ and 20 mM

of Fe3þ aqueous solution and were fed at 1 ml s1 onto the

rotating grooved disc. A high rotation speed of 2500 rpm

resulted in ultrafine (3–5 nm) particles with asymmetrical

shape. A lower speed of 500 rpm, resulted in spherical

nanoparticles of around 10 nm in size, Figure 3. The fluid layer

of the SDP has a plug flow characteristic, where the particles

produced are constantly being removed from the nucleation

and growth zones by radial propagation across the disc. At high

rotation speeds the non-linear wave regime is dominant in the

fluid film, which in turn ensures greater gas adsorption into the

thin film. Consequently the nucleation process becomes

dominant, thereby resulting in a number of ultra-small

magnetic nanoparticles. At lower spin speeds the wave regime

is no longer predominant, and the velocity of the traversing

waves are highly reduced, and so is the absorption of the

corresponding NH3 gas into the flowing film. The number of

nucleation sites is thereby decreased and the growth process

becomes dominant at lower speeds resulting in the formation

of larger particles. Selected area electron diffraction, Figure 3,

shows sharp, well-defined diffraction rings, which reveals the

nanocrystallinity of the samples. The corresponding inter-

planar spacings calculated from the diffraction pattern are

presented in Table 1. The values obtained are consistent with

those obtained from the corresponding standard bulk Fe3O4

(Joint Committee on Powder Diffraction Standards, JCPDS,

card 19-0629).

At constant speed, samples synthesized using the grooved

disc had a narrower particle size distribution compared to the

smooth disc. The particle size for the samples synthesized on

the grooved disc ranged from 3–5 nm (Fig. 3a), whereas the

particle size distribution of a sample prepared using the smooth

disc ranged from 3–12 nm (Fig. 4a). When using the grooved

disc, the shear forces and viscous drag between the moving

fluid layer and the periodic grooved surface give rise to more

efficient turbulent mixing within the fluid layer and this in turn

results in homogeneous reaction conditions in the thin film.

The grooves on the disc also result in the formation of waves on

the flowing film, thereby amplifying the amount of the NH3 gas

adsorbed. However, on a smooth disc the micro-mixing is not

as efficient resulting in a broader size distribution. Increasing

the temperature of the disc offers scope to overcome the

aforementioned size distribution issues associated with the

smooth disc. At higher temperature the absorption efficiency

of the gas is reduced in the flowing fluids, however the growth

conditions become highly amplified.[17] As a result, the size

distribution of the samples prepared on the smooth disc

became very narrowwith particle sizes in the range 8–10 nm for

the reactions at 120 8C, Figure 4b.


Figure 3. TEM images of Fe3O4 nanoparticles (10 mM Fe2þ) synthesizedat 25 8C on grooved disc with disc rotating speed: a) 2500 rpm andb) 500 rpm.

Table 1. The d-spacing values (nm) calculated from the electron diffractionpatterns and the standard atomic spacing for Fe3O4 along with respectivehkl indices from the (JCPDS file No.19-629).

Calculated d spacing JCPDS data for Fe3O4 hkl

0.480 0.4852 111

0.299 0.2967 220

0.260 0.2532 311

0.210 0.2099 400

0.175 0.1714 422

0.166 0.1615 511

Figure 4. TEM images of Fe3O4 nanoparticles (10 mM Fe2þ) synthesizedon smooth disc with disc rotating speed of 500 rpm, at a) 25 8C and b)120 8C.

924 www.afm-journal.de 2008 WILEY-VCH Verlag GmbH & Co. KGaA,Weinheim Adv. Funct. Mater. 2008, 18, 922–927

FULLPAPER

In order to investigate the effect of precursor concentration

on the nanoparticle size and size distribution, the precursor

concentration in the feed was increased five times (Disc:

grooved; Feed rate: 1 ml s1). An increase in the concentration

resulted in a broader size distribution (comparing Fig. 5a vs.

Fig. 3a). Nevertheless, only 2 % of the particles are 10 nm,

while 98% of the particles are still around 5 nm. The

appearance of the larger particles may result from Ostwald

ripening of the particles, where the larger particles grow at the

expense of the smaller particles. Presumably this relates to less

uniform local concentration fields within the thin fluid film at

higher precursor iron concentrations where the driving force

for Ostwald ripening becomes more dominant resulting in the

appearance of larger particles. Increasing the disc temperature

to 120 8C resulted in spherical particles in the size range 10–15

nm, Figure 5b (Supporting Information: Fig. S1).

In the absence of surfactant, Fe3O4 nanoparticles tend to

agglomerate due to their extremely high surface energies

(>100 dyn cm1) coupled with the magnetic dipole–dipole

attractions between the particles.[18] Stabilization of nanopar-

ticles with biopolymers has generated enormous interest

recently, especially in imparting the bioavailability of such

particles for medical applications, in particular for drug

delivery. Alginic acid is a natural biopolymer of marine brown

algae which contains –COOH group and can bind covalently to

iron oxide nanoparticles,[19] and has been widely used in

biotechnological and biomedical applications.[20] Several

researchers have added alginic acid to preformed Fe3O4

nanoparticles in order to stabilize and improve their bio-

compatibility.[21] In the present study, we coated alginic acid

onto Fe3O4 nanoparticles in situ in a single-step during the

synthesis, resulting in the formation of stable suspensions of

superparamagnetite nanoparticles. The particles are spheroi-

dal with a diameter of approximately 10 nm, Figure 6a. The

isoelectric point of the sample is shifted to pH 1 (Fig. 6b),

which indicates that the alginic acid is coated on the Fe3O4

nanoparticles. FTIR spectra further confirmed the coating of

alginic acid on Fe3O4 nanoparticles, with the carboxylic acid


Figure 5. TEM image of Fe3O4 nanoparticles (50mMFe2þ) synthesized at2500 rpm, a) 25 8C, b) 120 8C.

Figure 6. a) TEM image of alginic acid coated Fe3O4 nanoparticles. b)Zeta potential of alginate coated Fe3O4 nanoparticles and uncoated Fe3O4

nanoparticles.

Table 2. Specific saturated magnetization of various Fe3O4 samples.

Sample a b c d E

Specific saturated magnetization, Ms [emug1] 68 76 70 78 73

Figure 7. Room temperature magnetization curves of various Fe3O4

samples synthesized at a) 2500 rpm, 25 8C, grooved disc; b) 500 rpm,25 8C, grooved disc; c) 500 rpm, 25 8C, smooth disc; d) 500 rpm, 120 8C,smooth disc; e) 500 rpm, 25 8C, grooved disc, coated with alginic acid.


FULLPAPER

functional groups converted to carboxylates, vCO 1610, and

1384 cm1 (Supporting Information: Fig. S2). It is noteworthy

that the alginic acid coated Fe3O4 nanoparticles are highly

stable in solution at physiological pH (pH¼ 7.4). This arises

from the high surface charge (>30 mV) and electrostatic

repulsion associated with conversion of the alginic acid

carboxylic acid moieties to the corresponding carboxylates

in the presence of ammonia. It also relates to steric

stabilization associated with covalent binding of carboxylate

moieties of reacted alginic acid with metal ion centers. The

hydrodynamic diameter of the alginic acid coated Fe3O4 as

measured from the DLS was approximately 24 nm (Supporting

Information: Fig. S3).

Figure 7 shows the room temperature magnetization curves

of as-prepared magnetite nanoparticles. The absence of

hysteresis loops with zero coercivity and zero remanence

indicate a typical of superparamagnetic behavior. Their

Ms values vary in the range from 68–78 emu g1, Table 2.

These values are lower than, but close to, the theoretical

Ms value for bulk magnetite of 92 emu g1,[22] but are higher

than previously reported experimental values of 30–50 emu g1

for similarly sized magnetite nanoparticles.[23] The effect of

particle size on the magnetic behavior of magnetic nano-

materials has been investigated by several researchers,[24–27]

and our results are consistent with the expected reduction in

Ms value associated with reduced particle size. The varia-

tion in our measurements could be a reflection of differences in

particle size between the samples. The remarkably high

Ms value of our samples suggests there is no chemical change at

the surface[28] and our observed values may be approaching the

limit of Ms for Fe3O4 nanoparticles with diameters less than

20 nm.

3. Conclusions

In conclusion, the particle size of Fe3O4 nanoparticles can be

controlled by judicious choice of the operating parameters of

SDP technology. Surfactants can be readily coated onto the

nanoparticles during the process intensification so that

stabilized nanoparticles can be generated in a single pass

under continuous flow, which is without precedent. The

capabilities of SDP in controlling particle size and stability

have been clearly demonstrated with the ability to produce

ultra-small superparamagnetic magnetite nanoparticles with

high saturation magnetizations, and with narrow size distribu-

tions. This approach is cost effective and environmentally

friendly throughout, including the choice of surfactant, and can

be scale up for commercial purposes, i.e., the research reactor

becomes the production reactor while maintaining narrow size

distributions.

4. Experimental

All the chemicals were of reagent grade andwere usedwithout furtherpurification. FeCl3 6H2O and FeCl2 4H2O were purchased from

Fluka. Alginic acid was purchased from Sigma. In a typical synthesis,aqueous solutions of Fe2þ/3þ precursors were prepared by dissolvingFeCl2 4H2O (10 mM) and (20 mM) FeCl3 6H2O (1:2 molar ratios) indeoxygenated ultrapure Mili-Q water. Higher concentrations of Fe2þ

(50 mM) and Fe3þ (100 mM) aqueous solution were used to study theeffects of concentration on the size of the nanoparticles. The SDPwas aProtensive 100 series with integrated feed pumps to direct the reactantsonto the rotating disc. The above solutions were delivered onto the discsurface using one feed jet at 1.0 ml s1, using continuous flow gearpumps (MicroPumps). Grooved and smooth stainless steel discs with100 mm diameter were used which were manufactured from 316stainless steel with the grooved disc having 80 concentric engineeredgrooves equally spaced at 0.6 mm depth. For the samples prepared inthe presence of alginic acid, the acid was fed onto the SDP using one ofthe feed jets during the synthesis of the Fe3O4 nanoparticles. Theweight of Fe3O4 nanoparticles that can be made depends on theconcentration of the reactants used. In a typical synthesis using a 100mlstarting solution of 10 mM of Fe2þ and 20 mM of Fe3þ, 0.3–0.4 gof Fe3O4 can be made with a flow rate of 1 ml s1.

The reactor chamber was purged with argon gas before the reactionto remove oxygen. Ammonia gas was then fed into the sealed reactorchamber at a constant flow rate. Black suspensions of Fe3O4

nanoparticles were collected from beneath the disc through an exitport.

The samples collected were immobilized with a permanent magnetand supernatant solutions were decanted. Samples were re-dispersed indeoxygenated ultrapure Mili-Q water. This process was repeated atleast three times to remove chloride salts and excess alginic acid.

The size and morphology of the Fe3O4 nanoparticles weredetermined using transmission electron microscopy (TEM, JEOL3000F) at an accelerating voltage of 300 kV. Size distributions weredetermined based on the average of the diameters from differentangles on the basis of at least 500 particle measurements per sample.

Zeta potentials and the average particle size of the of Fe3O4

nanoparticles coated with alginate were measured by dynamic lightscattering (Zetasizer Nano ZS series; Malvern Instruments).

The magnetic properties were measured using a Quantum Design 7T MPMS superconducting quantum interference device (SQUID)magnetometer at temperatures between 5 and 300 K. The iron contentof each samplewas determined usingAtomicAbsorption Spectroscopy(AAS) and the measured magnetization data was normalized withrespect to the iron content of the samples.

Received: September 24, 2007Revised: November 27, 2007

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[7] L. T. Vatta, R. D. Sanderson, K. R. Koch, J. Magn. Magn. Mater. 2007,

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[8] O. K. Matar, C. J. Lawrance, Phys. Fluids 2005, 17, 052102.

[9] a) K. V. K. Boodhoo, R. J. J. Jachuck, Green Chem. 2000, 2, 235.

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[16] G.M. Sisoev, O. K.Matar, C. J. Lawrance,Chem. Eng. Sci. 2005, 60, 7,

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PAPER www.rsc.org/loc | Lab on a Chip

Fabrication of carbon nano-tubes decorated with ultra finesuperparamagnetic nano-particles under continuous flow conditions†

Suk Fun Chin, K. Swaminathan Iyer and Colin L. Raston*

Received 19th October 2007, Accepted 3rd January 2008First published as an Advance Article on the web 31st January 2008DOI: 10.1039/b716195f

Ultra fine (2–3 nm) magnetite (Fe3O4) nano-particles are uniformly deposited on single-walledcarbon nano-tubes (SWCNTs) pre-functionalised with carboxylic acid groups using microwaveradiation. The deposition process involves chemical precipitation associated with continuous flowspinning disc processing (SDP), as a rapid, environmentally friendly approach which is readilyscalable for large scale synthesis. The resulting decorated SWCNTs are superparamagnetic withspecific saturated magnetization of 30 emu g−1.

Introduction

Carbon nano-tubes (CNTs) receive much attention becauseof their outstanding electronic, mechanical, thermal, chemicalproperties and significant potential applications in nanoscienceand nanotechnology.1 Recently functionalizing CNTs with ironoxide superparamagnetic nano-particles (magnetite Fe3O4 andc-Fe2O3 maghemite) have featured in many studies. This re-lates to their promising applications in the fields of fillers inpolymeric materials, nanoprobes for magnetic force microscopy,wastewater treatment, biosensors, and drug delivery.2–4 Variousmethods have been explored to attach iron oxide nano-particlesto CNTs. In the case of Fe3O4 nano-particles, this includes theuse of a carboxylic derivative of pyrene as an interlinker forthe attachment of Fe3O4 nano-particles to the carbon nano-tube surface, and combining polymer wrapping and layer-by-layer (LbL) assembly techniques and the electrostatic attractionbetween templated amino-functionalized carbon nano-tubesand Fe3O4 nano-particles.5–7 However, all these methods involvedown-stream processing of purification/separation and theattachment of iron oxide nano-particles onto the carbon nano-tubes is poorly controlled. Therefore scaling up the synthesis ofsuch composite material using this approach is difficult.

