Assessment of Mixed Monolayer-protected Gold Nanoparticles
Assembly in Solution: Study and Characterization
Ana Isabel Martins Tiago Fernandes
in fulfillment with thesis requirement for the degree of
Master of Science in Biological Engineering
Examining Committee
Chair:
Prof. Dr. Luís Joaquim Pina da Fonseca (IST)
Advisors:
Prof. Dr. João Pedro Rodrigues Estrela Conde (IST)
Prof. Dr. Molly M. Stevens (IC)
Reviewer:
Prof. Dr. Duarte Miguel de Franca Teixeira dos Prazeres (IST)
September 2010
Assessment of Mixed Monolayer-protected Gold Nanoparticles
Assembly in Solution: Study and Characterization
Ana Isabel Martins Tiago Fernandes
in fulfillment with thesis requirement for the degree of
Master of Science in Biological Engineering
Examining Committee
Chair:
Prof. Dr. Luís Joaquim Pina da Fonseca (IST)
Advisors:
Prof. Dr. João Pedro Rodrigues Estrela Conde (IST)
Prof. Dr. Molly M. Stevens (IC)
Reviewer:
Prof. Dr. Duarte Miguel de Franca Teixeira dos Prazeres (IST)
September 2010
2 of 94
Acknowledgements
Firstly I would like to express my deepest gratitude to Prof. Molly Stevens, my supervisor at Imperial
College, for giving me the opportunity to work on this interesting project. Her constant encouragement,
interest and insightful support had a considerable impact on my success in the course of my work. She
gave me the freedom to explore new ideas while keeping me focused on my goals. It was a privilege to
have worked in Prof. Molly Stevens’ group.
I am also thankful to Vanessa LaPointe for her brilliant ideas and continuous support and to Nia Bell, my
synthesis mate, for all her availability and sympathy given at laboratory!
I also want to express my sincere gratitude to Dr. Nicolas Schaeffer he was la lumière au bout du tunnel
when I most needed it. Merci beaucoup!
My experience at IC was definitely improved by the constant sense of mutual aid and friendship with the
greatest office mates (e.g. Air, Benji, David, Kristy, Jess, John, Maria, Mathew, Pinyuan, Stuart…!)
I want to thank Prof. Stellacci and Dr. Jeffrey Kuna at MIT for their availability and for their encouraging
and helpful advices.
I would also like to thank Prof. João Pedro Conde from IST, for his support, patience and
encouragement.
I am short of words for the immeasurable emotional and financial support of my family, especially of my
parents, brother, sister and grandmother. To all my friends who helped me in one way or another.
Obrigada Gonçalo, Carmo e Rui.
Finally, I dedicate this work to Francisco, he is still too young to understand the importance he had and
the inspiration and strength he gave during the past year.
3 of 94
“Things on a very small scale behave like nothing that you have any direct experience about. They do
not behave like waves, they do not behave like particles, they do not behave like clouds or billiard balls,
or weights on springs, or like anything that you have ever seen.”
Richard Feynman
4 of 94
Abstract
With the recent progress in nanoscience, gold nanoparticles (NP) have found widespread use in many
areas of scientific research ranging from biology to physics and medicine. These particles are easily
synthesized and can be readily coated with a self-assembled monolayer (SAM) of thiolated ligand
molecules that, depending on the synthesis conditions, phase-separate into ordered domains (rippled
domains) that encircle the gold core. This ligand shell confers stability against coalescence and controls
the particle's interactions with its environment (e.g. assembly, electron transfer ability). Furthermore,
the ability to manipulate and assemble these nanomaterials through the controlled functionalization of
their ligand shell is crucial for their incorporation into and development of new nanoparticle based
materials and devices. The engineering process behind the creation of those new nanostructures is
strongly dependent on the methods used to position these particles in specific locations in relation to
their neighbouring counterparts. Besides the individual NP properties, the gl obal properties and
functions of the assembled nanostructured systems are also ruled by the interparticle distances and
interactions between each of the cons tituent nanoparticle.
Herein, we describe the synthesis and characterization of near monodisperse homo and mixed
monolayer-protected gold NPs. Crucial information about these particles spontaneous assembly in
solution is also provided and a procedure to prevent this unwanted aggregation is presented.
Furthermore, in order to investigate how, depending on their ligand shell morphology (rippled or not),
these particles assemble, studies (UV-Vis and NMR spectroscopies and transmission electron microscopy
(TEM)) conducted on five different series of monolayer-protected metal nanoparticles (MPMNs) in the
presence of dithiol containing molecules are reported. High linker to gold NPs ratios (~2500) induce fast
and high rates of place-exchange reactions between the ligands in the SAM and the incoming linker
molecules, and three-dimensional ball-type aggregates form. In contrast, lower ratios (~125) do not
induce NPs assembly. We conclude it is very challenging to have a precise control over the number of
functional groups attached to each particle, therefore further studies with different solvents, surfactants
and cross-linker molecules should be performed.
Keywords:
Gold Nanoparticles, Self-assembled monolayer, Phase-separated ordered domains,
Nanoparticles assembly, Dithiol, Trasmission electron microscopy
5 of 94
Resumo
Com os recentes progressos em nanociência, as nanopartículas de ouro têm sido alvo de estudo
intensivo apresentando uma ampla utilização em diversos campos da investigação científica que se
estendem por áreas como a biologia, a física e a medicina. Estas partículas são facilmente sintetizadas,
sendo espontaneamente revestidas por uma monocamada auto-agregada de moléculas ligantes que,
dependendo das condições de síntese, se separam e organizam formando domínios ordenados à volta
da partícula de ouro. Este revestimento de ligandos confere estabilidade contra coalescência e controla
as interacções destas partículas com o ambiente local (i.e. agregação, capacidade de transferência de
electrões). Além disso, a capacidade de manipular e agrupar estes nanomateriais através da
funcionalização controlada da monocamada de ligandos é crucial para o desenvolvimento e/ou sua
incorporação em novas nano-estruturas e dispositivos. O processo de criação destas nano-estruturas
está dependente dos métodos utilizados para posicionar estas partículas em localizações específicas em
relação às partículas vizinhas. Para além das propriedades individuais de cada nanopartícula, as
características e propriedades globais das nano-estruturas agregadas dependem igualmente das
distâncias e interacções estabelecidas entre cada uma das partículas constituintes.
Na presente tese, a síntese e caracterização de nanopartículas protegidas por monocamadas compostas
por um (homogéneas) ou mais tipos (mistas) de ligandos é descrito. Informação crucial sobre a
agregação espontânea destas partículas em solução é fornecida e um processo para prevenir esta
agregação indesejada é apresentado.
A fim de investigar o modo de agregação destas partículas na presença de um linker (ditiol), estudos
(Espectroscopia de absorção UV-Vis, espectroscopia RMN e microscopia de transmissão electrónica) em
cinco séries de nanopartículas com diferentes morfologias na monocamada de ligandos (ordernada ou
não ordenada) são descritos. Rácios linker por nanopartícula de ouro elevados (~2500) induzem rápidas
trocas entre os ligandos da monocamada e as moléculas do linker, formando-se agregados tridimensionais de forma circular. Por outro lado, rácios baixos (~125) não induzem a agregação das
partículas. Conclui-se que é muito difícil controlar o número de grupos funcionais na superfície de cada
partícula e estudos com outros solventes, surfactantes e linker deverão ser seguidos.
Palavras-chave:
Nanopartículas de ouro,
Monocamada auto-agregada, domínios separados
ordenadamente, Organi zação de nanopartículas, Ditiol, Microscopia de transmissão electrónica
6 of 94
Contents
Acknowledgements ................................................................................................................3
Abstract................................................................................................................................5
Resumo ................................................................................................................................6
Contents ...............................................................................................................................7
List of Tables .........................................................................................................................9
List of Figures ...................................................................................................................... 10
List of Abbreviations and Symbols........................................................................................... 13
1.
Introduction ................................................................................................................. 15
2.
Literature Review .......................................................................................................... 16
2.1.
Nanoscience ...................................................................................................... 16
2.2.
Nanoparticles (NPs): Nanoscience Building Blocks .................................................... 17
2.2.1.
Gold Nanoparticles...................................................................................... 18
2.2.1.1.
Historical Background on Gold Nanoparticles ............................................... 18
2.2.1.2.
Stabilization of Gold Nanoparticle Dispersions .............................................. 19
2.2.1.3.
Properties and Characteristics of Gold Nanoparticles. .................................... 19
2.2.1.4.
Surface Plasmon Resonance ...................................................................... 20
2.3.
Monolayer-Protected Metal Nanoparticles.............................................................. 20
2.4.
Self-assembled Monolayers (SAMs) ....................................................................... 20
2.4.1.
2.4.1.1.
Experimental Observations ....................................................................... 22
2.4.1.2.
Properties of rippled MPMNs .................................................................... 25
2.4.2.
2.5.
Phase-Separated Ordered Domains ................................................................ 22
Place-Exchange Reactions. ............................................................................ 25
MPMNs Assembly............................................................................................... 27
2.5.1.
Polar Defects.............................................................................................. 27
2.5.2.
Dithiols ..................................................................................................... 29
2.5.3.
Other types of Assembly .............................................................................. 30
3.
Thesis Outline ............................................................................................................... 31
4.
Materials and Methods .................................................................................................. 32
4.1.
Characterization Techniques for the Nanoscale ........................................................ 32
4.1.1.
UV/Visible Spectroscopy............................................................................... 32
4.1.2.
Nuclear Magnetic Resonance (NMR) Spectroscopy............................................ 34
4.1.3.
Transmission Electron Microscopy ................................................................. 35
4.2.
NP Synthesis ...................................................................................................... 36
4.3.
NP Characterization ............................................................................................ 37
4.3.1.
UV/Visible Absorption Spectroscopy............................................................... 37
7 of 94
4.3.2.
Transmission Electron Microscopy (TEM) ......................................................... 38
4.3.3.
Nuclear Magnetic Resonance (NMR) Spectroscopy............................................ 38
4.4.
5.
NP Assembly...................................................................................................... 38
4.4.1.
UV/Visible Absorption Spectroscopy............................................................... 39
4.4.2.
Transmission Electron Microscopy (TEM) ......................................................... 39
Results and Discussion.................................................................................................... 40
5.1.
Synthesis and Analysis of Monolayer-Protected Metal Nanoparticles ........................... 40
5.2.
MPMNs Molarity................................................................................................ 48
5.2.1.
5.3.
Number of Gold Atoms: ............................................................................... 48
Cross-linking of MPMNs in solution........................................................................ 52
5.3.1.
UV/Visible Analysis...................................................................................... 52
5.3.2.
TEM Analysis .............................................................................................. 56
5.3.2.1.
Hypothesis ............................................................................................. 56
5.3.2.2.
TEM Sample Preparation .......................................................................... 58
5.3.2.3.
Stability of Particles in Solution.................................................................. 59
5.3.2.4.
Assembly Variations with Time .................................................................. 66
5.3.2.5.
Assembling Behaviours with Cross -linker Concentration................................. 70
5.3.2.6.
Assembly after Purification Steps ............................................................... 75
5.3.2.7.
New cross-linker approach ........................................................................ 77
6.
Conclusions and Future Work .......................................................................................... 81
7.
References ................................................................................................................... 83
Appendix ............................................................................................................................ 92
8 of 94
List of Tables
Table 1 Amounts of ligands used in the synthesis of NPs with various compositions .......................... 37
Table 2 Table of average core size ± Standard deviation for MH/OT MPMN series: ........................... 44
Table 3 Total yield and efficiency of MPMNs synthesis................................................................. 45
Table 4 Plasmon peak values and intensities for each of the MH/OT MPMNs series. ......................... 46
Table 5 Estimated gold clusters characteristic dimensions. ........................................................... 49
Table 6 Linear formula, molecular weight, molar concentration and average number of nanoparticles
per gold cluster (NNPs ) for each set of MPMNs. ........................................................................... 50
Table 7 TEM grid and TEM image considerations. ....................................................................... 50
Table 8 1,9-nonanedithiol and 1,16-hexadecanedithiol chemical structure and molecular weight ........ 51
Table 9 Number of mols (N) and number of cross-linker molecules (Ncross-linker) for different molar
concentrations considering a final volume of 0.5 ml of cross -linker solution..................................... 51
Table 10 Molar concentration (C), number of mols ( N) and number of NPs and thiols (NNanoparticles and
Nthiols respectively) correspondent to two different MPMNs final concentrations, considering a final
MPMNs solution volume of 0.5 ml. .......................................................................................... 51
Table 11 Cross-linker to gold NP and cross -linker to thiol ratios ..................................................... 52
Table 12 Different amounts of time samples were left under stirring after the purification steps......... 61
Table 13 Plasmon peak wavelengths and intensity variations with time for MPMNs solution .............. 65
Table 14 Table with the calculated linker to particle and linker to thiol ratio values........................... 70
Table 15 Different conditions used in the assembly study, when the solutions analysed were submitted
to the purification protocol..................................................................................................... 75
9 of 94
List of Figures
Figure 1 Growth of manufacturer-identified, nanotechnology products. ......................................... 17
Figure 2 The Lycurgus Cup ...................................................................................................... 18
Figure 3 Faraday’s colloidal ruby gold ....................................................................................... 19
Figure 4 Representative diagram of an SAM of alkanet hiolates supported on a gold NP .................... 21
Figure 5 STM image of MPMNs showing phase-separated ordered domains .................................... 22
Figure 6 Surface plot of the ligand shell presenting the contours of the ripples................................. 23
Figure 7 Three dimensional rendering of gold NPs STM height images ........................................... 23
Figure 8 Schematic representation of the free volume available to ligands on curved surfaces ............ 24
Figure 9 Equilibrium arrangements of binary SAMs adsorbed on NPs with different degrees of curvature
determined through mesoscale simulations............................................................................... 24
Figure 10 Illustration of the processes involved in ligand place -exchange reactions on gold MPMNs .... 26
Figure 11 Illustration of a rippled MPMN showing one of the two diametrically opposed polar defects . 28
Figure 12 TEM images of MPMN chains. ................................................................................... 28
Figure 13 TEM images of linear hierarchical assemblies of MPMNs ................................................ 29
Figure 14 Schematics of MPMN ligand shell chemistry and morphology.......................................... 31
Figure 15 Schematic depicting the promotion of an elect ron from an orbital in the ground state (π) to an
unoccupied orbital at a higher energy level (π*) ......................................................................... 33
Figure 16 Light passing through the cuvette containing the sample. ............................................... 33
Figure 17 Components of a transmission electron microscope (TEM). Adapted from [122] ................. 36
Figure 18 Chemical structures of 1-octanethiol and 6-mercapto-1-hexanol...................................... 40
Figure 19 Representative TEM images obtained for four different MH/OT MPMNs synthesized showing
the size of the gold core of the different particles. ...................................................................... 41
Figure 20 Representative TEM images obtained for three different MH/OT MPMNs synthesized showing
the size of the gold core of the different particles. ...................................................................... 42
Figure 21 Histograms obtained for five different MH/OT MPMNs synthesized de picting core size
