Magnetic Magnetic bioferrofluids bioferrofluids with tailoredwith tailored

properties for biomedical applicationsproperties for biomedical applications

F. Palacio, A. Millán, E. Natividad, A. Arizaga, A. Urtizberea,

N.J.O. Silva, R. Piñol, G. Ibarz, M. Castro and A. Mediano

Instituto de Ciencia de Materiales de Aragón.CSIC - Universidad de Zaragoza. Zaragoza

NanoBio-Europe2008Barcelona, 9-13, june 2008

OutlineOutline

•• Introduction: biomedical applications ofIntroduction: biomedical applications ofmagnetic nanomagnetic nanoparticlesparticles

•• Nano vsNano vs. . NanoNano

•• Superparamagnetic particlesSuperparamagnetic particles

•• HyperthermiaHyperthermia

•• Polymeric route to the Polymeric route to the prproduction of iron oxideoduction of iron oxidenanoparticles and nanoparticles and ferrofluidsferrofluids

Interest of Magnetic Nanoparticles inInterest of Magnetic Nanoparticles inBiomedical ApplicationsBiomedical Applications

Controllable sizes (Controllable sizes (≥≥ 100 nm - 100 nm - ≤≤ 10 nm) and 10 nm) andshapes (spheres, needles, beads)shapes (spheres, needles, beads)

Magnetic functionality => Magnetic functionality => magnetic fieldsmagnetic fieldspenetrate human tissuespenetrate human tissues

They obey Coulomb They obey Coulomb’’s law => can be controlleds law => can be controlledat a distanceat a distance

They transfer energy from an ac magnetic field They transfer energy from an ac magnetic fieldand convert it into heatand convert it into heat

Biomedical Applications of MagneticBiomedical Applications of MagneticNanoparticlesNanoparticles

DiagnosisMRI • Sensors

• Cell separation• Enzyme tests• Immunoassays

Therapy

· Hyperthermia (MFH)· Radiotherapy/ /Chimietherapy- -combined MFH· Thermal ablation

• Drug delivery• Arthritic treatments• Chronic disseases

Magnetic (Magnetic (NanoNano)Particles)Particles

Magnetic domains

+ Magnetic field + Magnetic field HH

MagnetisationM = m/Vm

Magnetic forceMagnetic force

Fm = Vm (M/H)(B·H/2)

Atomic World ruled byAtomic World ruled bythe laws of Quantumthe laws of QuantumMechanicsMechanics

Macroscopic World asMacroscopic World aswe perceive it, ruled bywe perceive it, ruled byClassical Mechanics andClassical Mechanics andElecElectromagnetism lawstromagnetism laws

Introduction to Introduction to nano-sizenano-size

From the physics point of viewFrom the physics point of view Nano-Nano- is the interfaceis the interfaceseparating the atomic from the macroscopic scalesseparating the atomic from the macroscopic scales

NANONANOWORLDWORLD

In the atomic world there are propertiesIn the atomic world there are propertieswhose critical lengths are at the nano-scalewhose critical lengths are at the nano-scale

Introduction to Introduction to nano-sizenano-size

At the scale below a critical length new propertiesAt the scale below a critical length new propertiesarise that can give rise to new materials and newarise that can give rise to new materials and new

applicationsapplications

Critical lengths such asCritical lengths such as

one electron Fermi wavelength one electron Fermi wavelength

exciton exciton Bohr radiusBohr radius

single magnetic domain length single magnetic domain length

lead to Quantum Dots and Superparamagnetic particleslead to Quantum Dots and Superparamagnetic particles

Single-domain particles =>Single-domain particles =>superparamagnetssuperparamagnets

When the size of a particle is smaller than theWhen the size of a particle is smaller than theminimum allowing the formation of domainsminimum allowing the formation of domains

((≈≈ 20 - 30 nm), it becomes single-domain and 20 - 30 nm), it becomes single-domain and

superparamagneticsuperparamagnetic

Biomedical Applications ofBiomedical Applications ofMagnetic NanoparticlesMagnetic Nanoparticles