The alternative in situ synthetic approach is more promisingand efficient in coating CNTs with magnetic nano-particles.This includes solvothermal and high temperature thermaldecomposition of iron(III) acetylacetonate in the presence ofmultiwall carbon nano-tubes (MWCNTs) in polyol solutionleading to the formation of Fe3O4 nano-particles attached to thecarbon nano-tubes.8,9 Even though Fe3O4 nano-particles havebeen reported to be uniformly attached to the CNTs surfacewith high coating density, the major drawbacks of this approach

Centre for Strategic Nano-Fabrication, School of Biomedical,Biomolecular and Chemical Sciences, The University of WesternAustralia, 35 Stirling Highway, Crawley, WA, 6009, Australia.E-mail: [email protected]; Fax: +(618) 64881005;Tel: +(618) 64881572† Electronic supplementary information (ESI) available: Experimentaldetails of Fe3O4 synthesis using SDP, TEM images and magnetizationcurve of Fe3O4. See DOI: 10.1039/b716195f

are that the reaction temperatures are very high, toxic/expensiveorganic solvents are used, high time/energy consumption, andscaling up may be problematic. Some effort has focused on theuse of in situ chemical precipitation from solutions of Fe2+/3+

using NH4OH or NaOH to directly coat CNTs with Fe3O4

nano-particles, as a more environmentally benign, faster andmore economic method. However, due to the lack of control ofthe synthetic conditions associated with typical bench-top batchchemistry, the size distribution of Fe3O4 nano-particles attachedto the CNTs is very broad, ranging from 25–80 nm with lowcoating density. Moreover, the composite Fe3O4/CNT materialis ferromagnetic rather than superparamagnetic.10 The abilityto control both the particle size and size distribution of Fe3O4

nano-particles and their loading behavior on CNTs is of primaryimportance for tailoring the physical and chemical properties ofthese materials,11 yet it remains a synthetic challenge.

Spinning disc processing (SDP) involves a continuous flowreactor which has been recently shown to be effective in gainingaccess to a narrow size distribution of superparamagnetic Fe3O4

nano-particles in aqueous media using Fe2+/3+ and base (seeESI).† The efficient SDP capability of fabricating metal (Ag,Au, Pt) plated CNTs also has been demonstrated with themetal uniformly layered around single wall carbon nano-tubes(SWCNTs) or metal nano-particles of narrow size distributiondecorated on CNTs.12,13 Thus SDP has potential for preciselycontrolling the size and size distribution of magnetic nano-particles attached to CNTs. In this paper we describe a novel yetsimple method to coat superparamagnetic Fe3O4 nano-particlesof narrow size distribution on SWCNTs in situ by modifiedchemical precipitation method using SDP in aqueous mediaat room temperature under continuous flow conditions.

Experimental

Spinning disc processor

SDP, Fig. 1, is a rapid flash nano-fabrication technique with allreagents being treated in the same way, and is in contrast totraditional batch technology where conditions can vary acrossthe dimensions of the vessel.14 The reagents are directed towards

This journal is © The Royal Society of Chemistry 2008 Lab Chip, 2008, 8, 439–442 | 439

Fig. 1 Schematic representation of the synthesis of Fe3O4 coatedSWCNTs.

the centre of the disc, which is rotated rapidly (300 and 3000 rpm)resulting in the generation of a very thin fluid film (1 to 200 lm).The thinness of the fluid layer and the large contact area betweenit and the disc surface facilitates very effective heat and masstransfer. The drag forces between the moving fluid layer andthe disc surface enable very efficient and rapid mixing. Thegreatest strength of SDP synthesis is the broad range of controlpossible over all the operating parameters involved in nano-particle formation, enabling the simultaneous and individualoptimization of many interdependent operating mechanisms,with the ultimate goal of achieving very narrow particle sizedistributions.15 Another feature of SDP is that it is continuousflow, readily allowing scale-up of the ensuing product formation.

Materials and methods

All the chemicals were of reagent grade and were used withoutfurther purification. FeCl3·6H2O and FeCl2·4H2O were pur-chased from Fluka. Ultra pure water with a resistivity greaterthan 18 MX cm was obtained from a Milipure Mili-Q system.Single-walled carbon nano-tubes (Cheap Tubes Inc.) were usedas obtained.

For purification and functionalization of SWCNTs, 10 mg ofSWCNTs were dispersed in 5 ml of 1 : 1 mixture of 70% HNO3

and 98% H2SO4 aqueous solutions in the reaction chamber. Thereaction was carried out in a CEM Focused Microwave SynthesisSystem, Discover Model. The microwave power was set at 300 W,and the pressure was 12 bar, and the temperature was set at130 C for 30 minutes. After the reaction, the SWCNTs werefiltered, washed and re-dispersed in 100 mL of ultra pure Milli-Q water and sonicated for 15 min. The functionalized SWCNTswere dispersed in water and the container purged with N2 gasto remove oxygen. Then 10 mM of FeCl2·4H2O and 20 mM ofFeCl3·6H2O (1 : 2 molar ratios) were added and the mixturestirred for 1 h. After that, the solution was filtered to removeexcess Fe2+/3+ and the resulting carbon nano-tube and Fe2+/3+

complex was re-dispersed in deoxygenated Mili-Q water.A spinning disc processor 100 series (Protensive, Inc) was

used. Integrated feed pumps were used to feed (0.5 ml s−1) the re-actants on the rotating disc at 1000 rpm. Solutions/suspensionsof CNTs and Fe2+/3+ were fed from one feed and the deoxy-genated NH4OH aqueous solution was fed from the other

under an atmosphere of high purity argon gas (99.9%, BOCGasses), within the sealed reactor chamber. The disc surfacewas manufactured from 316 stainless steel. The 10 cm grooveddisc was used for the current study, with 80 concentric engineeredgrooves equally spaced approximately 0.6 mm in height. Sampleswere collected from beneath the disc through an exit port.

The size and morphology of the samples were determinedusing transmission electron microscopy (TEM, JEOL 3000F).Diffraction patterns were recorded using a TEM JEOL 2000FX II instrument. The FTIR spectra ware recorded using aPerkin–Elmer Spectrum One instrument. FTIR spectra wereobtained with KBr pellets and were recorded in the range4000–500 cm−1. The magnetic properties were measured usinga Quantum Design 7 T MPMS superconducting quantuminterference device (SQUID) magnetometer at temperaturesbetween 5 and 300 K. The iron content of each sample wasdetermined using atomic absorption spectroscopy (AAS) andthe measured magnetization data was normalized with respectto the iron content of the samples.

Results and discussion

The carboxylic functionalized carbon nano-tubes were readilydispersed in water for growth/attachment of Fe3O4 nano-particles. Conventional carboxylation of CNTs is carried outby refluxing a suspension of the CNTs in strong oxidizingagents such as concentrated HNO3 or H2SO4 for 10–50 h.16

However, the prolonged and vigorous acid treatment can resultin severe reduction in the length of the CNTs. Herein thecarboxylation (and purification) of SWCNTs involved the use ofmicrowave radiation for 30 min, thereby dramatically reducingthe reaction time and minimizing damage to the carbon nano-tubes.17 The SWCNTs are stable indefinitely in solution. SuchSWCNTs undergo loss of functionalized groups at 500 C withthe structural order on the sidewall being restored.

The functionalized SWCNTs are readily dispersible in water,with FTIR (Fig. 2) spectra confirming the presence of –COO− moieties. The peak at 1512 cm−1 corresponds to theasymmetric stretching mode of COO−, which as expected shiftsto 1452 cm−1 after coating with Fe3O4.18 The characteristic peakcorresponding to the stretching vibration for the Fe–O bondis also shifted to higher wavenumbers, 698 cm−1 compared to570 cm−1 reported for the stretching mode for Fe–O in bulkFe3O4,19 suggesting that Fe3O4 is bound to the COO− on theCNTs surface.

Fig. 2 FTIR spectra of COOH-functionalized SWCNTs and Fe3O4

coated SWCNTs.

440 | Lab Chip, 2008, 8, 439–442 This journal is © The Royal Society of Chemistry 2008

Ultrafine (2–3 nm) of Fe3O4 nano-particles with very narrowsize distribution were observed to be uniformly coated onto theCNTs surface, Fig. 3(a), (b) and (c). As can be seen fromthe TEM images, the distribution of Fe3O4 nano-particles onthe SMWNTs surface is very uniform and no local aggregationis observed with a very high coverage density. The ability toeffectively decorate the SWCNTs with Fe3O4 nano-particlesrelates to the functionalisation of the nano-tubes with COO−

moieties that can bind directly to Fe2+/3+ ions.20 The use of thissimple yet strategic approach reduces the down-stream process-ing dramatically. This is associated with purification/separationsteps to remove excess nano-particles that grow independentlyin solution and are not associated with the CNTs.

Fig. 3 TEMs of Fe3O4–SWCNT composite at (a and b) low resolutionand (c) high resolution, and (d) associated SAED pattern for materialsynthesized using SDP.

In stark contrast, when the experiment was carried out usingthe same chemistry via traditional batch processing, significantlylarger Fe3O4 nano-particles (8–10 nm) were observed to form onthe CNTs, Fig. 4(a) and (b). The particles formed random aggre-gates along the CNTs. It is believed that the non-homogenousmixing environment in a traditional reaction vessel coupledwith the tendency of CNTs to clump induces uncontrollednucleation and growth of nano-particles consequently resultingin the typical situation (Fig. 4a, b) that is well documented inthe scientific literature.

Fig. 4 TEM images of Fe3O4–SWCNT composite synthesized usingbench-top chemistry.

Table 1 The d-spacing values (nm) calculated from the electrondiffraction patterns and the standard atomic spacing for Fe3O4 alongwith respective hkl indexes from the JCPDS card (19-0629)


0.3093 0.2967 2200.2625 0.2532 3110.2165 0.2099 4000.1767 0.1714 4220.1673 0.1615 511

The strong shearing forces and viscous drag between themoving fluid layer and the disc surface of SDP gives rise to highlyefficient turbulent mixing within the fluid layer. The intensityof mixing plays a pivotal role in the precipitation mechanismand, hence, the nano-particle size and size distribution. Thefluid on the surface of the disc has a short and controllableresidence time. This results in an extremely rapid induction timefor nucleation and growth of the nano-particles. This, coupledwith the plug flow conditions on the disc, results in the nano-particles having similar growth conditions after nucleation. Thisis of paramount importance for synthesizing nano-particles withnarrow size distribution and avoiding the precipitation of non-magnetic iron oxide nano-particles.

Selected area electron diffraction (SAED), Fig. 3(d), showssharp, well-defined diffraction rings, which reveals the crys-tallinity of the nano-particles. The corresponding interplanarspacings calculated from the diffraction pattern are presented inTable 1. The values obtained are consistent with those obtainedfrom the corresponding standard bulk Fe3O4 (Joint Committeeon Powder Diffraction Standards, JCPDS, card 19–0629).

The magnetization curve of the SWCNTs material decoratedwith Fe3O4 nano-particles was measured at room temperature,Fig. 5. The sample exhibits superparamagnetic behaviour asevidenced by zero coercivity, zero remnance and the absence ofhysteresis loops. The specific saturation magnetization, Ms ofthe sample is 30 emu g−1. This value is smaller than the reportedvalue of bulk Fe3O4 of 92 emu g−1.21 The reduction in the valueof Ms could be attributed to the rather smaller size of the Fe3O4

nano-particles. However the Ms value is higher than the valuereported by others using the chemical precipitation method10

and comparable to the Ms value of those samples prepared bysolvothermal method.8

Fig. 5 Magnetization curve of Fe3O4–SWCNT composite at 300 K.

This journal is © The Royal Society of Chemistry 2008 Lab Chip, 2008, 8, 439–442 | 441

Conclusions

In summary, we have developed a novel, simple, rapid, costeffective and scalable method to decorate CNTs with 2–3 nmFe3O4 nano-particles using SDP. The resulting CNTs show highcoverage density. The approach also avoids the need for sub-sequent purification and separation of the composite materialfrom excess Fe3O4 nano-particles formed in solution, unlike inthe case of using batch reactions. Furthermore, the ability ofSDP to fabricate superparamagnetic SWCNT composites undercontinuous flow in scalable quantities is significant in any downstream applications.

Acknowledgements

The authors gratefully acknowledge support of this work bythe Australian Research Council, The University of WesternAustralia, and The University of Malaysia Sarawak. TheTEM work was carried out using facilities at the Centre forMicroscopy, Characterisation and Analysis, The University ofWestern Australia, which are supported by University, Stateand Federal Government funding. The authors also thank MrCameron Evans for help with Pov. Ray.

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2007, 7, 1121.14 P. Oxley, C. Brechtelsbauer, F. Ricard, N. Lewis and C. Ramshaw,

Ind. Eng. Chem. Res., 2000, 39, 2175.15 K. Swaminathan Iyer, C. L. Raston and M. Saunders, Lab Chip,

2007, 7, 1800.16 J. Liu, A. G. Rinzler, H. Dai, J. H. Hafner, R. K. Bradley, P. J. Boul, A.

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442 | Lab Chip, 2008, 8, 439–442 This journal is © The Royal Society of Chemistry 2008

Facile and Green Approach To Fabricate Gold and Silver CoatedSuperparamagnetic Nanoparticles

Suk Fun Chin, K. Swaminathan Iyer,* and Colin L. Raston*

Center for Strategic Nano-Fabrication, School of Biomedical, Biomolecular and Chemical Sciences,The UniVersity of Western Australia, Crawley, 6009 Australia

ReceiVed December 3, 2008; ReVised Manuscript ReceiVed February 6, 2009

ABSTRACT: Superparamagnetic magnetite nanoparticles are coated with homogeneous gold and silver shells using a simple aqueousbased seed mediated method at room temperature with dopamine as a surfactant. Nanoparticles of Au in the range 2-3 nm areattached to amine functionalized Fe3O4 nanoparticles, acting as seed for the growth of ultrathin Au or Ag shells. The monodispersedcore-shell nanoparticles Fe3O4@Au and Fe3O4@Ag have a particle size range of 10-13 nm with a shell thickness of approximately2-3 nm. They are magnetically purified and are superparamagnetic at 300 K with saturated magnetization values of 41 and 35 emug-1, respectively.