distributions. ....................................................................................................................... 43
Figure 22 Histograms obtained for two different MH/OT MPMNs synthesized depicting core size
distributions. ....................................................................................................................... 44
Figure 23 UV-Vis absorption spectra of MPMNs suspended in EtOH ............................................... 46
1
Figure 24 H NMR spectrum obtained for 2:1 MH/OT in CD 3 OD ..................................................... 47
Figure 25 Film of MH/OT 2:1 nanoparticles imaged by AM-AFM in ultrapure water........................... 47
Figure 26 Graph showing the relation between the number of layers of a gold NP and the total number
of gold atoms. ...................................................................................................................... 48
Figure 27 Change of absorbance in the 450-900 nm region of UV/Vis spectra for 1:1 MH/OT MPMN
solution upon addition of 1, 5 and 10 mM solution of cross -linker.................................................. 53
Figure 28 Change of absorbance in the 450-900 nm region of UV/Vis spectra for 5:1, 2:1, 1:2 and 1:5
MH/OT MPMNs upon addi tion of 5 mM of NDT solution. ............................................................. 54
10 of 94
Figure 29 SPR peak shifts for all the batch of the 5 sets of MPMNs tested at different times. .............. 55
Figure 30 Stabilization time .................................................................................................... 56
Figure 31 Observed assembly of the 1:1 MH/OT gold MPMNs. ...................................................... 57
Figure 33 Predicted structures of the assembled 2:1 and 1:2 MH/OT gold MPMNs ........................... 57
Figure 33 Expected structures of the assembled 5:1 and 1:5 MH/OT gold MPMNs ............................ 58
Figure 34 TEM images of 2:1 MH/OT MPMNs solution................................................................. 59
Figure 35 TEM images of 2:1 MH/OT MPMNs solution after sonication........................................... 60
Figure 36 TEM images of 2:1 and 1:1 MH/OT MPMNs solutions 2 hours after purification steps . ......... 60
Figure 37 TEM pictures of the 5:1 MH/OT MPMNs solution 3 days after purification steps ................. 61
Figure 38 TEM pictures of the 1:1 MH/OT MPMNs solution 1 hour after purification steps. ................ 62
Figure 39 TEM pictures of the 1:1 MH/OT MPMNs solution 5 hours after purification steps................ 62
Figure 40 TEM pictures of the 1:1 MH/OT MPMNs solution 3 and 14 days after purification steps ....... 63
Figure 41 TEM images of the 1:1 MH/OT MPMNs solution 5 days after purification steps................... 63
Figure 42 UV-Vis absorption spectra of 1:1 MH/OT MPMNs dissolved in EtOH The sample was kept
untouched during a period of twelve weeks, and during this time interval several UV/Vis samples were
taken periodically ................................................................................................................. 64
Figure 43 1:1 MH/OT in Methanol............................................................................................ 65
Figure 44 TEM images of 1:1 MH/OT MPMNs solution 1 hour after adding the cross -linker, NDT. ........ 66
Figure 45 TEM images of 1:2 MH/OT A MPMNs solution 1 hour after adding the cross-linker, NDT ...... 66
Figure 46 TEM images of 1:2 MH/OT B MPMNs solution 1 hour after adding the cross-linker, NDT ...... 67
Figure 47 TEM images of 5:1 MH/OT MPMNs solution 1 hour after adding the cross -linker, NDT ......... 67
Figure 48 TEM images of 1:1 A and 1:2 B MH/OT MPMNs solutions 1 minute after adding the crosslinker (NDT) solution ............................................................................................................. 68
Figure 49 TEM images of 1:2 B (same grid as figure 48 (b) but analysed two weeks after), 2:1 A and 5:1 A
MH/OT MPMNs solutions 1 minute after adding the cross-linker (NDT) solution............................... 69
Figure 50 TEM images of 1:2 MH/OT MPMNs solution 1 and 5 minutes after adding of cross-linker (NDT)
solution (1 mM) .................................................................................................................... 70
Figure 51 TEM images of 1:2 MH/OT MPMNs solution 1, 5 and 15 minutes after adding of cross-linker
(NDT) solution (0.5 mM)......................................................................................................... 71
Figure 52 TEM images of 2:1 MH/OT MPMNs 1 minute after adding the 1, 0.5 and 0.1 mM solution of
cross-linker (NDT). Scale bars: 50 nm ........................................................................................ 72
Figure 53 TEM images of 2:1 MH/OT sonicated MPMNs prepared 1 minute after adding the 0.5 and 0.1
mM solution of cross-linker, NDT. ............................................................................................ 72
Figure 54 TEM images of 2:1 MH/OT MPMNs 1 and 5 minutes after adding the “old” 1 mM solution of
cross-linker, NDT .................................................................................................................. 73
Figure 55 TEM images of 2:1 MH/OT MPMNs 1 and 5 minutes after adding the “fresh” 1 mM solution of
cross-linker, NDT .................................................................................................................. 73
Figure 56 TEM pictures of the 1:1, 2:1 and 5:1 MH/OT purified MPMNs solutions prepared 1 and 10
minutes after adding the 1mM cross -linker solution, NDT............................................................. 76
11 of 94
Figure 57 TEM pictures of the 1:1 MH/OT purified MPMNs solution prepared 10 minutes, 1 and 12 hours
after adding the 0.01 mM cross-linker solution (NDT) .................................................................. 77
Figure 58 Schematic illustrating the cross -linking process for the two different dithiol containing
molecules used: NDT and HDDT. .............................................................................................. 78
Figure 59 TEM images of 1:1 MH/OT MPMNs solution 1 and 15 minutes, 1, 3 and 18 hours after adding a
0.01 mM cross-linker solution (HDDT) ....................................................................................... 79
Figure 60 Schematic illustrating the incoming cross -linker molecule coiling around in the gold NP
forming two thiol-gold bonds in the same particle. ..................................................................... 79
Figure 61 TEM images of 1:1 MH/OT MPMNs solution 1 minute after adding a 5 mM cross -linker
solution (HDDT) .................................................................................................................... 80
12 of 94
List of Abbreviations and Symbols
AM-AFM
Amplitude modulation - Atomic force microscopy
CCD
Charge-coupled device
DCM
Dichloromethane
DNA
Deoxyribonucleic acid
Et 2 O
Diethyl ether
EtOH
Ethanol
FTIR
Fourier transform infrared spectroscopy
HDDT
1,16-hexadecanedithiol
MH
6-mercapto-1-hexanol
MPMN
Monolayer-protected metal nanoparticles
MW
Molecular Weight
NDT
1,9-nonanedithiol
NMR
Nuclear magnetic resonance
NP
Nanoparticle
OT
1-octanethiol
PDI
Polydispersity index
PMMA
Poly(methyl methacrylate)
SAM
Self-assembled monolayer
SPR
Surface plasmon resonance
STM
Scanning tunnelling microscopy
TEM
Transmission electron microscopy
UV
Ultraviolet
Vis
Visible
I0
Intensity of light entering the sample
It
Intensity of light exiting the sample
A
Absorbance
T
Transmittance
ε
Absorptivity
c
Concentration
l
Path length
r
Microscope resolution
λ
Wavelength
µ
Medium refractive index
α
Semi-angle of microscopes aperture
13 of 94
N Au
Total number of gold atoms
d
Particles’ average diameter
VAu
Particles’ volume
N thiols
Number of ligands per gold nanoparticle
L TEM image
Square side of TEM image
ATEM image
Observable area per TEM image
dTEM grid
TEM grid total diameter
ATEM grid
TEM grid total area
NP/ATEM image
Total number of particles per TEM image
NP/ATEM grid
Total number of particles per TEM grid
N cross-linker
Number of cross-linker molecules
14 of 94
1. Introduction
The idea of building structures from the bottom -up, molecule by molecule or atom by atom, is one of
the core concepts of nanoscience [1]. However, the success and potential behind this versatile “bottomup” approach is dependent on two main points: the first being the synthesis, at the nanoscale, of
building units and the second being the organization of these nanopieces together into a device or
material with predefined and sometimes sophisticated structures and properties. Generally
nanomaterials’ properties are not only dependent on each unit that is forming the final structure, but
also on the space and type of interaction existent between them. In the past thirty years a remarkable
success has been attained in the development of synthesis protocols for several types of nanobuilding
units, e.g. nanoparticles (NPs), nanofibers and nanorods [2]. In fact, these nanosized materials have
already been introduced in some commercial applications such as clothing and footwear (i.e. stainresistant trousers) [3, 4], titanium dioxide present in anti-aging cosmetic creams [5], and carbon
nanotubes present in stronger but very light tennis rackets and bicycle frames [3, 6].
A large effort was specifically focused on the study of metal -core nanoparticles because of their optical,
electronic, and surface properties [7]. Metal nanoparticles have s hown various properties, such as single
electron transistor behaviour and surface plasmon resonance tuning and sensing [8, 9]. In the case of
monolayer-protected metal nanoparticles (MPMNs), the ligand shell that coats the particle surface
prevents coalescence when in solution and sometimes in solid state [10]. Moreover, this organic
molecules coating provides most of the particles' surface related properties, including assembly and
sensing properties as well as solubility [7, 10, 11]. Depending on the nature of the coating t hey can be
dried from and re-dissolved in solvents many times, or can be purified via dialysis, chromatography or
filtration [11]. However, controlled and directional assembly of these particles is still a bottleneck for
present nanotechnoloy research. Due to their considerably small dimensions, the association of these
nanobuilding units into complex and sophisticated materials, with the expected structure, properties
and functions represents an extremely complex scientific challenge.
15 of 94
2. Literature Review
2.1. Nanoscience
In the past, macro and microsized structures of well-defined nature, shapes and functionalities have
been well documented. However over the last decades, there has been a considerable increase of
interest in the field of nanoscience and na notechnology [12] due to the development of new tools for
analysing, imaging and manipulating nanometer scaled objects, such as scanni ng probe and electron
microscopes [10]. This current tendency to shrink materials’ dimensions is strongly encouraged because
of the unique material properties and performance advantages (compared to the bulk) that arise when
their dimensions are decreased to the nanometer length scales [13].
Nanoscience deals with the control and manipulation of systems and objects in which at least one
dimension is between 1 – 100 nm [10]. It is a highly inter-disciplinary field where chemistry plays a
fundamental role in the development of nanostructures and new synthetic methods, physics is used to
explain and characterize changes in the properties of matter with size, engineering is essential in
applying the understanding of nano-scale materials into useful devices and biology often acts as the
source of inspiration, with biological systems offering many examples of sophisticated nanostructures
interacting in complex networks suggesting new strategies with which to build artificial nanosystems
[14].
Their nano-scaled dimensions impart these nanostructured systems with many exclusive properties and
therefore they represent promising candidates for a variety of applications such as drug delivery and
sensing in medicine, catalysts in materials science, electronic and optical devices in electronics and
magnetic storage media [15]. Additionally, the capacity to systematically modify their properties by
controlling the structure and the chemical properties of these nanostructured systems ma kes them
well-suited for uses in more fundamental scientific studies of nano-scale interactions. This is of special
interest considering that most of the biological processes occur at the nano-scale (e.g. interactions
between water molecules, cells and proteins).
As dimensions diminish to nanoscale, the properties of matter become scale dependent and materials
exhibit different properties from that of the bulk material leading to interesting physical behav iour
based on quantum-mechanical phenomena, such as electron affinity, optical effects, conductivity,
ionization potential, superparamagnetism, electron tunnelling and surface plasma resonance (SPR) [13,
16, 17].
Those unique optical, electronic, and thermal properties deliberately pursued in the nanometer length
scale are strongly related to the surface-to-volume ratio that is drastically increased compared to the
bulk material [13]. This size-dependent behaviour leads, in the most extreme situation, to structures
16 of 94
where almost every atom in the structure is interfacial. Atoms or molecules at the surface of a material
experience a different environment to that of the atoms situated in the bulk of the material and present
boundary properties that can be amplified in any interaction involving these nanosystems as the ratio of
atoms on the surface to atoms in the bulk becomes greater [18].
Over the last twenty years, several nanomaterials have been developed, some of them being already
industrially produced and commercially available (i.e. sunscreen, cosmetics, food packaging, clothing,
disinfectants and fuel catalyst [3]). Figure 1 shows the evolution of the “nanomarket” growth, depicting
the number of “manufacturer–identified, nanotechnology–enabled products” inventoried by the official
US Consumer Product Safety Commission and the Project on Emerging Nanotechnology [3].
1000
800
600
400
200
0
2005
2006
2007
2008
2009
Figure 1 Growth of manufacturer-identified, nanotechnology products listed on Project on Emerging
Nanotechnologies Consumer Products Inventory from 2005 to 2009 (in grey) showing products under possible
Consumer Product Safety Commission jurisdiction (blue) [3].
2.2. Nanoparticles (NPs): Nanoscience Building Blocks
Nanoparticles (NPs) represent an attractive category of nano-scaled materials, therefore they have been
intensively studied over the past few years.
They can be considered as zero-dimensional nanostructures. Several types of NP systems (i.e. metal,
metal-oxide, or semiconductor colloids and nanocrystals) have been prepared and studied [19]. They are
becoming an important group of nanomaterials due to the simplicity of their synthesis [11, 20, 21] and
their unique size- and shape-dependent [8, 22-24], electronic [25], optical [19, 26, 27] and catalytic [7]
properties. Moreover, chemical compositions and dimensions (~2 – 100 nm) [21, 28] makes them of
comparable size as biomolecules and biomolecular assemblies (e.g. proteins, nucleic acids) which makes
them well suited to investigate the biological interactions at the molecular -scale.
17 of 94
2.2.1. Gold Nanoparticles
In the field of nanosciences, a considerable effort was focused on the study of gold nanoparticles which
0
are made of the aggregation of a few metallic gold atoms (Au ), ranging from one to a few hundred
nanometers. To be stable in solution, these aggregates are generally surrounded by a protective layer
that can be a polymer, an organic or biologic al molecule, which prevents further aggregation or
coagulation between particles.
2.2.1.1. Historical Background on Gold Nanoparticles
th
Gold particles have been widely studied and used since the 5 century B.C for coloring glass and to cure
illness, first appearing in China, Egypt and in the Roman Empire. One of the most well -known
demonstrations that gold NP exclusive properties were popular is their insertion into the famous
th
Lycurgus cup (Figure 2) in the 4 century B.C.. This cup has a dichroic effect that makes it shine red in
transmitted light and green in reflected light [29].
The therapeutical effects of gold colloids were described for the first time in “panacea aurea auro
portabile” by Francisci Antonii in 1618[30]. Later, in 1676, the german chemist Johann Kunckels reported
the use of “drinkable gold” in his book “Nuetliche Observationes oder Anmerkungen von Auro und
Argento Potabili ” in which he states that “drinkable gold containing metallic gold in a neutral, slightly
pink solution exert curative properties for several diseases”[31, 32].
A more complete review on gold colloids was published in 1718 by Hans Heinrich Helcher, who
remarkably found the need for gold colloids to be stabilized with boiled starch [32, 33].
Figure 2 The Lycurgus Cup in (a) reflected and (b) transmitted light. Department of Prehistory and Europe, The
British Museum. Reproduced from [29].
However, it was not until the 19
th
century that research on this nano-scaled material has evolved
tremendously with the pioneering work carried by Michael Faraday, who by using a two phase synthesis
-
protocol (reduction of a chloroaurate (AuCl 4 ) with phosphorous in CS2 ) concluded that the ruby-red
color of certain tainted glass was a result of the presence of small gold colloids [34, 35] (see Figure 3).
18 of 94
Figure 3 Faraday’s colloidal ruby gold. Reproduced from [34]
Then, in 1994, Brust et al. showed that combining Faraday’s two-phase colloid synthesis with the known
self-assembly of thiol molecules on gold surfaces creates neutral small gold NPs relatively monodisperse
in size (~3.4 nm) coated with an alkanethiolate monolayer [20]. The resulting particles can easily be
isolated from and redissolved repeatedly in solvents without experiencing irreversible aggregation or
decomposition and are stable in atmospheric conditions.
2.2.1.2. Stabilization of Gold Nanoparticle Dispersions
The chemical stabilization of particles is essential to prevent degradation processes such as oxidation or
undesired sintering of NPs and to avoid agglomeration. One of the key aspects about colloidal chemistry
has to do with the means used to stabilize the particle suspensions in the medium in which they are
dispersed [28, 36].
NPs dispersion behavior is essentially dependent on the Van der Waals attraction and on the Brownian
motion [36]. Van der Waals forces alone are only signi ficant for short inter-particle distances, however
when combined with the Brownian motion, which ensures the continuous collision of particles in the
medium, these two forces lead to irreversible aggregation. This aggregation can be controlled and
stopped in the presence of repulsive forces capable of counteracting these attractive forces [28]. This
can be attained or by electrostatic stabilization which creates a distribution of charged species in the
system and/or by steric stabilization which involves the adsorption of molecules or m acromolecules
onto the particle surfaces.
2.2.1.3. Properties and Characteristics of Gold Nanoparticles.
Gold NPs are distinct from many other types of NPs because gold possesses some practical and
distinguishing advantages. First and foremost it is a reasonably inert material. Gold does not undergo
oxidation at temperatures below its melting point, it does not react with atmos pheric O2 and with most
chemicals. These properties make possible to handle and control it under atmospheric conditions [10].