•• Cell separationCell separation

•• Enzyme Enzyme inmunoassay inmunoassay teststests

•• Sensitive detection of viral agentsSensitive detection of viral agents

•• Drug deliveryDrug delivery

•• Contrast agents for magnetic imagingContrast agents for magnetic imaging

•• DMS as luminescent sensorsDMS as luminescent sensors

•• HyperthermiaHyperthermia

•• ··· the limit is the imagination ······ the limit is the imagination ···

H

FM

Md < critical

sizeSPM

Magnetic force-driven applicationsMagnetic force-driven applications

Transfected cell separation in gen therapy

Magnetic separation

Magnet

Blood vessel

Tissue

Drug delivery

Magnetic nanoparticles as contrastMagnetic nanoparticles as contrastagents for MRIagents for MRI

Huge magnetic moments as compared to Huge magnetic moments as compared to GdGdchelateschelates

Proton relaxation is affected by the largeProton relaxation is affected by the largemagnetic field heterogeneity in the magnetic field heterogeneity in the vecinity vecinity of theof theparticlesparticles

Can induce >10 fold increase in protonCan induce >10 fold increase in protonrelaxivitiesrelaxivities

Shortening of relaxation times, particularly TShortening of relaxation times, particularly T22((negative contrastnegative contrast))

Very high resolution, close to molecular levelVery high resolution, close to molecular level

HypertermiaHypertermia

Magnetic systems can convert energy into heat under the effectsMagnetic systems can convert energy into heat under the effectsof an alternating magnetic fieldof an alternating magnetic field

•• inductive heating (eddy currents) inductive heating (eddy currents)

•• hysteresis hysteresis losseslosses

No relevant at low ac magnetic fieldsNo relevant at low ac magnetic fields

!

PFM

= "µ0f HdM#

HypertermiaHypertermia

Superparamagnetic particlesSuperparamagnetic particles

ϖτ ~ 1 Emáx=>

Emáx (γ-Fe2O3, 10 nm)ϖ ~ 1 MHz

•• NeNeél él relaxationrelaxation

•• Rotational Brownian motion Rotational Brownian motion

-1

-0,5

0

0,5

1

-300 -200 -100 0 100 200 300 400

θ

U

220

1"

!"

"!##

+=

!

PSPM

= "1

2µ

0#"$H

0

2

Biological range: Biological range: 50 kHz 50 kHz ≤≤ f f ≤≤ 1200 1200kHz, H < 15 kHz, H < 15 kA/mkA/m and (H · f)max =and (H · f)max =

485 485 kHzkHz··kA/mkA/m

!

"B

=3#V

H

kT

!

"N

= "0 exp KV / kT( )

!

1

"eff

=1

"N

+1

"B

The shortest relaxation timeThe shortest relaxation timeconstant tends to dominate theconstant tends to dominate theeffective relaxation timeeffective relaxation time

HypertermiaHypertermia

HypertermiaHypertermia

coilscoils

thermalthermalinsulationinsulation

thermometerthermometer

samplesample

!

SAR(W / g) = C"T

"t#

1

mSPM

ΔT/ Δt is calculated fromthe initial slope

Non-adiabatic ----> heat losses ----> estimation of SARNon-adiabatic ----> heat losses ----> estimation of SAR

Adiabatic set-up for hyperthermiaAdiabatic set-up for hyperthermia

Eva Natividad et al., Appl. Phys. Letters, 92, 0991136 (2008)

Sample, Termometer

X

X

Coil (GEPGroup, UZ)

Vacuum

Electricalconnections

LN2

(no metallic components)

T

tacoff

acon

acoff

ΔT

Δt

Operation range:HH = 0 - 8kA/m = 0 - 8kA/m

ff = 50 = 50 –– 300 kHz 300 kHz

H < 15 H < 15 A/mA/m, f < 400 kHz, f < 400 kHz

Biological applicationsBiological applications::H · f < 4.85 · 10H · f < 4.85 · 1088 A/m·s A/m·s

Adiabatic set-up for hyperthermiaAdiabatic set-up for hyperthermia

(see also poster from Ana (see also poster from Ana ArizagaArizaga))

Superparamagnetic Particles forSuperparamagnetic Particles forBiomedical ApplicationsBiomedical Applications

functionalized magnetic nucleus (maghemite, other)

Protecting core,anchoring element

biological vector:antigens, etc.

Functionalized tailfor therapeuticuses: drugs,enzyms, etc.