Introduction

Superparamagnetic iron oxide nanoparticles (SPIONs) con-sisting of maghemite (γ-Fe2O3) or magnetite (Fe3O4) haveattracted much interest due to their potential applications inmagnetic guided drug delivery,1 specific targeting and imagingof cancer cells,2 hyperthermia treatment of solid tumors,3 andcontrast enhancement agents in magnetic resonance imaging(MRI).4 However, the extent of biomedical applications ofSPIONs strongly depends upon their stability in physiologicalsolutions and the extent to which their surfaces can bechemically functionalized. Techniques for coating magneticnanoparticles with biocompatible layers have been widelystudied. Coating SPIONs with organic shell such as macrocyclicsurfactants5 and polymers6 or inorganic shell7 can enhance theirstability, dispersibility, and functionality of the otherwise nakedmagnetic nanoparticles. While coating of magnetic nanoparticleswith polymers and silica shell has been extensively studied, thereare limited reports on the coating of SPIONs with metallic shells.For biomedical applications, elemental gold and silver are idealcoating targets due to their low reactivity, high chemicalstability, and biocompatibility, as well as their affinity forbinding to amine (-NH2) or thiol (-SH) terminal groups oforganic molecules.8 Moreover, Au and Ag shells also renderplasmonic properties to SPIONs,9 with gold coated particlesimparting a 6-fold enhancement in trapping efficiency anddetection sensitivity as compared to similar-sized polystyreneparticles. In addition, the absorption of irradiation by gold atthe most common trapping wavelength (1064 nm) imparts adramatic heating of the particles (266 °C/W),10 with Au or Agcoated SPIONs serving as excellent heating source followingmagnetically targeting a tumor.11,12

The synthesis of Fe3O4@Au/Ag core-shell nanoparticles ischallenging given the difference in surface energies of the twomaterials.13 Consequently, gold and silver metals tend tonucleate rapidly forming discrete nanoparticles in solutionwithout coating the surface of magnetite. γ-Fe2O3@Au orpartially oxidized Fe3O4@Au core-shell nanoparticles (∼60nm) have been reported by depositing Au onto the surfaces of9 nm particles involving the use of aqueous hydroxylamine.14

Core-shell nanoparticles of Fe3O4@Au and Fe3O4@Ag ranging

from 18 to 30 nm in size have also been prepared, using areverse micelle method,15 but are capped with organic surfac-tants which renders them unsuitable for biomedical applicationsas they disperse in organic solvents. Recently, hydrophobicFe3O4@Au and Fe3O4@Au@Ag core-shell nanoparticles havebeen prepared by reducing HAuCl4 and AgNO3 in a chloroformsolution of oleylamine as a surfactant. Following this they canbe transferred to the aqueous phase by mixing them with anaqueous cetyltrimethylammonium bromide (CTAB).16 Overallit is evident that a simple aqueous based synthetic method tocoat Fe3O4 nanoparticles with uniform and relatively thin layersof noble metal (Au, Ag) shells that have minimal purturbationson the magnetic properties for applications in biomedical fieldis an important objective. Herein, we report a strategic route tocoat SPIONs with 2-3 nm thick homogeneous Au and Ag shellsin aqueous solution at room temperature. This straightforwardapproach affords core-shell nanoparticles with superparamag-netic properties, at the same time rendering plasmonic propertiesto the nanoparticles. Refluxing nanoparticles of Fe3O4 with (3-aminopropyl)triethoxysilane (APTES) for a few hours resultsin functionalizing the surface of the particles with -NH2 groups.However, these nanoparticles are unstable in aqueous solution,and the synthetic procedure is time-consuming and requires hightemperatures.17 We recently used sulfonated calixarenes totemplate and stabilize nanoparticles of Fe3O4 where the phenolicgroups at the base of the calixarenes bind to surface metalcenters.5 Dopamine, Scheme 1(a), also has phenolic groupswhich can behave similarly, albeit with now two adjacant -OHgroups on the same aromatic ring which form a chelate ring toa metal center, as expected in the deprotonated form, with theirterminal primary amine groups poised to bind to other metalcenters.18,19 The utility of dopamine as such a surfactant featuresin the present study, as a linker molecule for metal centers onthe surface of Fe3O4 and Au or Ag, Scheme 1. We find that thedopamine can be easily attached to the iron oxide nanoparticlesin a single-step process by sonicating the nanoparticles withdopamine in aqueous solution at room temperature at pH ∼ 9.The resulting amine functionalized Fe3O4 nanoparticles arestable in aqueous solutions for months, with no evidence ofprecipitation, thereby circumventing the need for using othersurfactants. The presence of dopamine on the nanoparticles wasestablished using FTIR as evidenced by the peaks characteristicof -NH2 at 1617, 1600, and 1585 cm-1 (see Figure S1 in theSupporting Information). The Fe3O4 nanoparticles were prepared

* Corresponding authors. Fax: (618) 64881005. Tel: (618) 64881572. E-mail:[email protected]; [email protected].

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10.1021/cg8013199 CCC: $40.75 2009 American Chemical SocietyPublished on Web 04/10/2009

using spinning disk processing (SDP), which has been usedeffectively for size selective synthesis of Fe3O4 nanoparticles.20-24

Initial attempts to reduce Au or Ag precursor salts directly onto the -NH2 functionalized Fe3O4 nanoparticles (pH ∼ 9), usingglucose as a mild reducing agent, resulted in uncontrollednucleation of discrete nanoparticles of Au or Ag attached toFe3O4 nanoparticles, rather than forming a continuous layer ofthe metal (see Figure S2 in the Supporting Information). Wealso observed that when reducing agents such as hydrazine orascorbic acid were used, the large driving force for reductionof metals led to nucleation of free Au or Ag nanoparticlesevident by discrete Au or Ag nanoparticles in solution orattached to the surface of Fe3O4 nanoparticles.

The strategy used herein involves a “seed-mediated growth”method25 whereby preformed 2-3 nm Au nanoparticles wereattached to the -NH2 functionalized Fe3O4 nanoparticles, withAu3+ or Ag+ reduced onto the surface of the compositenanoparticles for complete encapsulation in ultathin shells. Thisinvolved the use of glucose as a mild reducing agent. Theseeding stage is necessary to overcome the difference in surfaceenergies of the Au or Ag, and Fe3O4 with the Au seeds acts asnucleation sites for deposition of Au or Ag shells. The resultingnanoparticles can be represented as Fe3O4@Au and Fe3O4@Ag,but only the former has a homogeneous layer of metal, withthe silver layer also containing the original nucleating Aunanoparticles. A permanent magnet was put at the side of avial containing the as-synthesized samples to magneticallyseparate the Fe3O4@Au and Fe3O4@Ag from any free Au orAg nanoparticles. The magnetically purified samples wereredispersed in phosphate buffer solution (PBS) by sonication,as stable solutions.

Experimental Section

Materials. Analytical grade ferric chloride (FeCl3 ·6H2O), ferrouschloride (FeCl2 ·4H2O) (Fluka), ammonia solution and silver nitrate(AgNO3) (Ajax Chemicals), chloroauric acid (HAuCl4) (AGR Matthey),and dopamine and tetrakis(hydroxymethyl)phosphonium chloride (THPC)(Aldrich) were used as obtained. Water with a resistivity greater than18 MΩ cm was acquired from a Millipure Milli-Q system.

Synthesis of Fe3O4 Nanoparticles. Fe3O4 nanoparticles weresynthesized using spinning disk processing with Fe2+/3+ in one jet feedand aqueous NH4OH in another, as detailed in the literature.20 Sizedistributions were determined based on the average of the diametersfrom different angles on the basis of at least 500 particle measurementsper sample. The average size of the as-synthesized Fe3O4 nanoparticlesis 8 ( 2 nm. Dopamine was added to the colloidal suspension of Fe3O4

nanoparticles, and the mixture was sonicated to form a stable suspensionof the ferrofluid.

Attachment of Au Seeds to Dopamine Functionalized Fe3O4

Nanoparticles. The 2-3 nm Au seed solution was prepared based onthe reported method with some modification.25 In a typical synthesis,5 mL of 1 M NaOH was diluted with 18 mL of Mili-Q water and thenmixed with 10 mL of tetrakis(hydroxymethyl)phosphonium chloride(THPC; 0.67 mmol) solution. The THPC stock solution was preparedseparately by diluting 0.6 mL of 80% THPC (3.375 mmol) aqueoussolution with 50 mL of Mili-Q water. The solution mixture was heatedto 50 °C and stirred vigorously to allow for complete mixing. Then,20 mL of 1% HAuCl4 solution was added dropwise to the above mixturekept under vigorous stirring. The color of the mixture turned deep red-brown, indicating the formation of Au seeds nanoparticles.

Twenty-five milliliters of the Au seed suspension was added to 25mL of dopamine coated Fe3O4 nanoparticles (1 mM), and the mixturewas stirred overnight, whereupon it was centrifuged to remove freeAu seeds. The precipitate was redispersed in Mili-Q water and sonicatedfor one hour.

Deposition of Au or Ag Shell onto Dopamine Coated Fe3O4

Nanoparticles. For shell growth, 5 mL (10 mM) of HAuCl4 aqueoussolution was added to the 50 mL Au seed decorated Fe3O4 (1 mM)nanoparticles under mixing. The pH of the mixture was adjusted toslightly basic (pH ∼ 9) to facilitate the reduction process. 1.5 timesexcess of glucose solution was added dropwise to the solution to reducethe HAuCl4 onto the Au seed decorated Fe3O4 nanoparticle surface.The same procedure was followed for Ag shell growth, with 5 mL (10mM) of AgNO3 solution in place of HAuCl4. The synthesizedFe3O4@Au or Fe3O4@Ag core-shell nanoparticles were magneticallyseparated from the free Au or Ag nanoparticles and redispersed inMili-Q water for further characterization.

Characterization

Transmission electron microscopy (TEM) images wererecorded using a JEOL 3000F at 300 kV.

Scheme 1. Schematic Illustration of (a) the Attachment of Dopamine onto the Surface of Fe3O4 Nanoparticles and (b) theSynthesis Process of Fe3O4@Au and Fe3O4@Ag Core-Shell Nanoparticles

2686 Crystal Growth & Design, Vol. 9, No. 6, 2009 Chin et al.

The FTIR spectrum was recorded in the transmission modeon a Perkin-Elmer Spectrum One spectrometer. The driedsample was ground with KBr, and the mixture was compressedinto a pellet. The spectrum was taken from 4000 to 400 cm-1.

X-ray powder diffraction patterns were recorded on a SiemensD5000 diffractometer with Bragg-Brentano geometry and CuKR radiation. The data were collected from 2θ ) 20° to 90°.

UV-visible spectroscopy was performed on a Perkin-ElmerLambda 25 UV/vis spectrometer using matched quartz cuvetteswith a path length of 1 cm.

The magnetic properties were measured using a QuantumDesign 7 T MPMS superconducting quantum interference device(SQUID) magnetometer at 300 K.

Results and Discussion

Figure 1a shows the TEM image of Fe3O4 nanoparticles andthe corresponding selected area electron diffraction pattern.Well-defined diffraction rings reveal the nanocrystallinity of thesamples. The corresponding interplanar spacings calculated fromthe diffraction pattern are presented in Table 1 in the SupportingInformation. The values obtained are consistent with thoseobtained from the corresponding standard bulk Fe3O4 (JointCommittee on Powder Diffraction Standards, JCPDS, card 19-0629). The synthesis of ultrasmall Au seeds is essential to adoptthe strategy presented in the current work (Figure 1b), whichinvolved synthesis at 2-3 nm in size with a narrow sizedistribution.26 It is also evident that the dopamine functional-

ization of the Fe3O4 nanoparticles resulted in a highly efficientattachment of the seeds on the individual nanoparticles (Figure1c).

The subsequent reduction of Au3+ or Ag+ using glucose asthe reducing agent in aqueous solutions resulted in the formationof a continuous thin Au or Ag layer at the surface of the Fe3O4

nanoparticles, Figure 2a,b. Analysis of the Fe3O4@Ag andFe3O4@Au core-shell nanoparticles and uncoated Fe3O4 nano-particles (Figure 1a) shows that both the Au and Ag shells havea thickness of 2-3 nm. The darker contrast in the central partof the particles corresponds to a size in the range of 8-10 nm(Figure 2a, inset) which is consistent with the size of the startingFe3O4 nanoparticles. The periphery of the particles shows theuniform Ag shells with a thickness of about 2-3 nm. Thecore-shell structure is harder to observe in Fe3O4@Au core-shellnanoparticles due to the much heavier atomic mass of Auresulting in a nondiscernible mass contrast between the coreand the shell. Both the Fe3O4@Au and Fe3O4@Ag nanoparticleswere observed to be more spherical in shape after being coatedwith Au and Ag relative to the uncoated Fe3O4 nanoparticles,which are irregular in shape. The average size of the core-shellparticles is 12 ( 3 nm. Clearly, the TEM images serve asimportant evidence supporting the formation of Fe3O4@Au andFe3O4@Ag core-shell nanoparticles.

Measurements of the surface plasmon (SP) resonance bandof the nanoparticles provide complementary evidence of theFe3O4@Au and Fe3O4@Ag core-shell structure. Excess elec-trons result in the plasmon absorption shift to shorter wave-length, whereas electron deficiency will shift the absorption to

Figure 1. Typical TEM images of (a) Fe3O4 nanoparticles (inset: SAD pattern), (b) Au seed nanoparticles, and (c) Au seed decorated Fe3O4

nanoparticles.