Gold is biocompatible, and binds thiols with high affinity, which is determinant for the formation of self assembled monolayers (SAMs) (vd. Section 2.4 – Self-Assembled Monolayers (SAMs)). Furthermore, gold
nanoparticles are amenable to “mixed monolayer” coverages , where different ligands can be affixed to
the particle surface in well-defined ratios [37].
19 of 94
Another reason for the common use of gold for the synthesis of nanoparticles is the well-defined
synthetic methods for their fabrication and manipulation. Indeed, they can be quickly synthesized by
several ways: either in aqueous [38-41] or organic solvent [20, 42] and following mono- [19, 42, 43] or
biphasic methods [20].
2.2.1.4. Surface Plasmon Resonance
Small metallic NPs exhibit strong optical absorption at one particular fre quency due to a collective
oscillation of the free electron gas in the NP [44]. This frequency known as surface plasmon resonance
frequency is a result of the propagation of electromagnetic waves along the surface of a conductor [45,
46]. When the dimensions of the conductor are reduced, boundary and surface e ffects become very
important [47] and since the wave is on the boundary of the metallic surface and the external medium,
the resonance frequency becomes extremely sensitive to any change on this boundary. This frequency
can be tuned by varying the composition of the NP core [48] and the dielectric constant of the
surrounding material which is dependent on the solvent and molecules adsorption to the metal surface
[46, 49]. This sensitivity of the plasmon resonance to the environment has driven the use of NPs as
biological and chemical sensors [15, 50].
2.3. Monolayer-Protected Metal Nanoparticles
Monolayer-Protected Metal Nanoparticles (MPMNs) are supramolecular assemblies consisting of a
nanoscale, crystalline, metallic core surrounded by an outer ligand shell, a sel f-assembled monolayer
(SAM see below) composed of thiol-containing molecules bond to the surface through a sulphur-metal
bond [11, 51]. These particles exhibit numerous useful and unique properties many of which arise and
are adjusted by the close spatial contact between the core and the shell. These particles characteristics
are conferred by their metallic core (e.g. surface plasmon absorption) [8], their SAM (e.g. solubility and
sensing) [11], and to both of the components (e.g. single electron transistor) [9] and can be easily
synthesized [52]. They have been recently exploited for many applications in all most of science fields
ranging from material science [53], to medicine [54], biology [55] , physics [56] and chemistry [57].
2.4. Self-assembled Monolayers (SAMs)
Understanding the relationship between the nanostructure of a material and its macroscopic properties
has always been a major goal for interfacial science [50]. Self-assembled monolayers (SAMs) constitute a
particularly good way to explore these interactions as they are structurally well defined and offer the
prospect of creating organic interfaces that can be tailored for different mechanistic studies.
SAMs are ordered monomolecular assemblies that adsorb spontaneously on the metal substrate due to
a surface energy minimization of the metal NP (a planar surface or highly curved nanostructure) [10, 58].
20 of 94
These molecules consist of a “head-group” which shows a special affinity for a substrate and, in the
other end of the molecule, a “terminal -group” a tail with a functional group that controls the surface
properties of the SAM (Figure 4) [59]. SAM adsorption on gold NPs involves the initial physisorption of
the molecules on the metal and the subsequent chemisorption of the “head -groups”. This strong
-1
covalent sulphur-gold (S-Au) bond (typically ~50 kcal mol ) is stable over a large range of temperatures,
solvents and pH [10, 60, 61]. Once adsorbed on the metal lattice, by losing the mercaptan’s proton, H,
SAM molecules can adopt energetically more favourable conformations, which allow high degrees of
van der Waals interactions (and in s ome cases hydrogen bonding) [62, 63] staying tightly packed. SAMs
composition can be easily and deliberately altered, for example, functional groups of the assembled
ligands can be tailored to exhibit for insta nce hydrophobic (e.g. methyl groups) or hydrophilic (e.g.
hydroxyl) ends [64].
It is also noteworthy that since the thiol molecules efficiently smooth the faceted and highly anisotropic
surface energy of the gold particles, once wrapped with this thiolated monolayer gold NPs acquire a
round and smooth shape [65, 66].
Figure 4 Representative diagram of an SAM of a lkanethiolates supported on a gold NP. Light grey circles represent
chemisorbed “head-groups”, which are thiol groups; dark grey circles represent the “terminal-groups”, which can
be a variety of chemical functionalities. Adapted from [10]
This protecting monolayer plays a critical role, controlling and ruling all the particles’ interactions with
the outside molecular environment and providing them with a long series of properties, such as stability
21 of 94
against aggregation into the thermodynamically preferable bulk state, solubility in many solvents,
assembly properties and sensing of specific biomolecules [51]. However, a detailed understanding of the
dynamics of the assembly still lacks of a complete and proper characterizatio n [67].
2.4.1. Phase-Separated Ordered Domains
SAMs composed of a mixture of ligands on flat surfaces have long been known to phase-separate into
randomly sized and shaped domains [68] or into worm-like structures [48] as observed by scanning
tunnelling microscopy (STM) [69-71].
Phase-separation also occurs on NPs. However the structure of SAMs on NPs is complicated by the fact
that the two-dimensional monolayer (the SAM) must be adsorbed onto a three-dimensional structure
(the NP). This additional complication leads to a unique situation of phase-separation into organized
(rippled) domains.
2.4.1.1. Experimental Observations
In 2004, Jackson et al. first confirmed the presence of ordered phase-separated domains in the ligand
shell of MPMNs. They found that those SAMs of alkanethiol ligands with varying tail groups phase
separate into stripes or ripples that encircle or spiral around the me tal nanoparticle (see Figures 5 and
6) [52]. The width of those ribbon-like domains was found to be as small as < 1 nm (often no more than
two molecules) and their presence was first confirmed by STM [52] and later using Fourier transform
infrared spectroscopy (FTIR) [72]. With FTIR, due to the perturbed intermolecular forces noticeable in
phase-separated domains an upward shift in the CH 2 symmetric stretching frequency was observed
confirming the presence of phase-separation. Unfortunately, those results could not completely prove
that the phase-separation was ordered.
Figure 5 a) STM image of MPMNs showing phase-separated ordered doma ins on b) STM image of a sing le gold
nanoparticle, arrows indicate ripple spacing. c) Schematic depiction helping to v isualise the arrangement of
molecules on the NP surface where the ra ised yellow regions symbolize the octanethiol molecules and the red ones
depict mercaptopropionic acid. Reproduced from [52]. Scale bar: 10 nm
22 of 94
Figure 6 Surface plot of the ligand shell presenting the contours of the ripples (scales in nm). The distance between
the two arrows shows the ripple spacing. Reproduced from [52].
The same group have also shown that changing the stoichiometric ratio of the ligands during synthesis
affects the SAM shell morphology, ranging from discrete phase -separated domains to highly ordered
ripples (see Figure 6). This phenomenon of ordered phase separation was then confirmed to form for a
wide variety of binary ligand shell compositions, including molecules with variable chain lengths and
molecules with different end groups and backbone structures (e.g. aliphatic vs. aromatic).
Figure 7 Three dimensional rendering of gold NPs STM height images. (a) (b) Ordered phase-separated domains (2:1
molar ratio of decanethiol/mercaptopropionic acid) a nd the schematic dra wing respectively, (c) (d) Discrete packed
phase-separated domains of the less abundant component (10:1 molar ratio of OT/ mercaptopropionic acid) and
the schematic drawing repectively. Reproduced from [52].
In 2007, Glotzer et al. carried out atomistic and mesoscale simulations in order to assess the origin of
the experimentally reported stripe formation [73].They predicted that ligands nanophase-separation
into striped patterns on NPs surfaces will depend on a balance between the enthalpic losses and
entropic gains [73].
23 of 94
As presented on Figure 8, phase-separation contributes to an increase in conformational entropy.
Basically, the longer or bulkier surfactant tails can occupy more space when surrounded by shorter or
less bulky molecules. When this entropy gain overcomes (a) the energy reduction that would be
observed in the case of bulk separation and (b) the energetic penalty of generating additional interfaces,
then nanophase-separated stripes will form. In this situation, the domain width of the ripples will
increase with the ligand chain length [73].
Figure 8 Schematic representation of the free volume (indicated by the shaded cones) available to ligands on curved
surfaces, and respective simulated cross-sectiona l v iews. (a) indicates ligands with s imilar lengths; (b) shows ligands
with cons iderable differences in lengths. The free volume and the consequent ga in in conf igurationa l entropy for
ligands with different lengths is clearly larger than that for ligands with the same length. Adapted from [73]
In addition, simulations predicted bulk phase separation for mixtures of short enough surfactants (e.g.
three carbon chains), or for ligands with small bulkiness difference. Bulk phase -separation was also
observed in the case of extremely high degrees of curvature (very small NPs) (see Figure 9 (a)). The
spheres, if small when compared to the surfactants length will not gain significant entropy by generating
extra interfaces (and therefore ripples) as the tails already possess enough conformational entropy by
moving radially outward in the sphere [73-75]. Increasing the NP radius results in ordered stri pes as
mentioned above (Figure 9 (b)). In larger spheres (Figure 9 (c)), disordered stripes and irregular domains
will form as predicted by atomistic simulations. Extending the radius to infinite (flat surfaces) the SAM
will form wormlike stripes for the referred tail length ratio - 4:7 (Figure 9 (d)) [73].
Figure 9 Equilibrium arrangements of binary SAMs adsorbed on NPs with different degrees of curvature determined
through mesoscale s imulations. Yellow and red lobes symbolize head termina ls of longer and shorter ligands,
respectively. Sphere radii: (a) 3 σ – sma ll NPs: binary mixtures of surfactants separate into two bulk phases, (b) 5 σ –
increas ing NP radius: phase-separation into ordered stripes occurs, (c) 10 σ - further increase in NP radius:
disordered stripes and patchy domains, (d) ∞ - f lat surface: same as (c). Sphere radius not in sca le. Reproduced
from [73]
24 of 94
Then, in 2008, Centrone et al. confirmed that when ordered phase-separated domains (ripples) are
present, the solubility of MPMNs in ethanol shows an unexpected behaviour, and does not increase
monotonically with the concentration of hydrophilic ligand [52, 76]. In this case, the longer of the two
ligands determines the solubility. On the other hand, as should be expected, when the particles have no
ripples the concentration of the hydrophilic ligand determines the solubility. These findings prove that
ligand shell morphology influences the solubility of these NPs almost as much as the chemical a nd
molecular composition of the SAM [76].
At the same time, in a combined STM and TEM study, Hu et al. showed that when ripples are present in
the ligand shell, they enhance the particle-particle interactions, leading to a stronger degree of
interdigitation when compared to their homoligand counterparts and are considerably less able to form
ordered supracrystals [77].
2.4.1.2. Properties of rippled MPMNs
This unprecedented molecularly defined arrangement imparts a variety of singular properties to a NP
system such as non-monotonic dependence of solubility [52, 78] on ligand shell composition and
demonstrated good resistance to protein nonspecific adsorption [78]. The latter occurs for NPs coated
with a mixture of hydrophobic- and hydrophilic-ligands. Once formed, the inter-distance between the
hydrophilic and hydrophobic domains in the ligand shell will be too small to allow the protein to find a
suitable conformation to adsorbe onto the NP [79]. This is the functional basis behind the dolphin skin
resistance to biofouling [80].
2.4.2. Place-Exchange Reactions
The development of versatile strategies to functionalize alkanethiolate gold MPMNs is vital in the
development of these materials as potential chemical reagents and catalysts [20]. In this context, ligand
place-exchange reactions are a key step in opening up MPMNs functionalization [11, 67].
MPMNs with alkanethiolate monolayers (R-S) can be changed by exposing them to a solution of another
kind of molecule (R’-S). This type of reaction is schematized below:
where x and n represent the numbers of new and former ligands, respectively.
Although the microscopic details remain unclear, reports of ligand exchange on two-dimensional-SAMs
on gold surfaces often show that many sites exchange extremely slowly or not at all, whereas others are
relatively reactive [81-84]. Similarly for gold MPMNs Hostetler et al. found that exchange occurs
25 of 94
preferentially at minority sites such as defects and pinholes (e.g. terrace edges, vertexes) as well as grain
boundaries and that terrace-bond ligands are both less reactive and, at best, only slightly mobile (see
Figure 10). These assumptions suggest the existence of a hierarchy of the different core surface binding
sites with associated susceptibility to place exchange [67].
The same group also indicated two different exchange regimes to explain such a site preference: (a) the
incoming ligand enters the monolayer in order to undertake place -exchange. This associative
mechanism would favour less crowded, nonterrace sites [67]; or (b) ligands will be desorbed
(preferentially at defect sites) followed by attachment of new thiol to the newly created surface vacancy
[85, 86]. Also important to mention is that these easily exchanged sites are not a static population (as
confirmed by a serial exchange experiments [87]) and are liable to migration within the ligand shell . (see
Figure 10) [67].
The same study also revealed that the MPMN monolayer only possess a limited number of ligands that
are bond weakly enough to be lost as disulfides. The kinetic re sults demonstrate that the rate of ligand
exchange on gold MPMNs depends on the concentration of the entering and exiting ligands, a rate that
is initially rapid but slows dramatically. Longer entering ligands and chain lengths in the protecting
monolayer will both decrease the rate of ligand exchange. Study of ligand place-exchange dynamics and
mechanism show that exchange has a 1:1 stoichiometry, which means that one molecule is adsorbed for
each molecule desorbed [67].
Figure 10 Representative illustration of the processes involved in ligand place-exchange reactions on gold MPMNs.
(a) Exchange of vertex thiolates (1) with solution thiol; (b) Exchange of edge and near-edge thiolates (2) with
solution thiol; (c) Exchange of terrace thiolates (3) with solution thiol; (d) surface migration among vertice and edge
thiolates; (e) Surface migration within edge (and near-edge) and terrace thiolates; (f) Surface migration within the
terrace thiolates. Adapted from [67].
26 of 94
2.5. MPMNs Assembly
To take advantage of the useful optical and electronic properties of MPMNs it is of big interest to be
able to predict and control the assembly of NPs into specific structures since this would expand the
range of potential applications of these particles [88]. In order to use NPs as nano blocks to build
complex devices, their positions relative to each other and to their environment must be precisely
controllable. Therefore, extensive research is evolving in order to try to organize MPMNs in a
predictable and ordered way.
NPs are inherently isotropic and therefore will not spontaneously form into stable anisotropic one
dimensional assemblies without experiencing some external driving force [89, 90]. Thus breaking the
symmetry of the inter-NP interactions represents one of the key challenges in nanomaterials research
[91]. In fact, efforts to control the assembly of MPMNs, based mainly on biomolecules [92-94] and other
templating molecules [95], have been delayed due to a lack of control in the number of receptors
interacting with the templating agent. Herein some examples of chemically directed assembly will be
reviewed.
2.5.1. Polar Defects
SAMs on flat gold surfaces form a two-dimensional crystal in which each ligand can be represented by a
vector corresponding to its projection in the surface normal (determined by the tilt angle) [10, 96]. In
the case of the SAM on a NP core, it is necessary to consider the assembly of a vectorial order
(projection of SAM ligands) onto a topological sphere [97, 98], which inevitably requires the formation
of two diametrically opposed defect points on the NP. This is consistent with the known “Hairy Ball
Theorem”, which states that it is impossible to arrange a vector field onto a sphere without the
formation of at least two diametrically opposed singularities. This theorem also explains quotidian
phenomena like the whirl present on our hair and also the existence of at least one cyclone in the
atmosphere at any given time [99].
Considering the last statements, the ordered domains present in the rippled MPMNs result in two
profoundly demarcated polar singularities [91]. Molecules in the poles, being not entirely stabilized by
intermolecular interactions with their neighbours , manifest themselves as highly reactive defect points
(Figure 11) and consequently are more vulnerable to be displaced by place-exchange reactions.
27 of 94
Figure 11 Schematic illustration of a rippled MPMN showing one of the two diametrically opposed polar defects
that need to exist to enable the alternation of concentric rings. This defect is highly reactive due to a lack of
intermolecular stabilizing forces. Reproduced from [91].