Superparamagnetic Particles forSuperparamagnetic Particles forBiomedical ApplicationsBiomedical Applications

The magnetic nucleus should preferable bebiocompatible, otherwise a strong protectivecoating (+ its validation!!) is required for invivo applications

• Maghemite

• Magnetite

• Co-ferrites and other (in vitro)

Tailoring magnetic nanoparticlesTailoring magnetic nanoparticles

Production methods must focus on:

• narrow size dispersion

• variable size control

• variable shape control

• control of organization andparticle-matrix interactions

Interest:Interest:• Important applications of magnetic nanoparticles are very exigent

regarding size and shape dispersion

• Polymers can provide a way to avoid agglomeration and easysurface functionalisation

DSAR

ΔD

Onlyparticleswithin anarrow ΔDare efficient

Size dependence of the SpecificAbsorption Rate (SAR)

ω, H

Tailoring magnetic nanoparticlesTailoring magnetic nanoparticles

1. Synthesize the magnetic particles,normally magnetite or maghemite

2. Avoid agglomeration by keeping themdispersed in a surfactant

3. Eliminate surfactant to stabilise theparticles either electrostatically orsterically

4. Functionalise the particles

Thermal decomposition of Fe(CO)5.Thermal decomposition of Fe(CO)5.

Oleic acid

Solvent: octhyl ether

ΔFe(CO)5

Maghemite, 10 nm Maghemite, 20 nmMaghemite, 14 nm

Size dispersion ≈ 5% Size ranges: 4-20 nm

Tailoring magnetic nanoparticlesTailoring magnetic nanoparticles

Fe(CO)5

Oleic acidOcthyl ether

Δ

Me3NO

γ-Fe2O3

(RO)3SiR’R’ Si OH

O

O

TEM EFTEM

Tailoring magnetic nanoparticlesTailoring magnetic nanoparticles

+

Hydrophilic PVP polymer

metal salt

(eventually)

disgregation ferrofluid

homogeneous gel nanocomposite

base

A. Millán and F. Palacio, Applied Organometallic Chemistry, 15, 396-400 (2001)A. Millán et al., Acta Materialia 55, 2201-9 (2007).A. Millán, F. Palacio, G. Ibarz, Patent PCT/EP2007/058312

Tailoring magnetic nanoparticlesTailoring magnetic nanoparticleswith polymerswith polymers

PEG

copolymerisation

Either spheres and rods can beselectively prepared

Simple and quick single-pot reactionSpheric particle sizes 2 - 15 nmSize dispersion ±10%No aggregation. Uniform distribution.

0

20

40

60

80

0 1 2 3 4 5 6 7 8 9 10D (nm)

Polymers can provide a wayPolymers can provide a wayto avoid agglomeration andto avoid agglomeration andeasy surface functionalisationeasy surface functionalisation

Tailoring magnetic nanoparticlesTailoring magnetic nanoparticleswith polymerswith polymers

Bioferrofluids, pH = 7.4

Adequate polymer design leads to nanocomposites which disperse ina phosphate buffer saline solution as nanoparticles coated with PEG

FerrofluidFerrofluidsamplesample

TestsTestssamplessamples

Maghemite ferrofluidsMaghemite ferrofluids

Maghemite ferrofluidsMaghemite ferrofluids

Ferrofluids Ferrofluids are are persistent for persistent for more more than one monththan one month

-4 10-7

0

4 10-7

8 10-7

1.2 10-6

0 50 100 150 200 250 300 350

MAGNETIC STABILITY

Freshly preparedAfter one month

M(e

mu

) a.u

.

T(K)

ZFC//FC(25Oe)

0

0.001

0.002

0.003

0.004

0.005

0

0.001

0.002

0.003

0.004

0.005

0.006

0 10 20 30 40 50 60 70 80

Maghemite rods

!''(e

mu

/Oeg

Fe

2O

3)

- 24h

!''(e

mu

/Oeg

Fe

2 O3 ) - 1

mo

nth

Temperature (K)

1 Hz

117 Hz

852 Hz

24 hours1 month

NMR NMR relaxometryrelaxometry

0 50 100 150 2000,1

1

100 100 200

100

200

300

R2 (

mM

-1

s -1

)

! (MHz)

T=297K

Fe = 11.2 mg / ml

R1 (

mM

-1

s -1

)

! (MHz)

Endorem PEI-25 PEI-COOH60 PEI-COOH75 PEI-500

• R1 and R2 relaxivities compared with commercial ENDOREM®

• PEI-25 R1 relaxivity compares to ENDOREM®

• PEI-25 R2 relaxivity higher over the whole field range.