Figure 2. TEM images of (a) Fe3O4@Ag core-shell nanoparticles and (b) Fe3O4@Au core-shell nanoparticles. Insets are the high magnificationof the core-shell nanoparticles.

Au and Ag Coated Superparamagnetic Nanoparticles Crystal Growth & Design, Vol. 9, No. 6, 2009 2687

longer wavelength in the case of Au. The interface betweenAu and Fe3O4 has been reported to lead to an electron deficientenvironment.27 Both Au seed decorated Fe3O4 nanoparticles andFe3O4@Au nanoparticles give rise to a plasmon resonance peakat 530 nm. The corresponding peaks showed a red-shift of about10 nm in comparison with the characteristic Au nanoparticlepeak at 520 nm. Similar shifts have been reported for Fe2O3@Aunanoparticles prepared using other synthesis methods28 andSiO2@Au nanoparticles.29 In the case of Fe3O4@Ag nanopar-ticles, the plasmon resonance peak shifted from the characteristicplasmon resonance peak of Ag nanoparticles at 420 to 450 nm,Figure 3. The red-shift in wavelength of Ag surface plasmonhas also been observed experimentally in silica@Ag core-shellnanoparticles.30

XRD patterns for both Fe3O4@Au and Fe3O4@Ag core-shellnanoparticles show characteristic peaks (at 2θ ) 30.0°, 35.3°,43.0°, 53.3°, 56.9°, 62.5°), marked with their indices (220),(311), (400), (422), (511) and (440), Figure 4. These indicatethat the nanoparticles were pure Fe3O4 with a crystalline inversespinel structure. Curve (a) in Figure 4 has diffraction peaks at2θ ) 38.2°, 44.4°, 64.6°, and 77.5°, which can be indexed to(111), (200) and (311) planes of gold in the cubic phase. Curve(b) shows peaks at 2θ ) 38.0°, 44.2°, 77.3°, 81.5° whichcorrespond to the (111), (200), (311) and (222) indices for puresilver in the cubic phase.

The room temperature magnetization versus applied field(M-H) measurements at 300 K for both Fe3O4@Au andFe3O4@Ag core-shell nanoparticles indicated zero coercivityand remanence, Figure 5. The enlarged view of the graph inthe inset indicates the absence of a hysteresis loop. This isconsistent with the superparamagnetic properties arising fromthe small size of magnetic core (<20 nm) used in thesynthesis. The coating of the Au and Ag shell on the Fe3O4

nanoparticles did not perturb the superparamagnetic propertiesof the Fe3O4 nanoparticles with the saturated magnetization,Ms, of Fe3O4@Au and Fe3O4@Ag being 41 and 35 emu g-1

respectively. These values are lower than the reported 92emu g-1 29 for bulk Fe3O4 and the 78 emu g-1 of bare Fe3O4

nanoparticle reported in our earlier studies.20 The decreasein saturated magnetization values could be due to thecontribution of overall mass from the nonmagnetic Au andAg shells.

Conclusions

In conclusion, we report a strategic route to prepare mono-dispersed core-shell Fe3O4@Au or Fe3O4@Ag nanoparticlesusing a simple aqueous method at room temperature. A layerof thin and uniform Au and Ag shells is formed on the surfaceof Fe3O4 nanoparticles. Furthermore the synthetic methodemployed herein attempts to address sustainability metrics atthe inception of the science in using water as the solvent andreadily available dopamine as the surfactant, and glucose as amild reducing agent. The resulting nanoparticles are superpara-magnetic with good magnetization values. This coupled withtheir plasmonic properties gives them potential in diagnosticand therapeutic applications.

Acknowledgment. The authors are grateful for the financialsupport for this work by the Australian Research Council andThe University of Western Australia, and for the gold from AGRMatthey. S.F.C. acknowledges postgraduate scholarship fromThe University of Malaysia Sarawak. The microscopy analysiswas carried out using facilities at the Centre for Microscopy,Characterisation and Analysis, The University of WesternAustralia, which are supported by University, State and FederalGovernment funding.

Figure 3. UV-vis absorption spectra of the nanoparticles.

Figure 4. X-ray diffraction (XRD) pattern of (a) Fe3O4@Au and (b)Fe3O4@Ag nanoparticles. * ) Fe3O4, # ) Au, 4 ) Ag.

Figure 5. Magnetization curves of Fe3O4@Au and Fe3O4@Ag core-shellnanoparticles at 300 K. Inset is the enlarged view of the graph from-10000 to +10000 Oe and -20 to + 20 emu g-1.

2688 Crystal Growth & Design, Vol. 9, No. 6, 2009 Chin et al.

Supporting Information Available: FTIR spectra of dopaminecoated Fe3O4 and TEM images and d spacing values of Fe3O4. Thismaterial is available free of charge via the Internet at http://pubs.acs.org.

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CG8013199

Au and Ag Coated Superparamagnetic Nanoparticles Crystal Growth & Design, Vol. 9, No. 6, 2009 2689

DOI: 10.1002/chem.200802747

Encapsulation and Sustained Release of Curcumin using SuperparamagneticSilica Reservoirs

Suk Fun Chin,[a] K. Swaminathan Iyer,*[a] Martin Saunders,[b] Tim G. St. Pierre,[c]

Craig Buckley,[d] Mark Paskevicius,[d] and Colin L. Raston*[a]

Curcumin is a yellow polyphenol found in the rhizomes ofthe plant Curcuma longa. The compound has shown promisetowards wound healing, which is attributed to the presenceof myofibroblast, and in enhancing fibronectin and collagenexpression.[1] The treatment of wounds with curcumin hasbeen reported to increase the formation of granulationtissue, which includes greater cellular content; neo-vasculari-sation; and faster re-epithelialisation of wounds.[2] Thesefindings suggest that curcumin may be able to improve radi-ation-induced wound repair delay. Curcumin has also beenreported to show antioxidant,[3] anti-inflammatory,[4] antimi-crobial,[5] and anticancer capability.[6] Various animal modelsor human studies also showed that curcumin is extremelysafe even at a dosage as high as 12 g per day.[7] However, allof the above-mentioned properties of curcumin are yet tobe fully realised, due to its low water solubility, fast degra-dation, and poor bioavailability. Oral administration of cur-cumin at a dose of 2 g kg1 to rats, only resulted in a maxi-mum serum concentration of 1.35 (0.23 mg mL1 after ap-proximately 1 h, whereas in humans the same dose of curcu-min resulted in either undetectable or extremely low

(0.005 mg mL1 at 1 h) serum levels.[8] Attempts have beenmade to encapsulate curcumin in polymeric micelles, lipid-based nanoparticles, and hydrogels in order to improve itswater solubility, stability, and bioavailability.[9] However, or-ganic-based carriers such as polymeric nanoparticles, lipo-somes, and micelles are shown to suffer from poor stabilityowing to biochemical attack and swelling.[10] On the otherhand, silica-based drug-delivery carriers are chemically andthermally more stable, highly hydrophilic, and biocompat-ible. Furthermore, the high density of silanol groups locatedat the silica surface (pores and external surface) can bereadily treated with coupling agents to provide sites for teth-ering specific bioactive substrates including antibodies.[11]

Targeted drug delivery offers the advantage to safely andeffectively deliver desirable dosages of drugs to specific siteswithout adverse side effects. One strategy for targeting drugdelivery is to use an external magnetic field to guide mag-netically labelled drug carriers. For magnetic-targeting drugdelivery, a drug or therapeutic molecule is bound to a mag-netic material, introduced in the body, and then concentrat-ed in the target area by means of a magnetic field. Suchmagnetic carriers are an attractive approach for site-specificdelivery of drugs with the ability to concentrate them on de-sired cells or organs by an external magnetic field. They canbe subsequently removed after treatment thereby improvingthe efficiency of the treatment and reducing toxic side ef-fects. Moreover magnetic forces act at relatively long rangeand magnetic fields do not adversely affect most biologicaltissues. A few synthetic approaches have been reported forthe preparation of magnetic silica particles, such as the useof microemulsions,[12] sol-gels,[13] and backfilling.[14] However,in the case of microemulsions, much effort is required toseparate the particles from the large amount of organic sur-factants and solvents used. The backfilling method risksclogging the silica pores and consequently resulting in a de-crease of available surface area. Block co-polymer templat-ing methods have also been used to develop the internalmesopore structure, while simultaneously incorporating themagnetic nanoparticles into the silica matrix.[15] Here ther-

[a] S. F. Chin, Dr. K. S. Iyer, Prof. C. L. RastonCentre for Strategic Nano-Fabrication,School of Biomedical, Biomolecular and Chemical SciencesThe University of Western Australia, Crawley, WA 6009 (Australia)Fax: (+61) 86488-8683E-mail : [email protected]

[email protected]

[b] Prof. M. SaundersCentre for Microscopy, Characterisation and AnalysisThe University of Western Australia, Crawley, WA 6009 (Australia)

[c] Prof. T. G. St. PierreCentre for Strategic Nano-Fabrication, School of PhysicsThe University of Western Australia, Crawley WA 6009 (Australia)

[d] Prof. C. Buckley, M. PaskeviciusDepartment of Imaging and Applied PhysicsCurtin University of Technology, PO Box U1987Perth, Western Australia, WA 6845 (Australia)

Supporting information for this article is available on the WWWunder http://dx.doi.org/10.1002/chem.200802747.

Chem. Eur. J. 2009, 15, 5661 – 5665 2009 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim 5661

COMMUNICATION

mal treatment at 500–550 8C for several hours is needed toremove the co-polymers surfactants used in the syntheses,which can have a detrimental effect on the magnetic proper-ties of the composite materials.[16] In addition, the reportedsaturated magnetisation values of the silica mesoporouscomposites prepared as such is low due to the low loadinglevel of magnetic nanoparticles (<5 %), thereby limitingany practical applications. Overall, there is still considerableinterest in developing novel methods for preparing magnet-ic, porous, silica-based particles possessing useful magneticproperties, as well as exhibiting high surface areas.

Ozin et al. described the origami synthesis of well-formedmesoporous silica particles in the shape of discoids, spheres,or fibres. These particles were synthesised at room tempera-ture from TEOS/CTACl/HCl/H2O/HCONH2 sol and theirformation mechanism involved polymerisation-induced dif-ferential contraction of a patch of a hexagonal silicateliquid-crystal film formed at the air-water interface, whichcan fold into particles by somewhat mimicking “origami”folding.[17] Using this unique formation Sokolov et al., re-cently established a highly effective mechanism to storedyes in these mesoporous structures.[18] Recently we showedthat hydrophobic curcumin can also be effectively encapsu-lated into porous silica capsule through a simple multistepself-assembly approach.[19] The particles arise by the initialformation of surfactant micelles of cetyltrimethylammoniumbromide (CTAB), which aggregate and form micellar rods.The micellar rods loaded with curcumin template the forma-tion of porous silica by polymerisation/hydrolysis of tet-raethyl orthosilicate (TEOS) under acidic condition. Theabove encapsulation of curcumin in silica particles improvesits photostability and affords a controlled release mechanismunder physiological pH. In the present study, we report theformation of magnetic nanocomposites of silica with curcu-min in the CTAB micellar rod in the presence of superpara-magnetic Fe3O4 nanoparticles (8–10 nm). The currentmethod endows ultrahigh loading of magnetite inside theporous silica matrix. This high loading of Fe3O4 nanoparti-cles in the silica carriers will impart high magnetophoreticmobility for targeted delivery. Such magnetic carriers offeran attractive approach for site-specific delivery of curcuminwith the ability to concentrate them on desired cells ororgans by an external magnetic field and increase their resi-dence time in the vicinity of the target area.

The method herein for preparing the composite magneticparticles is different from other methods in the simplicity ofincorporating a high loading of Fe3O4 nanoparticles (37 %wt) and curcumin (30 % wt) into the porous silica matrixunder mild conditions at room temperature. Small-angle X-ray scattering patterns for the samples before magnetic sep-aration shows a distinctive Bragg peak corresponding to or-dered porous nonmagnetic silica structures (Figure 1 A). Thepeak is absent after magnetic separation to remove nonmag-netic material. This indicates that the porous silica matrixaround the magnetic aggregates is non-ordered. This wasfurther confirmed by TEM analysis of microtomed samples.In the absence of Fe3O4 nanoparticles the silica formed

highly ordered mesoporous structures (Figure 1 B), while inthe presence of the same, a silica sheath surrounds the mag-netic clusters (Figure 1 D). The TEM image of the micro-tomed samples also shows that the Fe3O4 nanoparticles aredensely packed inside the silica matrix. A polydispersedcore with constant shell thickness model from NIST[21] wasused to fit the SAXS data for the magnetically separatedsample (see Figure 1 C). The model assumes a polydisperseFe3O4 spherical core surrounded by a curcumin shell embed-ded in a silica matrix resulting in an average Fe3O4 core di-ameter of 7.13 nm (variance=1.89 nm) and a curcumin shellof thickness 2.590.07 nm. In the model, the single-particlescattering amplitude is appropriately averaged over theSchulz distribution of iron core radii.[22] The first six datapoints are not included in the fit as this scattering arisesfrom larger structures in the sample. It is therefore evidentthat the presence of nanoparticles interrupts the order re-sulting from the polymerisation-induced differential contrac-tion of the hexagonal silicate liquid-crystal film formed atthe air–water interface, which in its absence can fold intohighly ordered mesoporous structures.

SEM analysis revealed that the Fe3O4 nanoparticle con-taining silica particles were ellipsoidal in shape, Figure 2 A.The size of the particles ranged from 200 nm to 1 mm. Ele-mental mapping (Figure 2 B–E) of the microtomed sampleunder the TEM showed that each individual iron oxidenanoparticle is held together by a layer of carbon-rich mate-rial and a silicon- and oxygen-rich shell.