Stellacci and co-workers used these unstable singularities to functionalize MPMNs at two diametrically
opposed points [99] in order to form directional and controlled chains of NPs [91]. They positioned
carboxylic acid-terminated molecules at these unstable poles to create divalent NPs and then reacted
them with a diamine linker generating the linear chains of NPs. TEM was performed to assess and
characterize the chains of pole-functionalized rippled MPMNs (see Figure 12). The specificity of the
place-exchange reaction was demonstrated since a small number of branched chains and threedimensional aggregates were observed.
Figure 12 TEM images of MPMN cha ins. Chains were formed after adding a cross-linker to the solution. Most of the
chains do not have branches or 3D structures which is consistent with the fact that the two polar defects are the
most reactive points of the particles. Scale bars 200 nm, inset: left 25 nm; right 50 nm. Reproduced from [91].
They have also shown the ability to vary the distance between each particle in the linear chain by
altering the length of the linking diamine molecules [91].
In a subsequent study, Carney et al. demonstrated that the formation of linear chains occurs only with
MPMNs within a determined size range (in the referred case from approximately 2.5 to 8.0 nm) which,
28 of 94
by extension means that the ordered striped organization of the assembled monolayer only exists in
that specified size range as it was previously predicted by simulation (section 2.4.1.2) [73, 75].
Later on, in a UV-Vis study Vanessa LaPointe also found that in the case of rippled MPMNs (1:1 MH/OT)
a clear change in the absorbance spectrum was visible only 5 minutes after starting the reaction with
the linker molecules (1,9-nonanedithiol) and appeared to be finished approximately at the end of 20
minutes. However, in the 5:1 MH/OT series, which does not have these diametrically opposed and
highly reactive defects, the cross-linking reaction and the subsequent aggregation took much longer. It
took more than one hour before the reaction appeared to have ended and the aggregation level was
lower.
2.5.2. Dithiols
The simplest way of inducing controlled MPMNs aggregation is to use bi -dentate thiol ligands that cross link the gold particles together as a result of the strong sulphur-gold interaction. The common feature of
these materials is that they are completely insoluble because of the high degree of three-dimensional
cross-linking [56, 100, 101]. Brust group explored this method and demonstrated that the central
property behind the structures of the generated assemblies is the number of linking molecules per gold
NP. Adding 1,9-nonanedithiol molecules to the tetraoctylammonium bromide-stabilized gold NPs
(within a defined range of gold-dithiol molar ratios) assembled the particles together because thiols
form a stronger bond in the gold surface than the bromide molecules [102]. The resultant threedimensional ball-type assemblies precipitate at both high and low linker ratios, however at an
intermediate ratio (~60 - 14 000 ligands per MPMN) they remain dissolved in toluene. Curiously, the
round aggregates line up and generate relatively straight lines when ethanol is mixed with the toluene
solution (Figure 13) [102].
Figure 13 TEM images of linear hierarchical assemblies of MPMNs formed upon addition of ethanol to the toluene
solution. Reproduced from [102].
29 of 94
2.5.3. Other types of assembly
There is a wide range of methods to organize gold NPs besides the specific examples explained above.
Using the known covalent and strong non-covalent interactions between the ligands attached to the
NPs, various nanostructures, including mono- and multilayers, as well as composites with oligomers or
conducting polymers have been engineered [7, 103, 104]. Gold NPs have also been used as multivalent
cores to build dendritic structures [7] but the isotropic character of this type of binding represents a
problem for the bottom up fabrication of more complex nanoparticle -based structures.
Other groups studied monofunctionalized NPs bound to multidentate molecules with set up direc tional
interactions which permits a controlled assembl y. The required single valency in the NPs is prepared via
reaction with a functional thiol linked to a solid support [105-107]. For example, using the extreme
specificity of the interaction between DNA strands used as ligands, cyclic, linear, and discrete branched
structures of monofunctionalized gold NPs have been obtained [108, 109]. However, the introduced
single valency renders NPs spectator pendant groups, ra ther than building blocks for these nanoengineering challenges.
Another strategy to achieve one-dimensional aggregates of NPs is the use of pre -existing one
dimensional structures such as nanotubes, as templates onto which NP arrangments can be formed.
The one-dimensional assembly of nanorods is also viable since the ends of these structures are made of
a different material in relation to the center, therefore these ends can be functionalized with molecules
with reactive end groups and generate linear as semblies.
30 of 94
3. Thesis Outline
The possibility to understand, control and predict particles assembly has become a very attractive
research area since directional and isotropic assembly of this nanoscale building blocks could impart
upon them properties that enable their use for diverse ends such as sensing and electronic devices.
During the last decade a large effort was put into the study of gold NPs assembly. However the recent
finding of ordered phase-separated domains, ripples, and the consequent properties this morphology
confers (non-monotonic dependence of solubility and protein resistance properties) provided a new tool
for the study of these particles’ interactions within the solvents [76] and their neighbouring MPMNs
[91].
In this context and based on the work developed by Stellacci’s group (see Section 2.5.1 – Polar Defects)
the aim of this thesis is focused on the assembly study of gold NPs coated with an organic layer
assembly in the presence of a cross-linker in solution. This monolayer is composed by the same
molecules (6-mercapto-1-hexanol (MH) and 1-octanethiol (OT)) but in different ratios (as illustrated on
Figure 14). 2:1, 1:1 and 1:2 MH/OT present ordered phase -separated domains, ripples, in their ligand
shell while 5:1 and 1:5 MH/OT present unordered domains.
Figure 14 Schematics of MPMN ligand shell chemistry and morphology. Ratios indicate stoichiometric ratios of
thiolated ligands (6-mercapto-1-hexanol and 1-octanethiol) during synthesis. (Schematics courtesy of of Dr. Steve
Mwenifumbo
The present project was divided in two main stages. First, seven different types of gold MPMNs were
synthesized and characterized. This was attained by direct observation (the formation of a brownish
powder is related with the presence of smaller particles), solubility testing in EtOH (0:1 M H/OT was
1
tested in toluene), UV-Vis absorption spectroscopy, H NMR spectroscopy and TEM analysis for size
distribution. An adequate characterization of gold nanoparticles is preponderant for the future use of
the same. Gold NPs characterization was developed along with Miss Nia Bell and Miss Vanessa LaPointe.
The second part of this project was focused on the study of these MPMNs assembly. The assemblies
were studied by UV-Vis spectroscopy to determine the rate of assembly for each type of MPMNs and
TEM to observe the morphology of the aggregates. The state of aggregation observ ed on the TEM
images was used to determine the parameters that were used in the subsequent TEM analysis, such as
cross-linker molar concentration, time and MPMN concentration.
31 of 94
4. Materials and Methods
All reagents were purchased from Sigma-Aldrich Inc., UK (Dorset, UK) and used as received without
further purification, unless otherwise specified.
Solvents and acids such as absolute ethanol (EtOH), acetone, isopropanol, diethyl ether, (Et 2 O),
dichlorometane (DCM), toluene, benzene, hexane, nitric acid (HNO 3 ) and hydrochloric acid (HCl) were
purchased from VWR, UK (Lutterworth, UK)
Carbon coated Copper TEM grids (300 mesh) were obtained from Agar Scientific.
TEM Images were obtained using a JEOL 2010. The images were analyzed using NIH Image Software
ImageJ. UV-Vis absorbance measurements were acquired using a Perkin-Elmer spectrometer.
4.1. Characterization Techniques for the nanoscale
Nanostructures are “inconveniently small” [110] – too small to be observed and studied directly. For this
reason a “spy” versatile enough to report on a wide range of molecules and capable of relaying the
information on the structures, motions and chemical reactions of these systems without significantly
altering those properties is needed [110].
These nanostructures chemical and physical properties are mostly governed by their composition, size,
structure, shape and surface properties, thus their precise characterization is essential to ass ess these
parameters and understand their effect on the intrinsic properties of the materials. For these purposes ,
a series of analytical tools have been developed. Examples like electron microscopy [111], atomic force
microscopy (AFM) [112], scanning probe microscopy (STM) [113] are used for structural analysis of these
nanostructures. Spectroscopic techniques, such as UV-Vis absorption spectroscopy or Raman
spectroscopy are used to obtain information on the optical properties of the material [114-116]. The
different techniques used to characterize the nanoparticles mentioned in this thesis are described
below.
4.1.1. UV-Visible spectroscopy
A UV-Visible (UV-Vis) spectrophotometer is used to determine the absorption or transmission in both
the UV and visible wavelength [20]. Absorption of photons results in the promotion of an electron from
an orbital of a molecule in the ground state to an unoccupied orbital at a higher energy level (see Figure
15).
32 of 94
Figure 15 Schematic depicting the promotion of an electron from an orbital of a molecule in the ground state (π) to
an unoccupied orbital at a higher energy level (π*)
In general, UV-Vis spectroscopy is used to study how a sample responds to light as it measures the
attenuation of a beam of light after it passes through a sample (Figure 16). Ultraviolet and visible light
are energetic enough to promote outer electrons to higher energy levels, and UV/Vis is usually applied
to molecules or inorganic complexes in solution [117].
Figure 16 Light passing through the cuvette containing the sample.
A UV-Vis spectrophotometer consists of a radiation source which uses an incandescent bulb for visible
wavelengths and a deuterium lamp in the ultra -violet wavelengths, a sample holder, a monochromator
or diffraction grating (which enables the selection of a narrow band of wavelengths), a photodetector to
measure the intensity of light which is transmitted through the sample, and an output device [117].
When the beam passes through a sample, some light gets absorbed, while some continues through the
sample to the transmitter. The ratio of the intensity of light beam entering the sample (I 0 ) and coming
out (I t) at a specific wavelength is defined as transmittance (T).
Absorbance (A) is the negative logarithm of T
33 of 94
The Beer-Lambert law states that for a given ideal solution, there is a linear relationship between
concentration and absorbance provi ded that the path length is kept constant. The extinction coefficient
(ε) is a constant for each molecule for each wavelength.
where ε, an intrinsic property of the species, represents the extinction coefficient of the substance (L
-1
-1
-1
mol cm ), c is concentration of absorbing species (mol L ), and l is the absorption path length (cm)
through the sample. Therefore, provided that ε and l are kept constant, there is a linear relationship
between concentration and absorbance.
Gold NPs, specifically, exhibit distinct colors (and thus SPR bands) that are a characteristic of particles
nature, shape, size or assembly state on the refractive index of the surrounding medium and on the
nature of their protective layer [114, 118] The SPR peak intensity and broadness is dependent on the
NPs solution concentration (and extinction coefficient) and its size dispersion respectively. [114, 118]
For instance, particles smaller than 3 nm in diameter do not show any SPR peak.
4.1.2. Nuclear Magnetic Resonance (NMR) Spectroscopy
Nuclear magnetic resonance is a property that magnetic nuclei have in a magnetic field, which cause the
nuclei to absorb energy from the electromagnetic pulse and radiate this energy back out at a specific
resonance frequency which, among other factors, depends on the strength of the magnetic field.
NMR Spectroscopy is a powerful technique that can provide useful and detailed information on the
topology, dynamics and structure of molecules and biomolecules in solution. This spectroscopic
technique relies on the magnetic properties of the atomic nucleus and can be used to investigate the
quantum mechanical properties of molecules. When placed in a strong magnetic field, certain nuclei
1
(e.g., H,
13
C,
15
N – the ones with an even number of electrons ) resonate at a characteristic frequency
(characteristic of the isotope and electronic/electromagnetic environment ) in the radio frequency range
of the electromagnetic spectrum. Slight variations i n this resonant frequency give detailed information
about the molecular structure in which the atom resides, providing us structural information about the
global system [119]. NMR always provides “local” information; i. e. the world is studied from the
perspective of a single atom in a molecule where this atom can only “s ee” 5 Å (approximately three
bonds away). However this perspective can be tuned so we could “see” the world from each constituent
atom in the molecule. The NMR data consists of a series of relationships between the atoms of the
molecule, where the intensities of the signals are directly proportional to the concentration. With the
right information about these relationships we can construct an unambiguous model of the molecular
structure [119].
34 of 94
4.1.3. Transmission Electron Microscopy
Transmission Electron Microscopes today are capable of achieving a point to point resolution of better
than 0.1 nm. In fact, a major attraction to the early developers of the TEM was that, since electrons are
smaller than atoms, it would be possible, at least theoretically, to build a microscope capable of
“seeing” details below the atomic level [120].
In this context, transmission electron microscopy (TEM) is a powerful imaging technique used to
determine the size and shape of materials at the nanometer length scale that uses, in essence, the same
working principles of the light microscopy technique, both based on the foc us of an electron or a light
beam through magnetic or optical lenses, respectively. The resolution of the microscopy, r, is defined
by:
where λ represents the wavelength of the incident beam, µ is the medium refractive index and α stands
for the semi-angle of the microscope aperture [121, 122]. Since the wavelength of electrons is about ten
thousand times smaller than the photons wavelength, theoretically the TEM resolution will be
approximately ten thousand ti mes smaller than an optical microscope resolution, enabling the
observation of nanometer scale objects [121].
Figure 17 shows the components of a conventional TEM. The electron gun provides an intense beam of
high energy electrons that pass through an ultra thin specimen (10 to 100 nm), interacting with it. An
image is formed from the interaction of t he electrons transmitted through the specimen. The electron
beam can be generated either by thermionic discharge or by field emission and TEMs use an
accelerating voltage between 100kV to 400 kV. To avoid scattering of electrons by air, TEMs are
operated in vacuum. The first condenser lens is used to create a de -magnified image of the gun
crossover and to control the minimum spot size available in the rest of the condenser system. The
second condenser lens affects the convergence of the beam at the specimen and the diameter of the
illuminated area of the sample. The condenser aperture controls the fraction of the electron beam
which is allowed to hit the specimen. The objective lens then forms an inverted initial image of the
sample. The objective aperture is used to select the electrons which will contribute to the image i.e. to
control the contrast of the initial image. The project lens is used to magnify the initial image formed by
the objective lens to a desired magnification for viewing. A viewing screen is an integral part of any
electron microscope. It can be coated with materials such as ZnS, which translates electron intensity to
light intensity, which we observe and record on a layer of photographic film or on a CCD camera [123].
35 of 94
Figure 17 Components of a transmission electron microscope (TEM). Adapted from [123]
4.2. NP synthesis
Seven types of MH and OT coated gold NP were synthesized following a previously published [43] onephase method and the general procedure was as described below.
All the glassware was previously cleaned with aqua regia (3:1 HCl to HNO3 ), deionized water, acetone,
toluene, acetone, dried with N2 gas and finally rinsed with the respective reaction solvent
(dichloromethane for all particles except the 0:1 MH/OT which was rinsed using benzene).
In a 300 ml round bottom flask, 80 ml of the reaction solvent was stirred in a water bath held at 55°C in
which 496 mg (~1 mmol ) of AuPPh3 Cl was dissolved. Once totall y dissolved (10 min), the thiol -containing
ligands were added (see Table 1) and the solution was then stirred for 5 minutes before adding 870 mg
(~10 mmol) of the reducing agent, in this case a borane tert-butylamine complex. The reaction vessel
was then left for one hour and the condenser system was turned on.
After one hour the solution was allowed to cool at room temperature. The particles were precipitated
overnight at room temperature in 80 ml of Et 2 O. The supernatant was removed and the particles were
collected and washed in Et 2 O. This purification process was repeated until particles were fully
precipitated and the solution was clear. The remaining supernatant was then removed. The particles
were dried in air, weighed and stored for future use.
36 of 94
Table 1 Amounts of ligands used in the synthesis of NPs with various compositions
Volume (µl) 1
Ratio
1
(MH/OT)
MH
OT
1:0
273
0
5:1
227
58
2:1
181
116
1:1
136
174
1:2
91
231
1:5
45
290
0:1
0
347
A 1:2 ligand to gold salt molar ratio was used in
the synthesis of the MH/OT coated particles.
Surfactants are added to the reaction vessel during nanoparticle formation in order to control the rate
of growth and limit aggregation [43]. The adsorption of surfactant-like molecules to nucleated
nanocrystals lowers the free energy of the surface and, therefore, the reactivity of the particles. The
ratio of surfactant to metal precursor can control the size distribution of the particles.