Relaxivities R1 and R2 quantifiesthe increased nuclear relaxationrates 1/T1 and 1/T2 of nuclei(typically protons) due to thepresence of magnetic centers

Hyperthermia valuesHyperthermia values

A. Millán et al (in preparation)

Fortin et al, J. Am. Chem. Soc., 129 (9)(2007) 2628-2635

G. Glöckl, R. Hergt, M. Zeisberger et al.,JP: CM 18 (2006) S2935–49

Wang et al, J. Magn. .Magn. Mat., 293(2005) 334-340

ibid., 280 (2004) 358-368

Hergt et al, J. Magn. .Magn. Mat., 270(2004) 345-357

Ma et al, J. Magn. .Magn. Mat., 268(2004) 33-39

Hilger et al, Academic Radiology, 9(2002) 198-202

Brunsentsov et al, J. Magn. .Magn.Mat., 225 (2001) 113-117

Hiergeist et al, J. Magn. .Magn. Mat.,201 (1999) 420-422

Ref.

144

9.2

14.7

93.7

22.9

27.4

0.8

2.1

12.9

10.8

27.211.3

SAR *(W/g)

158

1650

98

85

900

600

28.9

75.6

93

210

200

78

SAR(W/g)

1.1

35.8

6.6

0.9

12.7

9.5

5.4

5.4

5.4

13.2

5.5

1.2 - 5.3

Hf/(Hf)max

150

700

400

63

410

410

80

80

400

880

410

410

f(kHz)

5

24.8

8

7

15

11.2

32.5

32.5

6.5

7.2

6.5

1.4 - 6.3

H(kA/m)

d ≈12needles

16.5 - 24.8

7.9 - 22.3

6 - 10

7 - 18

15

81 - 416

7.5 - 46

10

10 - 12

100

8

Size (nm)

maghemite

maghemite

maghemite

magnetite

maghemite

maghemite

magnetite

magnetite

magnetite

maghemite

magnetite

magnetite

Material

(*) Referred to standard H = 4.85 (*) Referred to standard H = 4.85 kA/mkA/m, f = 100 kHz values, f = 100 kHz values

Nanomagnets for Nanomagnets for theragnostictheragnostic

MRIMRIPAVIAPAVIA

χχ acac at at ra

dio-radio-

Frequencies

Frequencies

Florence

Florence

Hyperthermia

Hyperthermia

Zaragoza

Zaragoza

Low NMRfrequencies (few kHz- 10 MHz) down to 4.2 K, multinuclear operatinal mode,

Adiabatic and non adiabatic magnetothermia set ups (50 - 500 kHz,0 - 8 kA/m, down to 80K

χac susceptibi-

lity set up50 - 500 kHz

1.5 - 350 K

Interested Interested may may contactcontact: : amillan@unizaramillan@unizar.es.es or palacio@unizaror palacio@unizar.es.es

Group TERMOMAGGroup TERMOMAG

Ana ARIZAGAAna ARIZAGA

Ramon BURRIELRamon BURRIEL

Javier CAMPOJavier CAMPO

Miguel CASTROMiguel CASTRO

Clara GONZALEZClara GONZALEZ

Miguel A. GONZALEZMiguel A. GONZALEZ

Gemma Gemma IBARZIBARZ

Ricardo LOPEZRicardo LOPEZ

Emma LYTHGOEEmma LYTHGOE

Eva NATIVIDADEva NATIVIDAD

Angel MILLANAngel MILLAN

M Carmen MORON M Carmen MORON

Lucas OLOLucas OLO

Elias PALACIOSElias PALACIOS

Rafael PIÑOLRafael PIÑOL

Ines Ines PUENTEPUENTE

Alberto RODRIGUEZAlberto RODRIGUEZ

Clara RODRClara RODRIGUEZIGUEZ

Cristina SAENZ de PIPAONCristina SAENZ de PIPAON

Jorge SANCHEZJorge SANCHEZ

Nuno Nuno J. O. SILVAJ. O. SILVA

Leticia TOCADOLeticia TOCADO

Ainhoa Ainhoa URTIZBEREAURTIZBEREA

ACKNOWLEDGEMENTSACKNOWLEDGEMENTS

MEC, MAT2004-03395-C02-01

CONSOLIDER - MAT2007-61621

European Network ofExcelence, 6FP

Programa CENIT - Ingenio 2010Industrial Consortium

Proyecto de colaboración industrialEscalado de producción

MolecularMolecularNanoscienceNanoscience

Programa ConsoliderIngenio 2010