Encapsulation of curcumin within the silica matrix wasconfirmed by confocal laser scanning microscopy and UV/Vis spectroscopy. As curcumin is naturally fluorescent in thevisible green spectrum, the silica particles appeared fluores-cent under the confocal microscope. Furthermore, UV/Visspectra of the particles showed a typical absorbance band of

Figure 1. A) SAXS spectra of magnetically separated silica particles andas-synthesised silica particles; upper curve, curcumin/Fe3O4/silica; lowercurve, magnetically separated sample; B) TEM image of a free silica par-ticle. C) Polydispersed core with constant shell-thickness model fit to themagnetically separated sample. + : curcumin/Fe3O4/silica; cmodel fit.D) TEM image of microtomed sample showing Fe3O4 particles surround-ed by a silica matrix.

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curcumin at 420 nm. (see the Supporting Information S1and S2). These results indeed confirm that curcumin was en-capsulated inside the silica sheath. The concentration of thecurcumin in the mesoporous silica was back-calculated bycomparing to the standard curve of curcumin in ethanol,being established at 30 % wt. The Brunauer–Emmett–Teller(BET) analysis showed that the surface area of silica parti-cles in the presence of both curcumin and Fe3O4 nanoparti-cles is 55.7 m2 g1. However, the silica particles loaded withFe3O4 nanoparticles in the absence of curcumin had a BETsurface area of 328.7 m2 g1. The reduction of BET surfacearea of the curcumin-loaded silica particles suggests that thecurcumin is successfully loaded inside a porous silica matrix(pore size determined from BET is 5.1 nm), with the curcu-min surrounding the magnetite nanoparticles, as identifiedusing TEM, Figure 2. It is worth noting that the as-formedcomposite materials were washed with water and ethanol(see experimental section) prior to analysis. It was previous-ly reported that ethanol treatment is a crucial step in leach-ing the dye from mesoporous silica particles. It is believedthat ethanol reacts with silica surface partially breakingopen the coiled mesopores.

The controlled release capability of the magnetic silicacapsules was analysed by leaching the curcumin in phos-phate buffer at physiological pH (pH 7.6). The UV spectraof the buffer shows an increase in absorbance with time ofleaching the curcumin at 420 nm (Figure 3). The curcuminlocated inside the channels of CTAB slowly diffused outfrom the silica capsules through passive diffusion processes.These surfactant molecules (CTAB) also act as stabilisingcages, decelerating the diffusion rates of the drugs locatedinside the channels, thereby maintaining a steady-state diffu-sion out into the buffer solution. Magnetisation versus ap-plied field measurements at room temperature indicatedzero coercivity and remanence, Figure 4. The magnetic be-haviour is consistent with the expected superparamagneticproperties owing to the small size of the magnetic cores(<20 nm) embedded in the mesoporous silica bodies. Themass-specific saturation magnetisation (Ms) of the magnetic

silica particles was around 22 emug1 before and after therelease of curcumin. The Ms was around 60 emug1 whennormalised to the Fe3O4 content. This value is close to thespecific saturation magnetisation of bare 8–10 nm Fe3O4

nanoparticles reported in our earlier publication.[23] It is alsoworth noting that the encapsulation of Fe3O4 nanoparticlesin silica did not change the superparamagnetic property ofthe Fe3O4 nanoparticles.

Figure 2. A) SEM image of magnetic silica particles. B) TEM image ofmicrotomed sample of magnetic silica particles. C) Oxygen map. D) Ironmap. E) Silicon map. F) Carbon map of the circled area in B.

Figure 3. A) UV absorbance for a supernatant liquid containing the silicacomposite material under physiological pH at different time intervals—from bottom to top: 0, 1, 24, 48, 96 h. B) Release profile of curcuminfrom superparamagnetic silica particles.

Figure 4. A) Magnetisation curve of as synthesised magnetic silica parti-cles (black) and magnetisation curve of magnetic silica particles after therelease of curcumin (grey).

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In conclusion, we have developed a simple and strategicroute to encapsulate curcumin inside superparamagneticporous silica particles. Such structures are potential proto-types for materials for use in targeted drug delivery. Whilstcurcumin has been reported for its excellent antitumor/anti-cancer and anti-inflammatory properties, along with its abili-ty to enhance wound healing, its use is limited, because ofits otherwise poor bioavailability. The new approach herein,directed towards encapsulation in magnetic porous silica, af-fords a targeted and sustained release mechanism of curcu-min under physiological pH with high magnetic mobility(magnetic nanoparticle loading as high as 37 % wt). The newsynthetic methodology for preparing composite functionalmaterials is likely to be applicable to other biologicallyactive materials, beyond that of the highly topical curcuminmolecule.

Experimental Section

Materials : Ferric chloride (FeCl3·6H2O), ferrous chloride (FeCl2·4H2O)(Fluka) tetraethylorthosilicate (TEOS, 98 %, Aldrich), cetyltrimethylam-monium bromide (Aldrich,), formamide (99.5 %, Ajax), hydrochloricacid (32 %, Ajax), and curcumin powder (Sigma) were used as received.

Synthesis of Fe3O4 nanoparticles : Fe3O4 nanoparticles were synthesisedby using spinning-disc processing with Fe2+ /3 + in one jet feed and aque-ous NH4OH in another, as detailed in the literature.[23a] The as-synthes-ised Fe3O4 nanoparticles were 8–10 nm in size.

Encapsulation of Fe3O4 nanoparticles and curcumin in mesoporous silicacapsules : A mixture of H2O/CTABr/HCl/formamide was stirred in ahigh-density polypropylene bottle (HDPP) at room temperature for2 days, after which curcumin (10 mg) and Fe3O4 (100 mg) nanoparticleswere added and the solution was stirred for a day. TEOS was then addedand the solution was stirred for about 5 min. The solution was then keptunder quiescent conditions for 3 days. The molar ratio of H2O/HCl/for-mamide/CTABr/TEOS was 100:7.8:10.2:0.11:0.13. The samples were cen-trifuged and washed with ethanol at least three times to remove anyexcess of curcumin that adhered at the surface of the silica capsule. Thewashed samples were then passed through a high field gradient magneticseparation column to separate the magnetic from the non-magneticsample.

Release experiment : The silica capsules loaded with curcumin and Fe3O4

nanoparticles were collected by a magnet, the solution was decanted, andthe samples were dried. A known amount of silica (100 mg) was dis-persed in phosphate buffer solution (pH 7.6). At predetermined intervalsof time, the sample was collected by using a magnet and a 1 mL aliquotwere taken for measurement. The released curcumin was redissolved inethanol and the absorbance was measured with a UV/Vis spectrometer.

Characterisation methods : Scanning electron microscopy (SEM) imageswere obtained on a Zeiss 1555 VP-FESEM instrument. Transmissionelectron microscopy (TEM) images were recorded using JEOL 3000F mi-croscope and the elemental maps was acquired on a JEOL 2100 TEM.The fluorescent image was recorded using a Leica TCS SP2 AOBS multi-photon confocal microscope. The FTIR spectrum was recorded in thetransmission mode on a Perkin–Elmer Spectrum One spectrometer. Thedried sample of was grounded with KBr and the mixture was compressedinto a pellet. The spectrum was taken from 4000 to 400 cm1. UV/Visspectroscopy was performed on a Perkin–Elmer Lambda 25 UV/vis spec-trometer by using matched quartz cuvettes with a path length of 1 cm.The magnetic properties were measured using a Quantum Design 7 TMPMS superconducting quantum interference device (SQUID) magneto-meter at 300 K. The iron content of each sample was determined usingatomic absorption spectroscopy (AAS) and the measured magnetisationdata were normalised with respect to the iron content of the samples.

SAXS patterns were measured with the Bruker NanoSTAR SAXS in-strument at Curtin University. The powder samples were mounted be-tween thin polymer films during measurements. Data was recorded at asample–detector distance of 65.0 cm, using a wavelength, l of 0.15418 nm(CuKa) resulting in a q-range of 0.11–3.27 nm1 (q=4psinq/l, in which 2q

is the scattering angle). The measured intensities were corrected forsample transmissions and background, and were converted to an absolutescale for modelling purposes by using a known standard by means of themethod of Spalla et al.[24]

Acknowledgements

The authors are grateful for the financial support for this work by theAustralian Research Council and The University of Western Australia.S.F.C acknowledges postgraduate scholarship from The University of Ma-laysia Sarawak. The microscopy analysis was carried out using facilitiesat the Centre for Microscopy, Characterisation and Analysis at The Uni-versity of Western Australia, which are supported by University, State,and Federal Government funding. The authors thank John Murphy forhelp with microtoming and confocal microscope work and Peter Duncanfor help with SEM work.

Keywords: curcumin · drug delivery · silica capsule ·superparamagnetics · sustained release

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Received: December 30, 2008Published online: April 25, 2009

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www.chemeurj.org

Suk Fun Chin, Mohamed Makha, Colin L Raston School of Biomedical, Biomolecular and Chemical Sciences

The University of Western Australia, Crawley, Western Australia 6009 Email: [email protected]

Telephone: (618) 64881572, Fax: (618) 64881005

Abstract—Magnetite nanoparticles were synthesized by coprecipitation of Fe2+ and Fe3+ with NH4OH using Spinning Disc Processing (SDP). Chitosan was then coated on the surface of magnetite nanoparticles using SDP. FTIR study and zeta potential measurement confirmed the absorption of chitosan unto the surface of magnetite nanoparticles. Transmission electron microscope (TEM) image showed that the particle sizes are in the range 10 – 200 nm.

Keywords-magnetite, nanoparticles; chitosan; coating

I. INTRODUCTION Magnetic nanoparticles have been extensively studied

because of their potential applications as contrast agents in magnetic resonance imaging (MRI) of tumors, cell and DNA separation, magnetically guided drug delivery, tumor hyperthermia etc. [1-3]. Among the magnetic oxides, magnetite nanoparticles are most suitable due to their low toxicity and good magnetic properties. Magnetite is a ferromagnetic iron oxide, Fe3O4 with an inverse spinel crystalline structure in which part of the iron atoms are octahedrally coordinated to oxygen and the rest are tetrahedrally coordinated to oxygen [4]. However, magnetite tends to aggregate due to strong magnetic dipole-dipole attractions between particles combined with inherently large surface energy.

In order to successfully prepare stable magnetite dispersions, any attractive forces between the nanoparticles must be overcome. Stabilizers or surfactants have been used to prevent sedimentation of these nanoparticles. The choice of stabilizer relates to its ability to interact with magnetite particles via functional groups and form a tightly bonded monomolecular layer around the particles. Another possible way of stabilizing iron oxide nanoparticles in aqueous solution is to encapsulate them with polymeric materials. The polymer coatings serve as a steric barrier which reduces magnetic attractions between the particles. Besides these, coatings on magnetite nanoparticles can also improve chemical stability by protecting the particles surface from oxidation.

In this study, encapsulation of magnetite nanoparticles using chitosan coupled with Spinning Disk Processing (SDP) technique is reported. As a natural biopolymer, chitosan is a hydrophilic, biocompatible, biodegradable and non-toxic polymer, making it attractive for biomedical applications [5-6]. The presence of a shell of this biopolymer around magnetite

nanoparticles will not only improve the biocompatibility of magnetite nanoparticles but also provides amino and hydroxyl groups for interplay with biomolecules. Furthermore, chitosan can be produced by deacetylation of chitin. Chitin is the main component of exoskeleton of crustaceans and is the second most abundant natural polymer after cellulose, and therefore is a cheap, renewable biopolymer [7].

II. MATERIALS AND METHOD A) Synthesis of magnetite nanoparticles

Magnetite nanoparticles were prepared by coprecipitation

of ferric and ferrous chlorides (1:2) with aqueous ammonia solution. Both solutions were purged with argon gas for at least 15 minutes and then continuously fed into the SDP at feeding rate of 0.5 ml/s. The disk spinning rate was 2000 rpm. The synthesis was carried out under an argon atmosphere. The conical flask contained the resulting magnetite was placed on a permanent magnet, the supernatant solution was discarded. Deoxygenated ultra pure deionized water was added to wash the magnetite nanoparticles. This process was repeated for several times until the pH of the suspension was nearly neutral. The magnetite nanoparticles were then re-dispersed in deoxygenated deionized water.

B) Coating of magnetite nanoparticles with chitosan

Chitosan solution was prepared by dissolved various

amount of chitosan in 1% of acetic acid. Chitosan coated magnetite nanoparticles were prepared by mixing the preformed magnetite nanoparticles with chitosan solution using SDP at feeding rate of 0.5 ml/s and at 1500 rpm disk spinning rate. The chitosan coated magnetite nanoparticles were then washed several times with deionized water to remove the excess chitosan in the suspension.

C) Characterization

The FTIR spectra of the samples were recorded on a Perkin

Elmer FTIR Spectrometer at 4 cm-1 resolution. Samples were ground-blended with KBr and the compressed to form pellet. All the spectra were recorded at the range of 400 – 4000 cm-1.

Zeta potentials of uncoated magnetite and chitosan coated magnetite as a function of pH were determined using a Zetasizer Nano ZS series (Malvern Instruments Ltd., UK).

Encapsulation of Magnetic Nanoparticles with Biopolymer for Biomedical Application

ICONN 20061-4244-0453-3/06/$20.00 2006 IEEE 374

A Philips 410 transmission electron microscope (TEM) was used to investigate the particle size and morphology of chitosan coated magnetite nanoparticles. The applied operating voltage was 80 kV.

III. RESULT AND DISCUSSION The FTIR spectra of chitosan, chitosan coated magnetite

and uncoated magnetite are shown in Figure 1. Both the chitosan and chitosan coated magnetite exhibited the characteristics peaks of chitosan. The absorption peak observed at 3436 cm-1 is due to the hydroxyl (OH-) group and the peak at 1637 cm-1 is due to the amine group (-NH2). The absorption peak at around 1072 cm-1 relates to C-C and C-O stretching modes of polysaccharides backbone, and the bending vibration of –CH2 group is at 1400 cm-1 [8] . The characteristic peaks of magnetite at 582 cm-1 and 565 cm-1 were also observed in the spectra of magnetite and chitosan coated magnetite. These spectra confirmed that the chitosan has coated onto the surface of magnetite. However, the spectra of chitosan coated magnetite nanoparticles showed very little difference from the pure chitosan indicating weak interactions between chitosan and magnetite nanoparticles.