The use of amineborane complexes is essential for the syntheses of monodisperse metallic
nanoparticles because compared to other reducing agents such as sodium borohydride (NaBH4 ) and
lithium borohydride (LiBH4 ), amineborane complexes have a weaker reducing ability, which can slow the
reducing rate of gold cations and allow control over the growth of nanoparticles [43].
4.3. NP Characterization
In order to characterize the size distribution and composition of the nanoparticles a series of techniques
were applied: UV-Vis spectroscopy, transmission electron microscopy and nuclear magnetic resonance
spectroscopy respectively. Besides the transmission electron microscopy (TEM) imaging, all
characterization was performed in solution and thus was an ensemble average of all the present
structures.
4.3.1. UV-Visible absorption spectroscopy
UV-Vis spectroscopy measurements were carried out to rapidly estimate the NPs size by ensuring that
all the MPMNs had similar surface plasmon resonance (SPR) peaks .
Measurements were acquired using a PerkinElmer Lambda25 UV-Vis spectrophotometer operating at
-1
room temperature and at a scanning rate of 240 nm.min from 350 to 900 nm with 1 nm resolution.
37 of 94
UV-Vis spectra were prepared in disposable poly(methyl methacrylate) (PMMA) cells for the particles
dissolved in EtOH or a 1 cm path-length quartz cuvette for 0:1 MH/OT NPs dissolved in toluene. NPs
solutions were prepared by dissolving 3 mg of nanoparticles in 12 ml of absolute EtOH or toluene for the
OT homo NPs, sonicating for 20 min and stirring for 2 or more days before performing the UV-Vis
spectroscopy. The UV-Vis samples were prepared by diluting 500 µl of the NP solution in 500 µl of EtOH
(or toluene in the case of 0:1 MH/OT particles).
4.3.2. Transmission Electron Microscopy (TEM)
Characterization of particle size and size distribution was carried out using transmission electron
microscopy (TEM). 7 µl of 0.125 mg/ml nanoparticle solution were pipetted onto a 300 mesh carboncoated copper TEM grid, wicked with a KimWipe and allowed to dry in air. Microscopy was performed
on a JEOL 2010 (JEOL I) operating at an accelerating voltage of 200 kV. NP size distributions were
obtained by analyzing a minimum of 300 parti cles from several TEM images using ImageJ software and
were plotted as % of Frequency vs Size.
4.3.3. Nuclear Magnetic Resonance (NMR) Spectroscopy
1
H NMR spectroscopy was performed on the NP solutions to determine the purity of the solution and
whether unbound ligands were present. The binding leads to a broadening of the peaks in the NMR
spectrum associated to the ligands (MH – R-CH2 OH at 3.4 ppm; OT – R-CH3 at 0.70 ppm). Because the
analysis must be performed in deuterated solvents, the product must be carefully dried before this
analysis in order to prevent product loss [37]. The sample tubes were cleaned following the same
glassware procedure described for the MPMNs synthesis except in the final where they were rinsed with
δ3 CD3 OD (deuterated methanol), then 10 mg of NPs were dissolved in 0.6 ml of CD 3 OD. The NMR was
performed on a Varian 500 MHz NMR (Brucker).
4.4. NP Assembly
To characterize the assembly process two different techniques were used. To assess the state of
assembly based on the optical properties of the nanostructures UV-Vis analysis was performed. To
observe the structural morphology of these assembled structures , TEM imaging was carried out.
38 of 94
4.4.1. UV-Visible absorption spectroscopy
To assess the time of aggregation, UV-Vis spectroscopy measurements were carried out. 0.5 ml of a 0.25
mg/ml NPs solution were mixed with 0.5 ml of different cross-linker solutions: 1, 5 and 10 mM. The
samples were left to run in the spectrophotometer taking UV-Vis spectra over 1 hour period. Each
sample was mixed at the end of each 10 min. A marked change in the absorbance spectrum is indicative
of aggregation, and the time they take to stabilize after adding the li nker will be taking into account.
4.4.2. Transmission Electron Microscopy (TEM)
To image the gold nanoparticles assemblies with the addition of the cross -linker, a 7 µl drop with the
gold nanoparticles solution mixed with the cross -linker solution was pippetted onto the TEM grid. Some
parameters were varied during the experiments like the gold nanoparticles solution concentration (0.25
g/l, 0.05 g/l and 0.025 g/l), cross-linker concentration (10, 5, 1, 0.1 and 0.01 mM), time (0, 1, 5, 10, 1
hour, 12 hour, 24 hour time) and also cross-linker type molecules. A more detailed description is
provided in the next session.
39 of 94
5. Results and Discussion
5.1.
Synthesis and Analysis of Monolayer-Protected Metal Nanoparticles
Seven types of MPMNs were synthesized using hydrophobic OT and hydrophilic MH as protective
ligands (see Figure 18). These capping agents were chosen due to their strong binding affinity to gold
surfaces because of the presence of a pending thiol group and their small head group size, which
prevents charged electrostatic interactions associated with large terminal groups of other typically used
ligands.
Figure 18 Chemica l structures of a) 1-octanethiol (OT) the hydrophobic ligand and b) 6- mercapto-1-hexanol (MH)
the hydrophilic ligand. MH/OT carbon chain ratio 6:8
The method, described in section 4.2, allowed the synthesis of nanoparticles coated with a homogenous
or heterogeneous SAM. The term homogeneous layers hereby refers to MPMNs functionalized with only
one type of thiol containing molecule (MH or OT), and heterogeneous particles to MPMNs coated with
various ratios of MH and OT. The terminology that will be used throughout this work is as follow: x:y
MH/OT, with x and y representing the molar ratios of MH and OT used during the synthesis of the
particles (i.e. 1:1 MH/OT implies 1:1 molar ratio of the MH and OT ligands; 1:0 MH/OT and 0:1 MH/OT
MPMNs represent gold NPs entirely coated with MH and OT molecules respectively). Seven series of
MH/OT MPMNs have been synthesized (1:0, 5:1, 2:1, 1:1, 1:2, 1:5 and 0:1 MH/OT respectively) and to
ensure some reproducibility, three different batch of each MPMNs synthesis have been produced.
By controlling the reaction temperature (55°C) and keeping the reagent ratios constant ( i.e. 1:2 ligand to
gold salt and 10:1 reducing agent to gold salt molar ratios) the synthesis produced MH /OT MPMNs with
relatively narrow size distributions.
Figures 19 and 20 present typical TEM images of the seven different MH/ OT MPMNs sets. Histograms of
the respective core size distributions are depicted in Figures 21 and 22. They provide critical information
about the size dispersion of the samples. Average MH/OT MPMNs series sizes ranged from 4.2 ± 0.9 to
4.9 ± 0.8 nm (see Table 2). Core sizes and size distribution were obtained by measuring at least 300
particles from several TEM images for each of the MPMN sets.
40 of 94
Figure 19 Representative TEM images obta ined for the three batch of four different MH/OT MPMNs synthesized
showing the size of the gold core of the different particles. Scale bars - 20 nm.
41 of 94
Figure 20 Representative TEM images obta ined for the three batch of three different MH/OT MPMNs synthesized
showing the size of the gold core of the different particles. Scale bars - 20 nm.
42 of 94
Figure 21 Histograms obtained for the three batch of f ive different MH/OT MPMNs synthesized depicting core size
distributions. Each MPMN system exhibit relatively monodisperse s ize distributions. NP core s izes were obta ined by
analyzing at least 300 particles from different TEM images for each of the MPMN systems.
43 of 94
Figure 22 Histograms obtained for the three batch of two different MH/OT MPMNs synthes ized depicting core size
distributions. Each MPMN system exhibit relatively monodisperse s ize distributions. NP core s izes were obta ined by
analyzing at least 300 particles from different TEM images for each of the MPMN systems.
Table 2 Table of average core size ± Standard deviation for MH/OT MPMN series:
MPMN
Core size (nm)
1:0 MH/OT
4.6 ± 0.8
5:1 MH/OT
4.7 ± 0.8
2:1 MH/OT
4.9 ± 0.9
1:1 MH/OT
4.4 ± 0.7
1:2 MH/OT
4.6 ± 0.8
1:5 MH/OT
4.2 ± 0.9
0:1 MH/OT
4.9 ± 0.8
Global average diameter
4.6 ± 0.8
Overall, the synthesis produced a relatively high yield of particles which ranged from 97.3 to 309.2 mg
which corresponds to efficiencies between 19.5 and 61.8% Table 3 shows the yield and efficiency data
for each of the MH/OT MPMN series.
44 of 94
Table 3 Total yield and efficiency of MPMNs synthesis.
Nanoparticles
1:0 MH/OT
5:1 MH/OT
2:1 MH/OT
1:1 MH/OT
1:2 MH/OT
1:5 MH/OT
0:1 MH/OT
Yield (mg)
Efficiency (%)
Batch A
271.5
54.3
Batch B
282.9
56.6
Batch C
309.2
61.8
Batch A
279.5
55.9
Batch B
258.4
51.7
Batch C
226.6
45.3
Batch A
124.6
24.9
Batch B
207.3
41.5
Batch C
153.5
30.7
Batch A
203.8
40.8
Batch B
175.8
35.2
Batch C
159.1
31.8
Batch A
118.1
23.6
Batch B
121.2
24.2
Batch C
166.9
33.4
Batch A
156.2
31.2
Batch B
206.3
41.3
Batch C
97.3
19.5
Batch A
185.8
37.2
Batch B
160.5
32.1
Batch C
166.0
33.2
2:1 MH/OT batch A were the first MPMNs synthesized which justifies the low yield obtained. Also a
decrease in the yield is noticeable for the particles with a higher ratio in the hydrophobic (OT) ligand.
This could be due to a loss of particles during the washing steps of the synthesis protocol or as a
consequence of OT solubility in EtOH.
UV-Vis absorption spectroscopy was used in the characterization of MPMNs. These particles SPR band
(and thus their colour in solution) is characteristic of their nature, size and shape, and also depends on
the refractive index of the surrounding medium and on the nature of their protective layer. Typically,
particles smaller than ca. 2 nm do not show any plasmon peak at all [7]. Thus, UV-Vis spectroscopy of
the prepared MPMN suspensions was carried out; the results are depicted in Figure 23. Surface plasmon
peaks are fairly similar with a maximum absorbance ranging from 504.2 to 514.0 nm. Due to limited
solubility of the different MPMN seri es, UV/Vis measurements were performed after at least a 2 days
settling period to enable slow floculation of the insoluble NPs, leaving in solution only the colloidaly
stable ones. As explained in section 2.4.1, the solubility of mixed ligand shell MPMNs (mixtures of
hydrophilic (MH) and hydrophobic (OT) ligands) does not increase linearly when increasing the fraction
of hydrophilic ligand in the ligand shell in a hydrophilic solvent such as EtOH. Since the intensity of the
45 of 94
peak of the plasmon band is directly proportional to the particles concentration and can be related to
the critical colloidal solubility, MPMNs solubility in EtOH was ranked as follows: 2:1 MH/OT (100%); 1:2
MH/OT (95 %); 1:5 MH/OT (81 %); 1:1 MH/OT (71 %); 1:0 MH/OT (56 %); 5:1 MH/OT (53 %). The relative
solubilities reported here have been normalized to highest intensity of the plasmon peak values: 2:1
MH/OT – 1.391. (0:1 MH/OT MPMNs were prepared in toluene since they are not soluble in EtOH so
they have not been considered for these c alculations). Since all the NPs are roughly the same size the
variations in plasmon intensity at the given concentration have not been considered.
Figure 23 UV-Vis absorption spectra of MPMNs suspended in EtOH (or toluene in the case of the OT Homo NPs) at
an initia l concentration of 0.125 mg/ml. For the MPMNs dissolved in EtOH, solubility follows a non-monotonic trend
with composition.
Table 4 Plasmon peak values and intensities for each of the MH/OT MPMNs series.
1
MPMN
Plasmon peak
Plasmon peak intensity
1:0 MH/OT
507.0 ± 3.0
0.81 ± 0.03
5:1 MH/OT
510.1 ± 3.2
0.77 ± 0.07
2:1 MH/OT
512.9 ± 5.1
1.46 ± 0.24
1:1 MH/OT
507.9 ± 4.6
1.03 ± 0.16
1:2 MH/OT
508.9 ± 6.7
1.39 ± 0.26
1:5 MH/OT
504.2 ± 5.6
1.19 ± 0.09
0:1 MH/OT
514.0 ± 2.0
0.20 ± 0.07
H NMR analysis confirmed the presence of unbound ligands, denoted by the presence of a sharp peak
in the NMR spectrum associated with the proton from the pending thiol of the ligand (MH – R-CH2 OH at
3.4 ppm) however the absence of a peak at 0.70 ppm confirms the absence of OT in solution [61]. This
excess of thiols in solution will have repercussions in the MPMNs stability in solution as discussed in
below (see section 5.3.2.5). Figure 24 shows a representative spectrum and respective analysis obtained
46 of 94
for one of the sets of MPMNs (2:1 MH/OT Batch B). Measurements were done for all MPMNs,
respective spectra and analysis can be seen in Appendix .
Figure 24 1 H NMR spectrum obta ined f or 2:1 MH/OT Batch B in CD 3 OD. The presence of a peak in the NMR
spectrum associated to the the hydrophilic ligand (MH at 3.4 ppm) shows the presence of free ligands in solution.
In order to achieve information on the structure of the ligand shells, Amplitude modulation – Atomic
Force Microscopy (AM – AFM) (performed and analysed by Dr. Kislon Voitchovsky at MIT) was
performed on the 2:1 MH: OT Batch A film. When high resolution was possible (see Figure 25) MPMNs
appeared normal and rough which citing Prof. Stellacci means that the few isolated particles present in
the film are indeed striped. A more detailed analysis was not possible since the films were co mposed of
~30-40 nm NPs aggregates which make it difficult to achieve a high imaging quality mandatory for a
proper analysis of the ligand shell stripes and their inter -distance.
Figure 25 Film of MH:OT 2:1 nanoparticles imaged by AM -AFM in ultrapure water. Right: XY: 40 nm, Z: 11 nm; Left:
Scale bar: 5 nm
47 of 94
AM-AFM was not performed for the other MPMNs series however morphological differences are
expected between homogeneous and heterogeneous-ligand particles shells. Pure MH and OT NPs were
assumed to have homogeneous hydrophilic and hydrophobic ligand shells. The 1:2, 1:1 and 2:1 MH/OT
particles displayed classical ripples which are attributed to hydrophilic/hydrophobic striated ligand shell s
(schematics of ligand shell structure are presented on Figure 14, section 3). The 1:5 and 5:1 MH/OT
MPMN were assumed to have homogeneous ligand structures, interspersed with discrete domains of
the minority ligand.
5.2.
MPMNs Molarity
In order to determine the best gold NPs and cross -linker concentrations to start the MPMNs assembly
study and vary these parameters in a controlled way, the particles molarity has to be determined. The
next section shows how we have assessed our MPMNs and cross -linker solutions molarity.
5.2.1. Number of Gold Atoms:
The gold atoms that form the gold NPs, are strongly and tightly packed, with each of the constituent
atom being surrounded by twelve neighbouring atoms [7]. Therefore, the smallest cluster has thirteen
2
atoms and the following contain 10 n + 2 atoms, where n represents the layer number. The increase of
the total number of atoms with the number of layers is represented in figure 26 [7].
4500
n of layer atoms
4000
3500
total n of gold atoms
3000
2500
2000
1500
1000
500
0
1
2
3
4
5
6
7
8
9
10
11
n of layers
Figure 26 Graph showing the relation between the number of layers of a gold NP and the tota l number of gold
atoms.
48 of 94
A facetted NP is considered a topological sphere [7] thus, in theory, the total number of gold atoms, NAu ,
existent in a NP can be calculated knowing the average diameter, d, of the core. Of course, the presence
2
of defects will allow NPs with an intermediate number of atoms to exist (i.e. not following the 10 n + 2
distribution).
Where VAu is the gold NP volume.
As specified on table 2, the average diameter (d) for all the considered NPs is 4.6 nm, We should note
that in the TEM images we can only see the gold core shell, so the ligand shell will not be considered.