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Figure 1. FTIR spectra of chitosan, chitosan coated magnetite and magnetite.

The zeta potential of magnetite nanoparticles with and without chitosan coating as a function of pH value is highlighted in Figure 2. The surface charge potential of iron oxides in water at various pH values could be explained by surface hydroxyl groups of iron oxides (FeOH) [9]. In a basic environment, the surface of iron oxide shows negative charge potential due to the dissociation of FeOH to form FeO-. In contrast, in an acidic environment, the positive surface charge is due to the formation of FeOH2

+. The measured isoelectric point (IEP) of uncoated magnetite is at ~pH 6.3. This value is very close to the reported value of magnetite which is at about pH 6 [9]. After coated with chitosan, the zeta potential of the magnetite are more positive below the IEP and less negative above the IEP. The coating of chitosan not only reduced the number of negative surface sites but also induced positive surface sites due to the protonization of –NH2 groups of

chitosan. As observed in Figure 2, the IEP of chitosan coated magnetite is shifted from pH 6.3 to 8.6. As the reported IEP of chitosan is at pH 8.7 [10], this shift of IEP can be explained by the formation of chitosan on the surface of magnetite nanoparticles.

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TEM imaging of chitosan coated magnetite nanoparticles is shown in Figure 3. The image indicated that the particle sizes range from 10 – 200 nm. The broad size distribution may be due to the formation of aggregation of the magnetite. It can be seen from the image that the magnetite nanoparticles are surrounded by the chitosan.

Figure 3. TEM image of chitosan coated magnetite nanoparticles

CONCLUSION In conclusion, we have successfully coated chitosan onto

the surface of magnetite by SDP. Future work will focus on optimizing the synthesis conditions for preparation of stable chitosan derivative coated magnetite nanoparticles with desirable particles size, surface and magnetic properties for biomedical applications.

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ACKNOWLEDGMENT The authors would like to gratefully acknowledge University of Malaysia Sarawak and The University of Western Australia for the support of this work.

REFERENCES [1] D.C.F. Chan, D.B. Kirpotin, P.A. Bunn. “ Synthesis and evaluation of

colloidal magnetic iron oxides for the site-specific radiofrequency-induced hyperthermia of cancer. J. Magn. Magn. Mater. Vol.122, pp 374 – 378, 1993,

[2] H. Chen, R. Langer. “Magnetically-responsive polymerized limposomes as potential oral delivery vehichles” Pharm. Res. Vol. 14, pp 537 – 540, 1997.

[3] D.K. Kim, , Y.Zhang, J. Kehr, Klason, T, B. Bjelke, M. Muhammed. “ Characterization and MRI study of surfacted-coated superparamagnetic nanoparticles into the rat brain”. J. Magn. Magn. Mater. Vol 225, pp 256 -261, 2001.

[4] R.M. Cornell, U. Schwertmann. The Iron Oxides; VCH: New York, 1996.

[5] Z.Ma, H.H, Yeoh, L.Y. Lim. “Formulation pH modulates the interaction of insulin with chitosan nanoparticles” J. Pharm Sc. Vol.91, No.6, pp1396 -1404, 2002.

[6] T. Chandy, CP, Sharma. “Chitosan beads and granules for oral sustained delivery of nifedipine: In vitro studies. Biomaterials. Vol.13, pp 949-952, 1992.

[7] Q. Li, E.T. Dunn, E.W. Grandmaison, M.F. Goosen. “Application and Properties of chitosan. In Applications of chitin and Chitosan; M.F. Goosen, Ed., Technomic Publishing: Lancaster, P.A, pp 3 -29, 1997.

[8] J.Brugnetto, J. Lizardi, F.M. Goycoolea, W. Argulles-Monal, J. Desbrieres, M. Rinaudo. “An infrared investigation in relation with chitin and chitosan characterization. Polymer. Vol.42, pp 3569 -3580, 2001.

[9] Z.X. Sun, F.W.Su, W. Forsling, P.O. Samskorg. “ Surface characteristics of magnetite in aqueous suspension”. Vol. 197, pp 151 – 157, 1998.

[10] C.Huang, Y. Chen. “Coagulation of colloidal particles in water by chitosan”. J. Chem. Tech. Biotechnology. Vol 66, pp227 – 232, 1996.

376

Process Intensification Strategies for the Synthesis of Superparamagnetic Nanoparticles and Fabrication of Nano-Hybrid

Suk Fun Chin1, K. Swaminathan Iyer1, Colin L. Raston1 and Martin Saunders2 1 Center for Strategic Nano-Fabrication, School of Biomedical, Biomolecular and Chemical

Sciences,The University of Western Australia, Crawley, 6009 Australia 2Center for Microscopy, Characterization and Analysis, The University of Western Australia,

Crawley, W.A. 6009 Australia

ABSTRACT

Continuous flow spinning disc processing (SDP), which has extremely rapid mixing under plug flow conditions, effective heat and mass transfer, allowing high throughput with low wastage solvent efficiency, is effective in gaining access to superparamagnetic Fe3O4 nanoparticles at room temperature. These are formed by passing ammonia gas over a thin aqueous film of Fe2+/3+ which is introduced through a jet feed close to the centre of a rapidly rotating disc (500 to 2500 rpm), the particle size being controlled with a narrow size distribution over the range 5 nm to 10 nm, and the material having very high saturation magnetizations, in the range 68–78 emu g-1. SDP also shown to be effective for fabrication of superparamagnetic carbon nanotubes composite. Ultra fine (2-3 nm) magnetite (Fe3O4) nanoparticles were uniformly deposited on single-walled carbon nanotubes (SWCNTs) in situ by modified chemical precipitation method using SDP in aqueous media at room temperature under continuous flow condition

Keywords: Spinning disc processor , superparamagnetic, continuous flow technology, magnetic nanoparticles, carbon nanutubes

1 INTRODUCTION Traditional fluid based synthesis techniques for the

production of nanoparticles have inherent limitations such as poor particle size distribution and reproducibility, and difficulties in scalability for commercial production. Process intensification, by means of spinning disc processing (SDP), potentially offers an avenue for the production of monodisperse nanoparticles with tuneable and controllable properties. SDP, Figure 1, is a rapid flash nano-fabrication technique with all reagents being treated in the same way, and is in contrast to traditional batch technology where conditions can vary across the dimensions of the vessel.[1] The reagents are directed towards the centre of the disc, which is rotated rapidly (300 and 3000 rpm) resulting in the generation of a very thin fluid film (1 to 200 μm). The thinness of the fluid layer and

the large contact area between it and the disc surface facilitates very effective heat and mass transfer. The drag forces between the moving fluid layer and the disc surface enable very efficient and rapid mixing. The greatest strength of SDP synthesis is the broad range of control possible over all the operating parameters involved in nanoparticle formation, enabling the simultaneous and individual optimization of many interdependent operating mechanisms, with the ultimate goal of achieving very narrow particle size distributions.[2] Another feature of SDP is that it is continuous flow, readily allowing scale-up of the ensuing product formation.

Figure 1. (a) Schematic representation of a SDP, (b)

Hydrodynamics of the fluid flow over a spinning surface.

The most common cost effective and convenient way to synthesize Fe3O4 nanoparticles is by co-precipitating ferrous and ferric salt solutions with a base, such as

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aqueous NaOH or NH4OH. [3] However, the size distribution of the Fe3O4 nanoparticles produced using this method is normally very broad. Consequently, the downstream purification and isolation process is more expensive and is time and energy intensive. Furthermore, scale-up of this method using conventional reactors can be problematic given the inhomogeneous agitation and areas of localized pH variations, resulting in the precipitation of non-magnetic iron oxides.[4] Herein we demonstrate the successful synthesis of Fe3O4 nanoparticles via co-precipitation using NH3 gas as a base source using spinning disc processing (SDP) under scalable and continuous flow conditions. To our knowledge, this is the first use of NH3 gas as a precipitating agent to make Fe3O4 nanoparticles in a thin fluid film. The technology offers a realistic route towards large scale synthesis of Fe3O4 nanoparticles with precise control within the 10 nm size range.

The efficient SDP capability of fabricating magnetic nanoparticles decorated single wall carbon nano-tubes (SWCNTs) were also demonstrated. A novel yet simple method to coat superparamagnetic Fe3O4 nano-particles of narrow size distribution on SWCNTs in situ by modified chemical precipitation method using SDP in aqueous media at room temperature under continuous flow conditions was reported in this paper.

2 EXPERIMENTAL

2.1 Synthesis of Fe3O4 nanoparticles

In a typical synthesis, aqueous solutions of Fe2+/3+

precursors were prepared by dissolving FeCl2.4H2O (10 mM) and (20 mM) FeCl3.6H2O (1:2 molar ratios) in deoxygenated ultrapure Mili-Q water. The SDP was a Protensive 100 series with integrated feed pumps to direct the reactants onto the rotating disc. The above solutions were delivered onto the disc surface using one feed jet at 1.0 mls-1, using continuous flow gear pumps (MicroPumps). Grooved and smooth stainless steel discs with 100 mm diameter were used which were manufactured from 316 stainless steel with the grooved disc having 80 concentric engineered grooves equally spaced at 0.6 mm depth.

The reactor chamber was purged with argon gas before

the reaction to remove oxygen. Ammonia gas was then fed into the sealed reactor chamber at a constant flow rate. Black suspensions of Fe3O4 nanoparticles were collected from beneath the disc through an exit port.

The samples collected were immobilized with a permanent magnet and supernatant solutions were decanted. Samples were re-dispersed in deoxygenated ultrapure Mili-Q water.

2.2 Fabrication of Fe3O4 decorated Carbon Nanotubes (CNT)

For purification and functionalization of SWCNTs, 10 mg of SWCNTs were dispersed in 5 ml of 1: 1 mixture of 70% HNO3 and 98% H2SO4 aqueous solutions in the reaction chamber. The reaction was carried out in a CEM Focused Microwave Synthesis System, Discover Model. The microwave power was set at 300 W, and the pressure was 12 bar, and the temperature was set at 130 oC for 30 minutes. After the reaction, the SWCNTs were filtered, washed and re-dispersed in 100 mL of ultra pure Milli-Q water and sonicated for 15 minutes. The functionalized SWCNTs were dispersed in water and the container purged with N2 gas to remove oxygen then 10 mM of FeCl2.4H2O and 20 mM of FeCl3.6H2O (1:2 molar ratios) was added and the mixture stirred for 1 hour. After that, the solution was filtered to remove excess Fe2+/3+ and the resulting carbon nano-tube and Fe2+/3+ complex was re-dispersed in deoxygenated Mili-Q water.

Solutions/suspensions of CNTs and Fe2+/3+ were fed

from one feed and the deoxygenated NH4OH aqueous solution was fed from the other under an atmosphere of high purity (99.9%, BOC Gasses) argon gas, within the sealed reactor chamber. The disc surface was manufactured from 316 stainless steel. The 10 cm grooved disc was used for the current study, with 80 concentric engineered grooves equally spaced approximately 0.6 mm in height. Samples were collected from beneath the disc through an exit port.

3 RESULTS AND DISCUSSION

(a) (b)(a) (b)

Figure 2. TEM images of Fe3O4 nanoparticles (10 mM Fe2+) synthesized at 25 oC on grooved disc with disc rotating speed: (a) 2500 rpm and (b) 500 rpm

The operating parameters of the SDP were effective in controlling the size, size distribution and shape of the Fe3O4 nanoparticles in the presence of NH3 gas feed as the base

783NSTI-Nanotech 2008, www.nsti.org, ISBN 978-1-4200-8503-7 Vol. 1

rather than NH4OH aqueous solution. In a typical synthesis, the samples were synthesized using 10 mM of Fe2+ and 20 mM of Fe3+ aqueous solution and were fed at 1mls-1 onto the rotating grooved disc. A high rotation speed of 2500 rpm resulted in ultrafine (3-5 nm) particles with asymmetrical shape. A lower speed of 500 rpm, resulted in spherical nanoparticles of around 10 nm in size, Figure 2.

The fluid layer of the SDP has a plug flow

characteristic, where the particles produced are constantly being removed from the nucleation and growth zones by radial propagation across the disc. At high rotation speeds the non-linear wave regime is dominant in the fluid film, which in turn ensures greater gas adsorption into the thin film. Consequently the nucleation process becomes dominant, thereby resulting in a number of ultra-small magnetic nanoparticles. At lower spin speeds the wave regime is no longer predominant, and the velocity of the traversing waves are highly reduced, and so is the absorption of the corresponding NH3 gas into the flowing film. The number of nucleation sites is thereby decreased and the growth process becomes dominant at lower speeds resulting in the formation of larger particles.

(b)(a) (b)(a)

Figure 3. TEM images of Fe3O4 nanoparticles (10 mM Fe2+) synthesized on smooth disc with disc rotating speed of 500 rpm, at (a) 25 oC and (b) 120 oC.

At constant speed, samples synthesized using the

grooved disc had a narrower particle size distribution compared to the smooth disc. The particle size for the samples synthesized on the grooved disc ranged from 3-5 nm (Figure 2a), whereas the particle size distribution of a sample prepared using the smooth disc ranged from 3 -12 nm (Figure 3a). When using the grooved disc, the shear forces and viscous drag between the moving fluid layer and the periodic grooved surface give rise to more efficient turbulent mixing within the fluid layer and this in turn results in homogeneous reaction conditions in the thin film. The grooves on the disc also result in the formation of waves on the flowing film, thereby amplifying the amount of the NH3 gas adsorbed. However, on a smooth disc the micro-mixing is not as efficient resulting in a broader size

distribution. Increasing the temperature of the disc offers scope to overcome the aforementioned size distribution issues associated with the smooth disc. At higher temperature the absorption efficiency of the gas is reduced in the flowing fluids, however the growth conditions become highly amplified [5]. As a result, the size distribution of the samples prepared on the smooth disc became very narrow with particle sizes in the range 8-10 nm for the reactions at 120 oC, Figure 3(b).

Figure 4. TEMs of Fe3O4–SWCNT composite at (a and b) low resolution; (c) high resolution and (d) associated SAED pattern for material synthesized using SDP

For the fabrication of Fe3O4-SWCNT composite, ultrafine (2-3nm) of Fe3O4 nano-particles with very narrow size distribution were observed to be uniformly coated onto the CNTs surface, Fig 4 (a), (b) and (c). As can be seen from the TEM images, the distribution of Fe3O4 nano-particles on the SMWNTs surface is very uniform and no local aggregation is observed with a very high coverage density. The ability to effectively decorate the SWCNTs with Fe3O4 nano-particles relates to the functionalisation of the nano-tubes with COO- moieties that can bind directly to Fe2+/3+

ions. [6] The use of this simple yet strategic approach reduces the down-stream processing dramatically. This is associated with purification/separation steps to remove excess nanoparticles that grow independently in solution and are not associated with the CNTs.

4 CONCLUSION

In conclusion, the particle size of Fe3O4 nanoparticles

784 NSTI-Nanotech 2008, www.nsti.org, ISBN 978-1-4200-8503-7 Vol. 1

can be controlled by judicious choice of the operating parameters of SDP technology. The capabilities of SDP in controlling particle size and stability have been clearly demonstrated with the ability to produce ultra-small superparamagnetic magnetite nanoparticles with high saturation magnetizations, and with narrow size distributions. We have also developed a novel, simple, rapid, cost effective and scalable method to decorate CNTs with 2-3 nm Fe3O4 nano-particles using SDP. The resulting CNTs show high coverage density. The apporach also avoids the need for subsequent purification and separation of the composite material from excess Fe3O4 nano-particles formed in solution, unlike in the case of using batch reactions. Futhermore the ability of SDP to fabricate superparamagnetic SWCNT composites under continuous flow in scalable quantities is significant in any down stream applications.

REFERENCES

1. P. Oxley, C. Brechtelsbauer, F. Ricard, N. Lewis

and C. Ramshaw, Ind. Eng. Chem. Res. 39, 2175, 2000.

2. K. Swaminathan Iyer, C. L. Raston and M. Saunders, Lab Chip, 7, 1800, 2007.

3. T. Fried, G. Shemer, G. Markovich, Adv. Mater. 13, 1158, 2001

4. L. T. Vatta, R. D. Sanderson, K. R. Koch, J. Magn. Magn. Mater. 311, 114, 2007

5. J. H. Wu, S. P. Ko, H. L. Liu, S. S. Kim, J. S. Ju, Y. K. Kim, Mater. Lett. 61, 3124, 2007

6. B. He, M. Wang, W. Sun, Z. Shen. Mater. Chem. Phys. 95, 289, 2006

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Chapter 4: Conclusions and Future Work

74

4 Conclusio

Although substantial progress has been etic

anoparticles with a wide range of synthetic procedures, scalability still remains one of

e ma

ns and Future Work

achieved in the synthesis of magn

n

th jor stumbling blocks hindering their commercialisation. Another factor that must

be addressed in any production process is the environmental impact, which includes

avoiding the generation or at the very least minimising the amount of waste. Overall,

such sustainability metrics need to be incorporated into the production at its inception.

These issues have been addressed in using of spinning disc processing (SDP) to

synthesise superparamagnetic Fe3O4 nanoparticles under continuous flow condition

involving an aqueous based chemical precipitation method.[179] The synthesis using SDP

is under continuous flow and can be easily scaled up for commercial production. The

quantity of sample generated is determined by the amount of time the processor is

running coupled with concentration of Fe2+/3+ and flow rates. Particle sizes can be

controlled in the range of 5 nm to 10 nm with narrow size distribution by judicious

choice of the operating parameters. These include reaction temperature, flow rates, disc

texture, disc rotating speeds and precursor concentrations, and their influence on the

size of the Fe3O4 nanoparticles have been established. Indeed the synthesis of Fe3O4

nanoparticles by the co-precipitation method using SDP is able to overcome the

limitations associated with the scaling up of conventional reactors such as

inhomogeneous agitation and areas of localised pH variations, resulting in the

precipitation of non-magnetic iron oxides and broad particle size distributions. In

addition, the high throughput of the system, low changeover times and low wastage

make this a very solvent efficient method of synthesis. Surfactants can be readily

coated onto the nanoparticles during the process intensification such that stabilised

nanoparticles can be generated in a single pass under continuous flow. This synthetic

approach is fast, cost effective, reproducible and environmental friendly with the


75

ability of SDP for preparing nanostructured materials has been extended

The extremely short reaction times of SDP (0.5s), typically limits the size of the

e3O4

resulting Fe3O4 nanoparticles are highly crystalline with good magnetisation values,

ranging from 68-78 emug-1.

The cap

to fabricate composite superparamagnetic carbon nanotubes.[180] An efficient method is

developed to decorate single wall carbon nanotubes (SWCNT) with spatially well-

separated ultrafine (2-3 nm) Fe3O4 nanoparticles. This involves pre-funtionalising

SWCNTs with COO- moieties that can bind directly to Fe2+/3+ ions. A more efficient

way for carboxylation has been developed by using microwave radiation which has

dramatically reduced the carboxylation reaction time down to 30 minutes in comparison

to the conventional reflux method which normally takes 10–15 hours, thereby

minimising any damage to the CNTs and reducing energy consumption. Moreover,

there is an additional benefit of the method in avoiding subsequent purification and

separation of composite material from excess Fe3O4 nanoparticles formed in solution. In

future, a continuous flow reactor equipped with microwave radiation capability will be

tried for the carboxylation reaction. The method developed in the study can also be

applied to decorate multiwall carbon nanotubes (MWCNT) with Fe3O4 nanoparticles.

The use of SDP to fabricate superparamagnetic SWCNT composites under continuous

flow in scalable quantities has significant implications towards the development of any

downstream applications.

F nanoparticles with diameters less than 10 nm. In future, a Rotating Tube

Processor (RTP), which is another form of process intensification, (Figure 4.1) can be

explored for synthesising Fe3O4 nanoparticles with larger diameters. The parameters of

the RTP are like SDP, but more importantly the residence time can be controlled. While

the centrifugal forces on the disc move the products off from the disc quickly, the

rotation of the tube swirls the reactants inside the long axis of the tube, thereby

affording longer residence times for the reactants. Furthermore, the ability to selectively

introduce reagents along the long axis of the reactor would enable better control in the


76

nucleation and growth process of the nanoparticles, as well as potentially coating

nanoparticles in situ which would be an important advancement in single step

processing to generate composite nano-materials.

Figure 4.1 Schematic diagram of RTP. A, B and C are jet feeds for controlling the

The feed pumps of the SDP used in the research were magnetic gears. As a

The development of a facile and simple method to modify the as-synthesised

processes using the RTP.[181]

result the surface modification of the pre-formed Fe3O4 nanoparticles using SDP was

not possible. Pumping solution of the preformed Fe3O4 nanoparticles through the SDP

results in clogging of the pumps. In the future, peristaltic pumps which are non-

magnetic are to be installed on the SDP and RTP to allow pumping of the preformed

Fe3O4 nanoparticles through the SDP for coating of Fe3O4 nanoparticles. The SDP and

RTP techniques offer potential opportunities to control the uniformity of the coating and

agglomeration of the magnetic nanoparticles. This is of paramount importance for

biomedical applications of the magnetic nanoparticles.

nanoparticles could render colloidal stability and new functionalities, opening up new

opportunities into novel and niche applications. Even though numerous techniques have

been developed to fabricate core-shell magnetic nanoparticles, difficulties associated in

controlling of the reaction conditions, especially the nucleation and growth of the shell

layer remains a major challenge. Without adequate control of the reaction conditions,


77

The surface of Fe3O4 nanoparticles have been modified with sulfonato-

lixar

aggregation of core particles, formations of separate particles of shell material or

incomplete coverage often occur. Various novel and improved aqueous based synthetic

protocols have been undertaken to coat Fe3O4 nanoparticles with appropriate shells to

stabilise and functionalise the nanoparticles for potential biomedical applications. The

properties of the resulting composite of Fe3O4 core-shell nanoparticles have been

elucidated using various characterization techniques involving Transmission Electron

Microscope (TEM), Scanning Electron Microscope (SEM), X-ray Diffractor (XRD),

Ultraviolet(UV) spectroscopy, and a Quantum Design 7 T MPMS superconducting

quantum interference device (SQUID) magnetometer.

ca enes macrocyles to impart colloidal stability and functionalities.[182] Stable

ferrofluids of sulfonato-calixarenes stabilise Fe3O4 nanoparticles formed by in situ co-

precipitation of Fe3O4 in the presence of the p-sulfonato-calixarenes and sulfonated p-

benzylcalixarenes. Fe3O4 nanoparticles prepared in the presence of p-sulfonato- calix[4

and 5]arenes did not form stable suspensions, in contrast to stable dispersed

nanoparticles using p-sulfonato-calix[6 and 8]arenes. The mechanism for anchoring the

calixarenes to the surface of the Fe3O4 nanoparticles and enhancing their stability has

been established. FTIR, zeta potential measurement and elemental mapping on energy

filtered TEM have been used to confirm the presence of calixarene on the surface of the

Fe3O4 nanoparticles. The p-sulfonato-calix[6 and 8]arenes and sulfonated p-

benzylcalix[4,5,6 and 8]arene stabilised Fe3O4 suspensions are highly stable at

physiological pH (pH=7.6) and exhibited superparamagnetic properties. As sulfonato-

calixarenes are well known to form host-guest inclusion complexes with a variety of

small hydrophobic molecules and drugs, the presence of p-sulfonato-calixarenes and

sulfonated p-benzylcalixarenes on the surface of Fe3O4 nanoparticles makes them

attractive candidates for biomedical applications. In future, the long term stability of

these nanoparticles under the human body’s physiological conditions, including pH and

electrolyte concentrations should be systematically and carefully evaluated before

proceeding to any in vitro applications. The potential applications of these sulfonato-


78

A novel and convenient aqueous based method has been developed to coat Fe3O4

example: Fe3O4@Ag@Au

calixarene Fe3O4 nanoparticles as drug delivery carriers should be evaluated by

incorporating a model drug in their cavities and studying their release profile under

physiological condition warrant thorough investigations.

nanoparticles with a thin and uniform layer of Au and Ag shells at room temperature. The

mild synthesis conditions employed water as the solvent, readily available dopamine as

the surfactant, and glucose as a mild reducing agent. The fabrication step had minimal

detrimental effects on the magnetic properties of the Fe3O4 nanoparticles. This has been

evidenced by the superparamagnetic properties and good magnetisation values of the

resulting Fe3O4@Au and Fe3O4@Ag core-shell nanoparticles. The utility of dopamine as

a surfactant to simultaneously stabilise and functionalise Fe3O4 nanoparticles in aqueous

solution with amine (-NH2) group circumventes the conventional tedious and time

consuming functionalisation process which involves refluxing solution of Fe3O4

nanoparticles with (3-aminopropyl) triethoxysilane (APTES) for 12-18 hours.

Optimisation of the concentrations of Au3+ and Ag+ precursors are very critical to ensure

that most of the Fe3O4 nanoparticles are fully coated, and minimise the formation of free

Au or Ag nanoparticles. These Au and Ag shells also serve as platforms for potential

derivatization with different multifunctional organic molecules, or specific targeting

ligands. This coupled with their magnetic and plasmonic properties should broaden and

strengthen the potential of these nanoparticles in biomedicine applications such as MRI,

hyperthermia and drug delivery. Future research includes investigating the fabrication

of multifunctional hybrid system incorporating bioconjugating specific antibodies,

targeting ligands or therapeutic agents onto the surface of Fe3O4@Au or Fe3O4@Ag

nanoparticles in order to achieve a more precise localization of targeted sites. Future

work also focuses on optimizing the properties of the multifunctional hybrid Fe3O4@Au

or Fe3O4@Ag nanoparticles for their performance as contrast agent in MRI,

hyperthermia treatment and as drug delivery carriers. Furthermore the synthestic

procedure can be applied to access “onion shell” structures with multiple coatings, for


79

weak magnetic moment resulting from the low loading of

agnet

The effective applications of magnetic silica capsules in biomedical applications

has been hampered by the

m ic nanoparticles in silica particles. It is noteworthy that the method reported in the

literature for loading of drugs into the channel of mesoporous silica involves mainly

post-synthesis of the nanoparticles with the mesoporous silica carrier was synthesised

first followed by the incorporation of the drug molecules. A novel method has been

developed to encapsulate curcumin in the mesoporous silica by surfactant template self-

assembly method. High loading of Fe3O4 nanoparticles (37 wt%) and curcumin (30

wt%) has been simultaneously incorporated into silica particles through this novel and

straightforward synthetic method at room temperature. Encapsulation of curcumin within

the silica matrix was confirmed by confocal laser scanning microscopy and UV visible

spectroscopy. The ability of these magnetic silica particles to release curcumin in a

controlled way at physiological pHs and using phosphate buffer, demonstrates that the

magnetic silica capsules hold great promise as controlled and target release carriers. The

encapsulation enhances the bioavailability, and imparts exceptional photostability of the

curcumin, and preventing them from being destroyed by enzymes and acid in the body,

thereby enhancing the likelihood of them being absorbed in the digestive tract. The

magnetisation values of the Fe3O4 nanoparticles were well preserved after the

encapsulation into the silica capsules. Also, the synthesis approach avoided the use of

expensive block copolymers as surfactant, which can substantially reduce the

production cost and at the same time producing a superior magnetic material. It is

believed that this method will have significant application for the controlled release of

therapeutic molecules using mesoporous silica capsules due to their stability,

controllable pore diameter, and excellent biocompatibility. Future work will focus on

optimising the quantity of curcumin/biomolecules loading by controlling the pore size

and internal surface chemistry to achieve a desirable slow release period. External

stimuli responsive features (e.g. pH, temperature, ionic strength, light) should be

incorporated into the magnetic carriers to avoid uncontrollable release of biomolecules

or therapeutic agents resulting from natural diffusion of molecules upon delivery in the

human body, thereby achieving precise targeting and treatment efficacy. In addition,


80

e aim of this study to develop a better synthetic route that is

ucible and cost effective for generating SPIONs has been achieved using

thorough and long term cytotoxicity studies of these magnetic silica particles need to be

carried out before in vivo applications can be implemented. This encapsulation and

controlled release method for curcumin also can be applied to other nutraceutical and

pharmaceutical drugs.