The grafting density of thiols (Nthiols ) bound to the NP surface depends on the surface area and t he space
occupied per each thiol on it (see Eq. 7)
Where the surface area can be calculated if we have the average diameter value (see Eq. 8) and the
2
space occupied per thiol in the surface of the NP which is known to be 22 Å [124].
Table 5 synthesises the results obtained:
Table 5 Estimated gold clusters characteristic dimensions.
d (nm)
4.6
3
VAu (nm )
0.51
NAu
2970
Surface Area (Å2 )
Nthiols
6647.6
302
Once estimated the total number of gold atoms (NAu), 2970, it was possible to estimate the molecular
weight of the five different sets of MPMNs. Then, considering the final volume and concentration of 12
ml and 0.25 mg/ml respectively, the total number of MPMNs in solution can be determined. The results
are presented in the table below:
49 of 94
Table 6 Linear formula, molecular weight, molar concentration a nd average number of nanoparticles per gold
cluster (NN P s ) for each set of MPMNs.
MPMNs
Linear Formula
Molecular
Molar concentration
Weight (g/mol)
(M)
5:1 MH/OT Au2 9 7 0(HS(CH2 )6OH) 25 1,7(CH3 (CH2 )6CH2 SH) 50,3
6.26E+05
2:1 MH/OT Au2 9 7 0(HS(CH2 )6OH) 20 1,3(CH3 (CH2 )6CH2 SH)100,7
6.27E+05
1:1 MH/OT Au2 9 7 0(HS(CH2 )6 OH) 1 5 1 (CH3 (CH2 ) 6 CH2 SH) 1 5 1
6.27E+05
1:2 MH/OT Au2 9 7 0(HS(CH2 )6OH) 10 0,7(CH3 (CH2 )6CH2 SH)201,3
6.28E+05
1:5 MH/OT Au2 9 7 0(HS(CH2 )6OH) 50 ,3(CH3 (CH2 )6CH2 SH) 251,7
6.29E+05
Average
3.99E-07
NNPs
2.88E+15
6.27E+05
Taking in consideration our TEM grid, it is a fair estimate to consider that of the 7 µl MPMNs solution
drop only 1 µl (0.125 mg/ml) stays in the carbon grid, and that at a 60 K magnification there is an
average of 300 particles per TEM image.
Knowing this and the details provided in the following table one can d etermine the average number of
MPMNs per TEM grid following two different ways: a) considering the molar concentration, the volume
that stays in the TEM grid (1 µl) and the Avogadro number; and b) the average number of particles per
image; Thus the values can be compared in order to verify the consistency of the approximations used
(a) molar concentration and final volume on the grid and b) number of particles per image).
Table 7 TEM grid and TEM image considerations.
LTEM image (nm)
2
150
ATEM image (m )
2.25E-14
dTEM grid (mm)
3
2
ATEM grid (m )
7.07E-6
NP/ATEM image
~300
1
NP/ATEM grid
9.5E+10
NP/ATEM grid2
1.2E+11
1
2
Considering an average of 300 particles per TEM image
Considering that only 1 µl (0.125 mg/ml) stays in the TEM grid
Where L TEM image is the square side of the TEM image, ATEM image is the observable area of the TEM image,
dTEM grid and ATEM grid represent the carbon grid’s total diameter and area respectively. NP/A TEM image and
NP/A TEM grid correspond to the total number of nanoparticles per TEM image area and TEM grid area
respectively.
Both values are in the same order of magnitude which indicates the approximation taken is valid and
significant.
50 of 94
Next, the ratio of cross -linker molecules used was determined. Two types of dithiol molecules (1,9nonanedithiol (NDT) and 1,16-hexadecanedithiol (HDDT)) were used at different stages of the work.
Their structure and characteristics are presented in the table below.
Table 8 1,9-nonanedithiol and 1,16-hexadecanedithiol chemical structure and molecular weight
1,9 – nonanedithiol
HS(CH2 ) 9 SH
MW (g/mol)
192.4
1,16 – hexadecanedithiol
HSCH2 (CH2 ) 1 4 CH2 SH
MW (g/mol)
290.6
Considering a 0.5 ml volume of cross -linker solution and the different molar concentrations used C (C=
xM, yM, ...) the total number of mols (N) and the number of cross -linker molecules (Ncross-linker) can be
calculated in the respective 0.5 ml solutions. The results are summarized in table 9.
Table 9 Number of mols (N) and number of cross-linker molecules (Ncross-linker) for different molar concentrations
considering a final volume of 0.5 ml of cross-linker solution.
C (mM)
N (mol)
Ncross-linker
10
5.0E-06
3.0E+18
5
2.5E-06
1.5E+18
1
5.0E-07
3.0E+17
0.1
5.0E-08
3.0E+16
0.01
5.0E-09
3.0E+15
During the TEM analysis two different concentrations (c) of MPMN solution were used (0.25 and 0.05
mg/ml). Considering a 0.5 ml volume for the NPs solution, the molar concentration ( C), the number of
mols (N) the total number of NPs and thiols (NNanoparticles and Nthiols respectively) in solution can be
determined for each of the MPMN concentration:
Table 10 Molar concentration (C), number of mols (N) and number of NPs and thiols (NNanoparticles and Nthiols
respectively) correspondent to two different MPMNs final concentrations, considering a final MPMNs solution
volume of 0.5 ml.
c (mg/ml)
C (M)
N (mol)
N Nanoparticles
Nthiols
0.25
4.0E-07
2.0E-10
1.2E+14
3.6E+16
0.05
7.97E-08
4.0E-11
2.4E+13
7.2E+15
51 of 94
Finally, knowing the total number of cross -linker molecules present in solution as well as the total
number of NPs and number of thiols in the ligand shell available to place exchange we can determine
the ratio of cross-linker molecules per a) each gold NP and b) each thiol bound to the gold NP surface:
Table 11 Cross-linker to gold NP ratio and cross-linker to thiol ratio for five different cross-linker molar
concentrations.
Ccross-linker
5.3.
Cross-linker to gold NP ratio
Cross-linker to thiol ratio
(mM)
0.25 mg/ml
0.05 mg/ml
0.25 mg/ml
0.05 mg/ml
10
2.50E+04
1.25E+05
8.33E+01
4.17E+02
5
1.25E+04
6.25E+04
4.17E+01
2.08E+02
1
2.50E+03
1.25E+04
8.33E+00
4.17E+01
0.1
2.50E+02
1.25E+03
8.33E-01
4.17E+00
0.01
2.50E+01
1.25E+02
8.33E-02
4.17E-01
Cross-linking of MPMNs in solution
5.3.1. UV-Visible Analysis
Since it is impossible to know exactly the number of NPs in solution (the molecular weight calculations
represent an estimation based on the TEM imaging) achieving the right cross-linker to gold NPs ratio is
very challenging. Having this in consideration for a first experimentation of the cross -linker effect, three
solutions with different molar concentrations of the linker molecules, 1,9 – nonanedithiol (NDT) were
added to all, but 1:0 and 0:1 MH/OT, NPs series and aggregation was observed for all types of MPMNs.
Soon after adding the cross -linker solution an insoluble black precipitate began to form and after one to
two hours the EtOH solution had lost most of its colour and the precipitation process reached
completion. These precipitates are indicative of big insoluble NPs agglomerates formation.
The extent and the kinetics of these processes can be studied with UV-Vis spectroscopy and are
reported on Figures 27 and 28 where the change of absorbance for the aggregation in the 450 -900 nm
region is shown.
Figure 27 shows the changes observed in the UV-Vis spectra when different concentrations of dithiol
were added.
52 of 94
Figure 27 Change of absorbance in the 450-900 nm reg ion of UV-Vis spectra for 1:1 MH/OT Batch A MPMN solution
upon addition of (a) 1 mM, (b) 5 mM and (c) 10 mM solution of cross-linker respectively. A dithiol alkane cha in was
added to the s olution of MPMNs causing the particles to link together as a place exchange reaction was taking
place. The legend represents a timecourse from 0 to 60 minutes after a ddition of cross -linker, curves were taken at
6 minutes intervals.
Following the cross-linker addition, the intensity of the SP band starts to decrease and a broadening of
the SPR band is observed. Furthermore the SP band maximum also slightly shifts to longer wavelengths
which is associated wi th the formation of NP clusters. However UV-Vis results are inconclusive in
determining whether this shift is caused by the formation of defined structures or non-specific
aggregation, a proper analysis is only achievable with the TEM (see section 5.3.2)
Figure 28 compares the different behaviours between the remaining particl es (5:1, 2:1, 1:2 and 1:5
MH/OT Batch A, respectively) upon addition of the cross-linker solution (5 mM). (The UV-Vis
measurements and analysis were repeated for all batches and sets of MPMNs for the three different
concentrations of cross-linker although here only Batch A is represented and for a representative
concentration of cross -linker, 5 mM)
53 of 94
Figure 28 Change of a bsorbance in the 450-900 nm reg ion of UV/Vis spectra for (a) 5:1, (b) 2:1, (c) 1:2 and (d) 1:5
MH/OT Batch A MPMNs upon addition of 5 mM of 1,9-nonanedithiol solution respectively. A dithiol a lkane cha in
was added to the solution of MPMNs caus ing the particles to link together as a place exchange reaction was taking
place. The legend represents a timecourse from 0 to 60 minutes after a ddition of cross -linker, the curves were
taken at 6 minutes intervals. The arrows point to the SPR peak, 0 and 60 minutes after adding the cross-linker
molecules and in a) and d) a smooth appearance of a second peak is also signed.
All graphs show MPMNs aggregate with time, however no clear difference between the three
concentrations of 1,9-nonanedithiol tested could be observed. On the other hand each set of these
particles shows a different behaviour.
2:1, and 1:2 MH/OT exhibit a straighter peak at 0 min (Figure 28, b) and c)) while 5:1 and 1:5 MH/OT
show a broader peak (Figure 28, a) and d)). For 2:1 and 1:2 MH/ OT particles one can only denote a
decrease in the peak intensity whereas along with that 5:1 and 1:5 MH/OT particles also show the arise
of a flatter curve. In fact for the 1:5 MH/OT particles at the end of one hour the plasmon peak almost
completely disappeared. It is also clear there is a shift of the SP band maximum to higher wavelengths
in the case of 5:1 MH/OT particles. It is also noteworthy the smooth appearance of a second peak in the
case of 5:1 and 1:5 MH/OT particles (Figure 28 a) and d)). These results meet the established theoretical
descriptions of Mie scattering for aggregates. This theory suggests that if the SPR absorption peak is still
present in the absorption spectrum of the cross -linked solution, small aggregate clusters of similar sizes
54 of 94
are probably forming. On the other hand, if the plasmon band completely disappears and a flatter
spectrum forms, aggregates are probably bigger and have a broad size distribution [125, 126].
The SPR peak variations with time for each set of MPMNs upon the addition of a cross -linker solution
are illustrated in Figure 29. A gradual increase of the SPR maximum value with increasing MH
concentrations is observed with this trend being clearer for the 5:1 MH/OT set.
Figure 29 SPR peak shifts for all the batches of the 5 sets of MPMNs tested at different times. Dark blue line: SPR
peak of MPMNs in EtOH; Medium and light blue lines: SPR peak of MPMNs solution measured immediately and 30
minutes after adding the 1,9-nonanedithiol solution, respectively; a) 1 mM b) 5 mM and c) 10 mM 1,9nonanedithiol concentration. The Figure clearly demonstrates a dependence on the ligand shell morphology. When
the ratio of hydrophilic ligand increases, more the SPR band shifts.
The advancement of the aggregation process was also followed with the naked eye as the solution
gradually changed from a light pink-red colour to a browner one, and finally, upon standing, to a dark
precipitate.
Since the differences between the five distinct MPMN solutions were no t too evident (as shown in
Figures 27 and 28 (a-d)) we opted to measure the stabilization time (Figure 30), which corresponds to
the time the difference between the curves at 800 nm takes to stabilize, i.e. reach zero. The stabilization
time indicates that the place exchange reaction rate is slowing down and the reaction is reaching the
equilibrium state.
55 of 94
Figure 30 Stabilization time, i.e. time, the difference between the curves at 800 nm along time, takes to stabilize
(reach zero), reaching the equilibrium.
Once again we can verify that the stabilization time is also dependent on the ligand shell composition
and morphology. Increasing MH concentration, the hydrophilic ligand, in the monolayer shell
composition appears to result in slower the stabilization time. This trend can arguably be explained
considering that the hydrophilic character of the ligand confers more stability in solution, prolonging the
place-exchange reaction. However, ligand place-exchange reactions are dependent on many
parameters, such as the ligands relative affinities to gold surface, their size and their solubility. Thus, the
detailed mechanisms of the ligand place exchange could not be fully rationalized.
5.3.2. TEM Analysis
After studying the optical properties of the different MPMNs assembl ies, their morphology was
examined by TEM. During the TEM analysis, several approaches have been used. TEM data is hereby
presented according to the different variables studied, i.e. colloidal suspension’s stability, time, crosslinker concentration and cross-linker type.
5.3.2.1. Hypothesis
TEM analysis was based on several assumptions based on the different behaviours and self-assembly
driving forces for each set of MPMNs.
DeVries et al. previously reported that when both the ligands are present in the same ratio in the ligand
shell, the resulting SAM will phase-separate into ordered (rippled) domains and therefore will present
fewer stable molecules in the poles. These ligands being not optimally stabilized by in termolecular
interactions will be the first molecules to be replaced in place -exchange reactions. Thus after
56 of 94
functionalizing these specific points, divalent NPs can, in theory, be generated and linear chains ranging
from three to twenty MPMNs in length wil l form (Figure 31) [91].
Figure 31 Observed assembly of the 1:1 MH/OT gold MPMNs. The unstable polar defects will be place-exchanged
with the dithiol molecules in solution originating linear assemblies. TEM image reproduced from [91].Scale bar: 50
nm.
In the case of 1:2 and 2:1 MH/OT the polar singularities will react faster than other defects that might
exist in the ligand shell. However these diametrically opposed polar defects are in this case more
stabilized by intermolecular interactions with their neighbouring counterparts which results in still linear
but smaller chains with occasional branched structures (see figure 32)
Figure 32 Predicted structures of the assembled 2:1 and 1:2 MH/OT gold MPMNs, respectively. The existence of less
reactive poles will be the driving force for the formation of linear but a few branched structures. Scale bar: 20 nm
The remaining sets of particles, 5:1 and 1:5 MH/OT gold MPMNs supposed to have an almosthomogenous ligand structure with discrete domains of the minority component are expected to form
unordered three-dimensional assemblies (Figure 33) since the linker molecules will place-exchange
randomly or at the occasional defect and pinhole sites that might exist anywhere in the ligand shell.
57 of 94
Figure 33 Expected structures of the assembled 5:1 and 1:5 MH/OT gold MPMNs, respectively. Instead of ripples,
the lesser abundant ligand will form discrete domains in the monolayer. There are no polar defects in these types of
ligand arrangements so the cross-linker molecules will bond to the meta llic surface randomly or at the occasional
defects that might exist so unordered and three-dimensional assemblies are expected. Scale bar: 10 nm
Considering their ligands ratio, 2:1 and 5:1 MH/OT series of MPMNs present a SAM with less amount of
OT, which is the longer molecule. According to Hostetler et al. studies, the rate of ligand exchange
decreases when the chain length of the protecting monolayer increases, because, according to the
previously explained associative mechanism it will be more difficult for the incoming molecules (dithiol)
to penetrate the monolayer and undertake place -exchange [67]. In this case the more abundant ligand
in the SAM is the MH (the smaller molecule in length) which favors the place exchange with the
incoming dithiol resulting in a quicker process and bigger structures.
5.3.2.2. TEM Sample preparation
Imaging of self-assembled NPs can be challenging, thus, TEM sample preparation is established
according to several parameters:
1.
If the NPs solution is too concentrated, the density of NPs in the grids will be too high and will
make it difficult to discern whether the structures formed were in fact a result of the cross linker action or formed by interparticle interactions upon drying.
2.
If the NPs solution is allowed to sit for too long on the TEM grid, it will be possible for the
MPMNs in solution to arrange or re-arrange themselves into ordered or random structures in
the TEM grid.