In conclusion, th

scalable, reprod

SDP involving a chemical precipitation method. The colloidal stability of the SPIONs

under physiological conditions also has been enhanced by modification of the SPIONs

surface with sulfonato-calixarenes. Hybrid nanomaterials of SPIONs namely:

SWCNTs/Fe3O4 and gold coated SPIONs and silver coated SPIONs have been

successfully fabricated using novel aqueous based synthetic methods. A strategic route

has also been developed to encapsulate high loading of SPIONs within mesoporous

silica capsule for drug delivery applications. The body of work represents a significant

contribution to nanoscience leading unto nanotechnology.

Chapter 5: Appendices

93

5 Appendices

1. Online supporting information for S.F.Chin, M.Makha, C.L.Raston,

M.Saunders, Chem. Commun., 2007, 1948-1950. DOI: 10.1039/b618596g.

2. Online supporting information for S.F.Chin, K.S.Iyer, C.L.Raston. Lab Chip,

2008, 8, 439-442. DOI: 10.1039/b716195f.

3. Online supporting information for S.F.Chin, K.S.Iyer, C.L.Raston, M.Saunders,

Adv. Funct. Mater., 2008, 18, 922–927.

http://www.wiley- vch.de/contents/jc_2126/2008/adfm200701101_s.pdf

4. Online supporting information for S.F.Chin, K.S.Iyer, C.L.Raston. Cryst.

Growth Des., 2009, 6, 2685-2689.

http://pubs.acs.org/doi/suppl/10.1021/cg8013199

5. Online supporting information for S.F.Chin, K.S.Iyer, M.Saunders, T.S.Pierre,

C.Buckley, M.Paskevicius and C.L.Raston. Encapsulation and sustained release

of curcumin using superparamagnetic silica reservoirs. Chem. Eur. J., 2009, 15,

5661-5665.

http://www3.interscience.wiley.com/journal/122363186/suppinfo

6. Highlights of Australian Chemistry, 2008, Vol.75, No.6

This journal is (c) The Royal Society of Chemistry 2007

Magnetite ferrofluids stabilized by sulfonato-calixarenes Suk Fun Chin a, Mohamed Makha* a, Colin L. Raston* a and Martin Saunders b Supporting Information 1(a)

4000.0 3600 3200 2800 2400 2000 1800 1600 1400 1200 1000 800 600 450.0-18.2

-10

0

10

20

30

40

50

60

70

80

90

96.6

cm-1

%T

Fe3O4

pb8SO3H_Fe3O4

pb4SO3H_Fe3O4

pb6SO3H_Fe3O4

pb5SO3H_Fe3O4

3411.9

1632.2

1400.4

1164.81121.9

1036.3

581.23429.2

1632.5

1400.91385.0

1162.71121.9

1035.51009.8

568.5

3436.3

1637.0

1400.51384.7

1121.9

578.9

3138.2

1634.8

1400.7

1175.41123.2

1036.91010.5

574.7

3401.2

1627.11400.2

583.2

1(b)

4000.0 3600 3200 2800 2400 2000 1800 1600 1400 1200 1000 800 600 450.0-35.3

-30

-20

-10

0

10

20

30

40

50

60

70

82.7

cm-1

%T

pb8SO3H

pb6SO3H

pb4SO3H

pb5SO3H

3455.0

1656.6

1460.4

1125.01039.2

865.1

686.7

3426.91468.9

1106.9

864.8

620.8

3433.8

1645.9

1400.2

1183.41121.4

1038.31011.3

619.1

3401.1

1460.3

1114.3

1038.41011.3

865.4

698.0

618.7

571.7


1(c)

4000.0 3600 3200 2800 2400 2000 1800 1600 1400 1200 1000 800 600 450.0-36.1

-30

-20

-10

0

10

20

30

40

50

60

70

80

90

100

108.6

cm-1

%T

Fe3O4

Scalix5-Fe3O4

Scalix4_Fe3O4

Scalix6_Fe3O4

Scalix8_Fe3O4

3400.6

1629.71400.51384.6

1169.8

1112.9

1043.7

581.93435.4

3153.2

1633.8

1401.1

1114.11047.4

582.6

3434.5

2917.22848.9

1629.8

1462.91400.6

1384.4

1175.9 1042.8

578.3

3432.4

2916.92848.9

1629.5

1463.01400.9

1384.4

1172.81116.6

1042.6

564.5454.9

3401.2

1627.11400.2

583.2

1(d)

4000.0 3600 3200 2800 2400 2000 1800 1600 1400 1200 1000 800 600 450.0-17.3

-15

-10

-5

0

5

10

15

20

25

30

35

40

45

50

56.5

cm-1

%T

NaSO3calix[6]arene

NaSO3calix[8]arene

3428.6

1645.8

1453.4

1112.1

1045.4

868.3619.0

3402.7

1454.8

1174.1 1041.8

865.7

619.7

Figure 1: FTIR spectra of (a) p-benzyl-calix[4,5,6 and 8]arene sulfonates coated magnetite; (b) p-benzyl-calix[4,5,6 and 8]arene sulfonates (c) p-sulfonato-calix[4,5,6,8]arene coated magnetite (d) p-sulfonato-calix[6,8]arene


Figure 2: Particle size distributions of p-sulfonato-calix[6,8]arene and sulfonated p-benzylcalix[4,5,6 and 8]arene coated magnetite

-80

-60

-40

-20

0

20

40

60

0 2 4 6 8 10 12 14

pH

Zeta

pot

entia

l (m

V)

pb8SO3Hpb6SO3Hpb5SO3Hpb4SO3HNaSO3calix[6]areneNaSO3calix[8]areneuncoated Fe3O4

Figure 3: Zeta potential of magnetite nanoparticles and p-sulfonato-calix[6,8]arene and sulfonated p-benzylcalix[4,5,6 and 8]arene coated magnetite nanoparticles as a function of pH value.

0

10

20

30

0.1 1 10 100 1000 10000

Number (%)

Size (d.nm)

Size Distribution by Number

pb4SO3H_ Fe3O4 pb5SO3H_ Fe3O4

pb6SO3H_ Fe3O4 NaSO3calix6_Fe3O4

pb8SO3H_ Fe3O4 NaSOcalix8_ Fe3O4

Figure 4: Typical Diffraction pattern of calix[n]arenes coated magnetite nanoparticles

1

Fabrication of Carbon Nanotubes Decorated with Ultra Fine Superparamagnetic Nanoparticles under Continuous Flow Conditions Suk Fun Chin,a K. Swaminathan Iyer,a and Colin L. Raston*a Supporting Information Synthesis of Fe3O4 nanoparticles using SDP In a typical synthesis, aqueous solutions of Fe2+/3+ precursors were prepared by dissolving

FeCl2.4H2O (10 mM) and (20 mM) FeCl3.6H2O (1:2 molar ratios) in deoxygenated

ultrapure Mili-Q water. The Fe2+/3+ precursors were reacted with deoxygenated NH4OH

aqueous solution. The SDP was a Protensive 100 series with integrated feed pumps to

direct the reactants onto the rotating disc. Grooved stainless steel disc with 100 mm

diameter was used which were manufactured from 316 stainless. The above solutions

were delivered onto the disc surface using feed jet both at 0.5 ml/s, using continuous

flow gear pumps (MicroPumps), under an atmosphere of high purity (99.9%, BOC

Gasses) argon gas, within the sealed reactor chamber. Samples were collected from

beneath the disc through an exit port. The samples collected were immobilized with a

permanent magnet and supernatant solutions were decanted. Samples were re-dispersed

in deoxygenated ultrapure Mili-Q water. This process was repeated at least three times to

remove chloride salts.

aCentre for Strategic Nano-Fabrication, School of Biomedical, Biomolecular and Chemical Sciences, The University of Western Australia, 35 Stirling Highway, Crawley, WA 6009, Australia. E-mail: [email protected] Fax: (618) 64881005; Tel: (618) 64881572

2

Figure S1: TEM images of Fe3O4 nanoparticles synthesized using SDP

-1.00E+02

-8.00E+01

-6.00E+01

-4.00E+01

-2.00E+01

0.00E+00

2.00E+01

4.00E+01

6.00E+01

8.00E+01

1.00E+02

-80000 -60000 -40000 -20000 0 20000 40000 60000 80000

Field (Oe)

Sp

ecif

ic m

agn

etic

mo

men

t (e

mu

/g)

Figure S2: Magnetization curve of Fe3O4 nanoparticles at 300K

Figure S2, magnetization curve of Fe3O4 nanoparticles synthesized by SDP showed

superparamagnetic behaviour at room temperature.

1

Size selective synthesis of superparamagnetic

nanoparticles in thin fluids under continuous

flow conditions

Suk Fun Chin1, K. Swaminathan Iyer1, Colin L. Raston1*, Martin Saunders2

1Center for Strategic Nano-Fabrication, School of Biomedical, Biomolecular and Chemical Sciences, The University of Western Australia, Crawley, 6009 Australia,

2 Center for Microscopy, Characterisation and Analysis, The University of Western Australia, Crawley, W.A. 6009 Australia

Supporting Information

50 nm

(a) (b)

50 nm50 nm

(a) (b)

Figure S1. TEM images of Fe3O4 (50 mM) synthesized using grooved disc, 2500rpm, at (a) 50 oC and (b) 80 oC

2

4000.0 3600 3200 2800 2400 2000 1800 1600 1400 1200 1000 800 600 450.0

-83.3

-60

-40

-20

0

20

40

60

80

100

120

144.4

cm-1

%T

Alginic acid coated Fe3O4

Alginic acid

33992975

29291740

16341417 1246

10881047

947880

812 603481

3435

24261610

1384

1271 11291088

839

594

Figure S2: FTIR spectra of algninc acid coated Fe3O4 and alginic acid

3

Figure S3: Size distribution of alginic acid coated Fe3O4


4000.0 3600 3200 2800 2400 2000 1800 1600 1400 1200 1000 800 600 450.0-17.3

-10

-5

0

5

10

15

20

25

30

35

40

45

50

55

60

65

7073.4

cm-1

%T

38443718

3344 30432959

2640

25412434

19391753

16171600

1585

1519

1501

1471

1393

13431320

1286

1261

11911176

11471115

1081

1014963

946936

876

814

791

773

751

724

599

555

476

Figure S1. FTIR spectra of dopamine coated Fe3O4 nanoparticles

Figure S2. TEM images of uncontrolled nucleation of (a) Au and (b) Ag unto Fe3O4 nanoparticles


0.480 0.4852 111

0.299 0.2967 220

0.260 0.2532 311

0.210 0.2099 400

0.175 0.1714 422

0.166 0.1615 511

Table 1. The d-spacing values (nm) calculated from the electron diffraction patterns and

the standard atomic spacing for Fe3O4 along with respective hkl indices from the (JCPDS

file No.19-629)

Encapsulation and Sustained Release of Curcumin using

Superparamagnetic Silica Reservoirs

Suk Fun Chin,[a] K. Swaminathan Iyer,*[a] Martin Saunders,[b] Tim St. Pierre,[c] Craig

Buckley,[d] Mark Paskevicius[d] and Colin L. Raston*[a]

[a] S.F.Chin, Dr.K.S.Iyer and Prof. C.L.Raston

Center for Strategic Nano-Fabrication, School of Biomedical, Biomolecular and Chemical Sciences, The

University of Western Australia, Crawley, 6009 Australia

Fax: +61 86488 8683

E-mail: [email protected]; [email protected]

[b] Prof. T.S.Pierre

Center for Microscopy, Characterization and Analysis, The University of Western Australia, Crawley, W.A. 6009

Australia.

[c] M. Paskevicius, Prof. C.Buckley

Department of Imaging and Applied Physics, Curtin University of

Technology, PO Box U1987, Perth, Western Australia, 6845, Australia.

7

Encapsulation and Sustained Release of Curcumin using Superparamagnetic Silica Reservoirs

Suk Fun Chin,[a] K. Swaminathan Iyer,*[a] Martin Saunders,[b] Tim St. Pierre,[c] Craig Buckley,[d] Mark Paskevicius[d] and Colin L. Raston*[a]


0

0.05

0.1

0.15

0.2

0.25

300 350 400 450 500 550 600

Absorbance

Wavelength Figure S1: UV absorbance silica particles loaded with curcumin

500 550 600 650 700

5

10

15

20

25

30

Inte

nsit

y

Wavelength (nm)

1 μm

(A) (B)

Figure S2: (A) Fluorescent image and (B) Emission spectra of silica particles loaded with curcumin

[a] S.F.Chin, Dr.K.S.Iyer and Prof. C.L.Raston Center for Strategic Nano-Fabrication, School of Biomedical,

Biomolecular and Chemical Sciences, The University of Western Australia, Crawley, 6009 Australia Fax: +61 86488 8683 E-mail: [email protected]; [email protected]

[b] Prof. T.S.Pierre Center for Microscopy, Characterization and Analysis, The University of

Western Australia, Crawley, W.A. 6009 Australia. [c] M. Paskevicius, Prof. C.Buckley Department of Imaging and Applied Physics, Curtin University of Technology, PO Box U1987, Perth, Western Australia, 6845, Australia.

Chapter 6: References

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