3.
The solvent used must be pure and miscible to avoid artifacts that are a result from the
nanoscale bubbles of phase separation (see figure 39 section 5.3.2.3.1).
For these reasons the solution was cast onto a carbon coater copper mesh grid and placed on a clinical
TM
tissue (KimWipe ) in order to rapidly wick away the excess of solvent and place the MPMNs onto the
58 of 94
TEM grid in the same arrangement as they were in solution, preventing the formation of unspecific
aggregation patterns that might artificially cause size selection and affect the size distribution results.
5.3.2.3. Stability of particles in solution
5.3.2.3.1.
Control experiments
Throughout the project several control experiments under different conditions have been tested. The
control experiments were made in order to find out the agglomeration state of the nanoparticles in
solution along time without the presence of cross -linker molecules. This was determinant to conduct the
work and to follow the approaches taken during this project.
Therefore, TEM grids were prepared with NP solutions in the absence of cross-linker as controls.
Representative images are shown below:
Figure 34 TEM images of 2:1 MH/OT batch A MPMNs solution (0.125 mg/ml). Scale bars: 100 nm
A general view over the TEM grid confirmed the presence of several small assemblies everywhere on the
grid. This was the first indication that, so far, suggests these particles are not stable in solution and are
spontaneously self-assembling.
In an attempt to eliminate the spont aneously formed small aggregates of NPs, s olutions were then
sonicated for 1 hour. The samples were then left to stir for a period of 24 hours, to let the eventual
persisting agglomerates to sediment. However, no significant improvement could be observed on the
TEM pictures, and small aggregates were still visible (Figure 35).
59 of 94
Figure 35 TEM images of 2:1 MH/OT batch A MPMNs solution (0.125 mg/ml). Grid prepared after sonicating the
sample for 1 hour period. Scale bars: 100 nm
Based on previous results, fresh solutions of nanoparticles were prepared. They were sonicated for two
hours and stirred for at least two days. Then, in order to effectively eliminate or at least reduce the
number and size of the spontaneous agglomerates, the solution was purified by filtration-centrifugation
using a syringe filter (ANATOP – 0.1 µm pore size at 15 000 rpm for 8 minutes).
Fresh solutions of 2:1, 1:1 and 5:1 MH/OT Batch B were thus diluted (1:10 to 0.025 mg/mL), filtered and
centrifuged. The TEM grids were prepared 2 hours after the purification for the first two types of
MPMNs (2:1 and 1:1 MH/OT). In the case of the 5:1 MH/OT MPMNs the solution was left standing for 3
days and the TEM grids were prepared at the end of the third day after the purification steps.
A general look over the TEM grid of the first two sets of NPs revealed that there were no aggregates in
solution (see figure 36). However samples analysed three days after purification a lready started to show
signs of aggregation, once again suggesting that the synthesized NPs were spontaneously selfassembling in solution with their neighbouring particles (figure 37). Nevertheless the different SAM
shells of the particles should also be considered as a reason for the spontaneous self-assembly of the 5:1
MH/OT molecules.
Figure 36 TEM images of (a) 2:1 and (b) 1:1 MH/OT batch B MPMNs solutions (0.025 mg/ml). Grids prepared two
hours after filtering and centrifuging the MPMNs fresh solutions. Scale bars: 50 nm.
60 of 94
Figure 37 TEM pictures of the 5:1 MH/OT batch B MPMNs solution (0.025 mg/ml). Grids prepared 3 days after the
filtration and centrifugation steps. Scale bars: 50 nm
In order to study the time-dependence of the aggregation process, a diluted batch of 1:1 MH/OT coated
particles (1:5 – 0.05 mg/ml) were filtered and centrifuged at different times (see table 12).
Table 12 Different amounts of time samples were left stirring after the purification steps and prior to TEM grid
preparation. Each figure letter corresponds to a figure, presented below, and represents the images obtained for
the different solutions of 1:1 MH/OT.
figure
Settling time
38
1 hour
39
5 hours
40
Left - 3 days
Right - 14 days
41
5 days
61 of 94
Figure 38 TEM pictures of the 1:1 MH/OT Batch B MPMNs solution f iltered and centrifuged 1 hour before the TEM
grid preparation. Upper images: 0.125 mg/ml; down images: 0.05 mg/ml. Scale bars: 50 nm.
Figure 39 TEM pictures of the 1:1 MH/OT Batch B MPMNs solution (0.125 mg/ml) filtrated and centrifuged 5 hours
before the TEM grid preparation. Scale bars: 50 nm.
62 of 94
Figure 40 TEM pictures of the 1:1 MH/OT Batch B MPMNs solution (0.05 mg/ml) f iltered a nd centrifuged 3 and 14
days before the TEM grid preparation, left and right images, respectively. Scale bars: 50 nm.
Figure 41 TEM images of the 1:1 MH/OT Batch B MPMNs solution (0.05 mg/ml) f iltered and 5 days before the TEM
grid preparation. Scale bars: 50 nm.
TEM grids prepared hours after the purification steps present a considerable number of small linear
assemblies (figure 38), which is interesting considering that these are the rippled particles. After three
days two and small three-dimensional structures could be observed (figure 39) however on the grids
prepared using “older” solutions no defined structure could be imaged. Those results are not consistent
with the previous results (figures 34 to 37). Working with different types of MPMNs that have different
ligand shell morphologies can make them have occasionally unexpected behaviours towards solvents. As
an example, the nanoscale rings highlighted in figure 39 could be explained as (a) a result of holes
opening up in the liquid film and pushing particles into their borders [127], (b) a result of water droplets
which condense on the surface of nonpolar solvents from humid air [128] or (c) impurities in the
solvent, such as other liquids droplets.
Several groups have focused on the study of the induced and spontaneous assembly of nanoparticles in
solution. Sidhaye et. al found that the size of the three dimensional lattices spontaneously formed in
solution depend on the chain length of the alkanethiol molecules that wrap around the NPs, i.e. when
the chain length of the ligands that cap the NP metal core increases, the interparticle attractive forces
decrease and so decreases the size and overlapping of the structures. [129]. The thiol-containing
63 of 94
molecules used in the present project are relatively short which can explain the formation of these big
structures.
Sidhaye et.al also proved that the formation of three-dimensional structures was only possible in the
presence of excess thiol molecules in solution, which are in dynamic equilibrium with those
chemisorbed onto the NP surface. Our NMR analysis revealed the presence of unreacted MH in solution
which can explain the instability of MPMNs in solution [67, 129].
5.3.2.3.2.
UV-Vis study
To test and confirm the MPMNs stability in solution over time, a solution of NPs was prepared as
described previously, sonicated for 20 minutes and left to stand for a period of twelve weeks. SPR peak
evolution was recorded periodically by UV-Vis spectrophotometry during this time interval. The results
are summarized in the graph presented below:
Figure 42 UV-Vis absorption spectra of 1:1 MH/OT Batch B MPMNs dissolved in EtOH at an initial concentration of
0.125 mg/ml. The sample was kept untouched during a period of twelve weeks, and during this time interval several
UV-Vis samples were taken periodically (2, 4, 6, 8 and 12 weeks after, respectively). A decrease in the intensity and
a slight shift of the SPR peak can be observed.
64 of 94
Table 13 Plasmon peak wavelengths and intensity variations with time for MPMNs solution (1:1 MH/OT Batch B –
0.125 mg/ml). This solution was kept untouched for twelve weeks period after a 20 minute - sonication step.
Analysis date
Plasmon
Peak
Plasmon peak
intensity
12th Nov 2009
508
0.88
26 Nov 2009
508
0.85
10th Dec 2009
th
509
0.81
st
510
0.76
th
512
0.66
21 Dec 2009
15 Jan 2010
As shown in Table 13 and Figure 42, there is a considerable decrease of the SPR peak intensities as well
as a slight shift of the SPR peak value, suggesting slow MPMN aggregation as described in section 5.3.1.
When particles aggregate they tend to sediment at the bottom leading to a reduction in the number of
particles in solution and thus a decrease in the peak intensity. This parallel study was important to
confirm the image-based finding that this type of particles were self-assembling in the ethanolic
solution.
5.3.2.3.3.
Solvent Dependence
MPMNs solubility was also studied using other solvents of various hydrophobicity (toluene,
dichloromethane (DCM) and methanol ). None but methanol confers stability to the particles (Figure 43).
All the MPMNs series are soluble in methanol and seem to be more st able than their counterparts in
EtOH in the first hours after solution preparation. However an increase in the number of smaller
aggregates (20-50 nm) in solutions aged two days was observed.
Figure 43 1:1 MH/OT Batch A in Methanol a) 2 and b) 48 hours after solution preparation. a) When the TEM sample
was prepared 2 hours after preparing the solution a homogeneous grid was observed. b) 48 hours after preparing
the MPMNs solution the grid presented a considerable number aggregates approximately 50 nm in size. The TEM
analysis was repeated a lso for the 2:1 and 5:1 MH/OT particles ( images not shown) revealing that both these
particles were stable in methanol. Scale bars: 20 nm
65 of 94
5.3.2.4. Assembly variations with time
In the first approach followed the assembly process was explored and analyzed at different times.
1 hour after adding the cross-linker solution - i=6
Initially TEM grids prepared 1 hour after adding the cross -linker molecules (5 mM solution of NDT in
EtOH) to the MPMN solutions have been analyzed. Representative results are presented in the images
below:
Figure 44 TEM images of 1:1 MH/OT Batch A MPMNs solution (0.125 mg/ml) 1 hour ( i=6) after a dding the cross linker, NDT. Ba ll-type aggregates are evidenced in the images. These ball-type aggregates were further cross-linked
due to a large excess of time and dithiol molecules or because of phase-segregation due to poor solubility. Scale
bars: 50 nm
Figure 45 TEM images of 1:2 MH/OT Batch A MPMNs solution (0.125 mg/ml) 1 hour ( i=6) after a dding the cross linker, NDT. Small ball-type aggregates are evidenced in the images. Scale bars: 50 nm
66 of 94
Figure 46 TEM images of 1:2 MH/OT Batch B MPMNs solution (0.125 mg/ml) 1 hour ( i=6) after adding the cross linker, NDT. Small ball-type aggregates further assembled are shown in the images. Arrows indicate the magnified
zones. Scale bars: 50 nm
Figure 47 TEM images of 5:1 MH/OT Batch A MPMNs solution (0.125 mg/ml) 1 hour after adding the cross-linker,
NDT. Ball-type structures are linked together forming larger structures. Arrows indicate the magnified structures.
Scale bars: 50 nm
TEM images show that one hour after adding the cross -linker solution large assemblies were formed
suggesting that many NPs have dithiol molecules at random sites in the ligand shell. These big structures
appear to have a somewhat chain-like character and i nterestingly they all seem to form large networks
of ball-type assemblies that link together originating chains. This is more evident on figures 44 and 47.
These spherical assemblies may be a result of (a) interparticle hydrogen bonding between the
hydrophilic alcohol termini of the ligands (so it depends on the MH/OT ratio in the ligand shell), (b) they
may be composed of long chains that are more stable when coiled together than when extended in an
all-trans configuration or (c) may be a result of the fully encapsulation of t he gold NPs assemblies by
dithiols. These ball -type aggregates are likely to be further cross -linked because of the large excess of
linker molecules.
Similar results have been achieved by Hussain et. al. In their work they have obtained uniformly sized
spherical assemblies of gold colloids in toluene by adding alkanedithiol solutions in different gold NPs to
dithiol molar ratios [102]. Moreover, they were able to further organize these ball -type assemblies into
relatively linear chains through the addition of EtOH.
1:2 MH/OT Batch A appears to behave differently when the cross -linker is added as no chains or big
assemblies were veri fied one hour after adding the linker solution. I nstead small isolated circular
67 of 94
assemblies were formed (figure 45). This resistant behaviour of the mentioned particles towards selfassembly was also previously predicted by UV-Vis analysis (see Appendix). Batch B of the same particles
was also analysed confirming the presence of linear chains of ball -like aggregates.
Overall, TEM analysis strongly suggests that NDT molecules are efficiently cross-linking the NPs.
However, focusing on the final aim of this project, how the different MPMNs start the assembly process,
it is necessary to observe intermediate structures. In order to try to assess this stage of the assembly
process TEM grids have been prepared one minute after pippeting the cross-linker solution. The 5 mM
solution of NDT was mixed with the 1:1 and 5:1 MH/OT Batch A and 1:2 MH/OT Batch B. Analysis of 1:2
MH/OT TEM grids was repeated two weeks later to check if the NPs stay stable and in the same
arrangement over time on the TEM grids. Representative images are presented below.
1 minute after adding the cross-linker solution - i=0
a)
b)
Figure 48 TEM images of (a) 1:1 A and ( b) 1:2 B MH/OT MPMNs solutions (0.125 mg/ml) one minute ( i=0) after
adding the cross-linker (NDT) solution. Three-dimens iona l structures are still v isible especia lly on f igure (a). Scale
bars: 50 nm
68 of 94
a)
b)
c)
Figure 49 TEM images of (a) 1:2 B (same grid as f igure 48 (b) but ana lysed two weeks after), (b) 2:1 A and (c) 5:1 A
MH/OT MPMNs solutions (0.125 mg/ml) one minute (i=0) after adding the cross-linker (NDT) solution. Threedimensional structures are still visible especially on figures (a) and (b). Scale bars: 50 nm
One minute after adding the cross -linker solution, there were already assemblies and small clusters of
MPMNs present in the TEM grid, except in the case of the 1:2 MH/OT batch B where only occasional two
and one-dimensional structures were seen. The large number of two and three-dimensional aggregates
is probably a result of the high degree of interdigitation which can be caused by the proximity of the
particles due to high concentrations (0.125 mg/ml).
69 of 94
5.3.2.5. Assembling behaviours with cross-linker concentration
Besides the specific chemical reactivity or the cross -linker structure, another important parameter that
determines the way the assembly process develops is the linker and NP relative molar concentrations.
Initially the assembly process was too fast, precluding the view of intermediate structures in the TEM
grids, so the cross-linker to gold NPs molar ratio was reduced. Instead of 12500 NDT/gold NP (5 mM) the
MPMN samples were tested with a less concentrated solution of cross-linker molecules (1 and 0.5 –
2500 and 1250 NDT/gold NP, respectively (see table 14)).
Table 14 Table with the calculated linker to particle and linker to thiol ratio values
Ccross-linker
Cross-linker to gold NP ratio
Cross-linker to thiol ratio
(mM)
0.25 mg/ml
0.25 mg/ml
5
1.25E+04
4.17E+01
1
2.50E+03
8.33E+00
0.5
1.25E+03
4.17E+00
0.1
2.50E+02
8.33E-01
The TEM grids analysed were prepared 1 (i=0), 5 (i=1) and 15 minutes (i=2) after adding the cross -linker
solution.
1 mM 1,9-nonanedithiol solution
Figure 50 TEM images of 1:2 MH/OT batch B MPMNs solution 1 ( i=0) and 5 minutes (i=1) after adding the crosslinker (NDT) solution (1 mM). Scale bars: 50 nm
70 of 94
0.5 mM 1,9-nonanedithiol solution
a)
b)
c)
Figure 51 TEM images of 1:2 MH/OT batch B MPMNs solution a) 1 ( i=0), b) 5 (i=1) and c) 15 minutes ( i=2) after
adding the cross-linker (NDT) solution (0.5 mM). Scale bars: 50 nm
TEM images show that one minute after adding the 1 m M cross -linker solution, mainly isolated MPMNs,
and occasional two-dimensional aggregates were observed. At the end of five minutes, big threedimensional structures (300-3000 nm) were formed but no intermediate arrangements were seen, i.e.
one and two-dimensional structures.
Opposed to what should be expected, TEM micrographs show that one minute after adding the 0.5 mM
cross-linker solution, two- and three-dimensional NPs were present in the sample. Five (i=1) and fifteen
(i=2) minutes after massive three-dimensional aggregates (300-3000 nm) were present in the grid. Ball like arrangements were not evident in these samples.
A different set of particles (2:1 MH/OT batch A) was analysed with the same solutions of cross -linker
plus a less concentrated one (1, 0.5 and 0.1 mM – 2500, 1250 and 250 NDT/gold NP, respectively (see
table 14)). The solutions pippeted onto the TEM were prepared one minute after adding the cross-linker
solutions (i=0). Results obtained before (Figure 52) and after (Figure 53) exposing the solution to
sonication for a period of one hour are shown.
71 of 94
a)
b)
c)
Figure 52 TEM images of 2:1 MH/OT batch A MPMNs 1 minute (i=0) after adding the (a) 1 (b) 0.5 and (c) 0.1 mM
solution of cross-linker (NDT). Scale bars: 50 nm
a)
b)
Figure 53 TEM images of 2:1 MH/OT batch A s onicated MPMNs prepared 1 minute ( i=0) after adding the (a) 0.5 and
(b) 0.1 mM solution of cross-linker, NDT. Upper images: 1, 2 and 3 – deta iled images showing ordered assemblies. 4
– a magnif ied example of structure lacking the degree of order observed on the previous examples . Lower images:
blue arrows pointing to linear assembly based arrangements; grey arrow: unordered three-dimens iona l assemblies.
Scale bars: 50 nm
72 of 94
A general overview over the TEM grid shows that the solution cross -linked with a concentration of 1 mM
of dithiol has small assemblies like the ones observed without the presence of cross-linker (see Figure 33
– Control experiments). However, despite the lower molar concentration (0.5 and 0.1 mM), one minute
after adding the cross -linker solution, the TEM images show for both the concentrations random
aggregates of aggregates (300 - 3000 nm), with inhomogeneous form, dispersed in the microgrid.
Taking in consideration the last unexpected results, a fresh 1 mM solution of 1,9 -nonanedithiol was
prepared. The soluti on previously used was two months old so it was hypothesized that the cross -linker
solution could deteriorate with time which could explain the fact that less concentrated dithiol solutions
were forming faster big three-dimensional structures. However the results (see Figures 54 and 55)
indicate that the cross-linker stays stable in solution since there were no differences between the TEM
grids prepared with the two 1 mM NDT solutions synthesized with two months of difference.
Representative TEM micrographs are presented below:
b)
a)
Figure 54 TEM images of 2:1 MH/OT batch A MPMNs (a) 1 minute (i=0) and (b) 5 minutes after adding the 1 mM
solution of cross-linker, NDT (prepared on the 4th of December 2009). Blue arrows point to details of the threedimensional assemblies where a linear-based structural arrangement is observed. Scale bars: 50 nm
a)
b)
Figure 55 TEM images of 2:1 MH/OT batch A MPMNs (a) 1 minute (i=0) a nd (b) 5 minutes after adding the fresh 1
mM solution of cross-linker, NDT (prepared on the 4th of February 2010). The blue arrow indicates a detail of the
assembly where a linear-based structural arrangement is observed. Scale bars: 50 nm
All the TEM images (Figures 44-55) show randomized chain-like structures. There are a variety of
justifications for the existence of such networks in the TEM grid: (a) secondary bonding occurs between
73 of 94
the alcohol groups in the ligand shells of neighboring NPs. (b) In the case of the rippled NPs, the pole
functionalization reaction conditions are not optimized and consequently place -exchange occurs not
only at the polar defects but also at other defect s ites in the ligand shell [91] and (c) the presence in
solution of excess thiol which was proven to play a major role in the formation of superlattice structures
[129]. However these explanations are not enough to clarify why are the 0.1 and 0.5 cross -linker
solutions making the MPMNs assemble faster than the 1 and 5 mM solutions.
It is also noteworthy to mention the two different types of arrangement that can be observed in the
aggregated structures. Blue arrows on Figures 53, 54 and 55 and images 1, 2 and 3 on Figure 53 point to
assembly structures where a very organized linear –based structure can be observed which suggests
that these structures have actually a high degree of organization. These observations suggest that this
type of aggregates observed on the TEM grids are composed of covalently bonded one- and twodimensional chains that are clustered into larger structures by van der Waals interdigitation forces. On
the other hand grey arrows on Figures 53, 54 and 55 and image 4 on figure 53 highlight structures
lacking the degree of organization observed on the examples mentioned on the last paragraph. Instead
unordered three-dimensional assemblies were formed.
Hussain et al. found that when the amount of cross -linker is reduced below a certain ratio (60 NDT/gold
NPs) the ligands at the gold NP surface will be easily replaced by dithiols, and the partial capping of gold
NPs with linker molecules would then result in cross -linking leading to insoluble and irregular aggregates
[102]. The ratios of cross-linker solution applied and the kinetics of the place -exchange reaction will
result in a fast assembling process. For these reasons it is very challenging to control the aggregation
and assess an intermediate stage of the process whe re small one- and two-dimensional structures can
be observed.
Based on the previous results, and on the control experiments made witho ut the cross-linker (section
5.3.2.3) fresh solutions of MPMNs were prepared, diluted (0.125 mg/ml), sonicated for two hours and
left to stir for a minimum period of two days.
In concentrated solutions (e.g. 0.25 mg/ml) particles are already too close so can easily interdigitate and
bind to the neighbouring equivalents. Thus, initially, 1:100 and 1:1000 dilutions of the source solutions
(0.25 mg/ml) were prepared and analysed however, for these concentrations there were only a few
particles in solution which means it was difficult to see them in the TEM. (Images not shown)
74 of 94
5.3.2.6. Assembly after Purification Steps
Once completed the stabilization study of the MPMN solutions (see section 5.3.2.3) and after verifying
that NPs were spontaneously assembling in solution it was decided to start doing purification steps
before preparing the TEM grids. After confirming the absence of big three dimensional structures in the
TEM samples prepared with the purified MPMN solutions, the effect of the cross-linker in these samples
was then studied.
The first set of TEM grids was prepared three days after the purification steps and the second one was
prepared just one hour after the purification protocol. For these two systems the cross -linker and source
solution concentrations were also varied. The different conditions used are described in the following
table:
Table 15 Different conditions used in the assembly study, when the solutions analysed were submitted to the
purification protocol
Purification steps
Syringe Filtration (0.1 µm) and centrifugation (15 000 rpm – 8 min)
1:1, 2:1 and 5:1 MH/OT Batch B
TEM grids prepared 3 days after the purification steps
Cross-linker
NDT – 1mM in EtOH
Concentration of AuNPs solution
1:10 dilution –0.025 mg/ml
Cross-linker to gold NP ratio
2.5E+04:1
Time after adding the cross-linker
i=0 (few seconds after) i=1 (10 mins after)
1:1 MH/OT Batch B
TEM grids prepared 1 hour after the purification steps
Cross-linker
NDT – 0.01mM in EtOH
Concentration of AuNPs solution
1:5 dilution –0.05 mg/ml
Cross-linker to gold NP ratio
125:1
Time after adding the cross-linker
i=1 (10 mins after) i=2 (1 hour after) and i=3 (12 hour after)
75 of 94
Figure 56 TEM pictures of the (a) 1:1 (b) 2:1 and (c) 5:1 MH/OT batch B purif ied MPMNs solutions (0.025 mg/ml)
prepared 1 minute ( i=0) after adding the 1mM cross-linker solution, NDT and (d) 1:1 (e) 2:1 and (f) 5:1 MH/OT batch
B prepared 10 minutes (i=1) after adding the same solution of cross-linker respectively.
In all cases, after a general view of the TEM grids there were a fe w aggregates. It was actually possible to
see these aggregates were slightly bigger 10 minutes after adding the cross-linker.
This slower kinetics of aggregation can be a result of the lower concentration of MPMNs solution used.
Decreasing the solution concentration increases the interparticle distance which leads to a reduced
probability of interdigitation or van der Waals interactions between neighbouring particles.
However, considering the ratio of 2.5E+04 molecules of cross-linker per gold NP which means 83 NDT to
thiol ratio, a higher disparity between the two different analysed times should be expected. This fact
combined with the lack of intermediate structures in the samples and the time factor (purification steps
performed three days before) drove the present work to the same questions: Are the existing
aggregates actually an effect of the cross -linker? Are the gold NPs self-assembling in solution? Or are
these structures a result of both mechanisms?
76 of 94
On the other hand, the results obtained after adding a low concentration cross -linker solution to a
MPMNs solution purified one hour before (see Figure 57) show that the assembly was only visible 12
hours after mixing these solutions, however intermediate structures were not observable in the samples
leading again to the same question: are the aggregates a result of spontaneous assembly or a result of
the cross-linker addition?
Figure 57 TEM pictures of the 1:1 MH/OT batch B purified MPMNs solution (0.05 mg/ml) prepared 10 minutes, 1
and 12 hours after adding the 0.01 mM cross-linker solution (1,9-nonanedithiol). Aggregation only visible after 12
hours. Scale bars: 50 nm
Interpreting these images is further complicated by the high interdigitation of gold NPs, in that it is
possible that the aggregates observed in the TEM grids are composed of cross-linked one- and twodimensional aggregates that are further assembled into larger structures by van der Waals forces and
cross-linker action.
5.3.2.7. New cross-linker approach
Taking in consideration the last results a new approach involving a different cross-linker was followed.
Considering that molecules in the ligand shell, MH and OT, are 6 and 8 carbons in length, respectively
and that the previous cross -linker (NDT) is 9 carbons, it is difficult to discern if the cross -linker is in fact
responsible for linking the neighboring particles together or is just helping or being an intermediate of
interdigitation and interparticle interactions between the ligands bond to the surface of each NP (see
Figure 58). For this reason a bigger cross-linker particle – 1,16-hexadecanedithiol (HDDT) was tested, in
order to achieve a faster place exchange reaction, using a l ower ratio of cross-linker per gold NP. This
longer cross-linker will enable the assembly of the particles together without needing to form any extra
interactions or interdigitation between the ligands.
77 of 94
a)
b)
Figure 58 Schematic illustrating the cross-linking process for the two different dithiol containing molecules used: (a)
NDT and (b) HDDT (9 and 16 carbons in length respectively). Since the ligands in the SAM are 8 and 6 carbons in
length (a) a 9 carbon in length dithiol molecule will only link 2 particles together by chang ing the conformation of
the ligand shell, which will require a high degree of interdigitation and intermolecular interactions between the
ligands of the neighboring particles. On the other hand, (b) a 16 carbon in length dithiol molecule will link the
particles together without needing any extra interactions or interdigitations, which translates in a more
energetically favorable reaction, and thus a faster and more effective assembly process.
It is evident that when a dithiol-containing molecule solution in a ratio of 2500:1 NDT:gold NP is added,
it is going to make particles aggregate. This fact is due to the direct competition between the cross linker solution and the thiols assembled to the surface of the gold NPs. In such a high ratio it will placeexchange imprecisely in the gold NP surface and not just on the defects. Thus, in order to see a
preferential place-exchange reaction, e.g. the place-exchange reaction occurring only in the defect spots
existent in the ligand shell we need to add the cross -linker in a ratio of 25 dithiol/gold NP (no more than
that) and be able to observe the reaction taking place in a countable time.
78 of 94
Figure 59 TEM images of 1:1 MH/OT Batch B MPMNs solution (0.05 mg/ml) 1 and 15 minutes, 1, 3 and 18 hours
after adding a 0.01 mM cross-linker solution (HDDT) (25:1 HDT/gold NP ratio) Scale bars: 50 nm
The TEM images revealed that aggregation was only visible 18 hours after adding the cross -linker
solution. Once again, this can be a result of spontaneous assembly of the particles in solution and not a
result of the cross-linker itself. The low HD DT/gold NP ratio used can be a cause for such a slow place
exchange kinetic, because it will not be energetically favourable for the SAM to lose its conformational
and stable morphology to let the cross -linker molecules bond to the particles’ surface. Also considering
that this cross-linker molecule is 16 carbons in length, the loop of it should be considered. In this case
the cross-linker molecule will form two thiol -gold bonds in the same gold NP and derail the further
linking with other particles (see Figure 60).
Figure 60 Schematic illustrating the incoming cross-linker molecule coiling around in the g old NP forming two thiolgold bonds in the same particle.
79 of 94
In order to test if the cross -linker was effectively linking the particles and also confirm that not all the
cross-linker molecules were coiling around the particle, a new 5 mM sol ution of HDDT (2500:1 HDT/gold
NP ratio) was added.
Aggregation was observed 1 minute after adding the cross -linker solution. The assembly process was
also followed with the naked eye. In approximately 2 hours the solution changed from a light pink -red
colour to a browner one and finally to a dark precipitate.
Figure 61 TEM images of 1:1 MH/OT Batch B MPMNs solution (0.05 mg/ml) 1 minute after adding a 5 mM cross linker solution (HDDT) Scale bars: 50 nm.
80 of 94
6. Conclusions and Future Work
With the advent of nanotechnology and nanofabrication techniques and the development of new tools
for analysing and imaging nanoscaled objects, nanostructured systems can now be easily created and
modified at both structural and chemical levels, and represent the subject of fundamental scientific
studies of nano- and molecular-scale interactions. However, the device application of nanoscaled
systems lacks of suitable methods to organize them into well-defined structures.
Founded on the Stellacci’s group studies [51, 52, 72, 75-77, 91], which have explored the induced
assembly into linear chains of MPMNs coated with a binary mixture of ligands that self-assemble in the
ligand shell forming sub-nanometer-ordered domains, the present thesis was developed regarding the
relationship between the ligand shell structure (presence or absence of sub-nanometer-ordered
domains in the SAM) and their assembly using a dithiol as cross-linker.
In the first part of this thesis, the systematic synthesis, using a relatively fast and cost -effective onephase method [43], and further characterization of seven series of differently coated MPMN systems is
reported. The method followed allows the production of MPMNs with a narrow size distribution on a
gram scale.
In the second part of this project, important information regarding these particles spontaneous
assembly in solution is provided. Both in MPMNs assembly studies and i n MPMN on surfaces for protein
and cells interaction studies, stable solutions of MPMNs are required (i.e. without the presence of
aggregates), therefore a purification method was developed, allowing the production of isolated
MPMNs in solution.
The third part describes the different approaches followed in the study of the particle assembly into
larger structures through induced place exchange reactions using a dithiol -containing molecule. Several
parameters were varied including the MPMNs solution and cross -linker concentrations, time and cross linker type molecule.
It was extremely challenging to control the number of functional groups attached to each parti cle. Given
the example of MPMNs with a core diameter around 4.6 nm, there are in total about 300 surface gold
atoms, and the total number of organic thiol ligands that may be attached to th e particles is also close to
300 [20, 67]. Thus, discarding the hypothesis of place-exchange reaction at the, if present, polar defects
due to the high cross -linker/gold NPs ratios employed, these particles will present multiple unknown
numbers of functional groups .
When the cross-linker concentration was decreased, evident NP assemblies were hardly observable due
to the poor place exchange reaction kinetics. In such low concentrations it was not energetically
81 of 94
favourable to change the stable conformational morphology of the ligand shell to place -exchange with
an incoming molecule that forms the same bond (sulfur -gold) in the gold NP surface.
Due to the lack of a precise control over the number of functi onal groups, any chemical reactions
conducted on such nanoparticles most likely will lead to the formation of large aggregates with
unknown and irreproducible structures and properties or no clear aggregation at all [130].
As a first step to overcome this challenge, one needs to be able to produce nanoparticles with a
controlled number of chemical functional group attached to the surface that can later link directly or
exchange with a cross-linker molecule inducing assembly in predictable and controlled way. The
development of such regioselective chemistry on NPs can then enable the production of wide variety of
complex nanostructures.
Improvements can still be made in the synthesis and assembly study procedures, and there are s everal
potential directions for this research to proceed. A few options are discussed here:
The choice of the solvent and incoming molecules should be carefully studied and analyzed in order to
shift the reaction dynamic equilibrium favouring the adsorbed state of the incoming molecule. For
instance, an incoming molecule that is only slightly soluble in the reaction solvent would exchange onto
poles or other eventual defects in the ligand shell but feel only a weak driving force to desorb. In this
situation a more effective and faster place exchange reaction rate would take place allowing us to work
with lower ratios of cross -linker/gold NP, and in the case of rippled NPs observe the formation of one dimensional chains.
The composition of the ligand shell could also be altered in order to favour the place exchange reaction
rates. For instance, tetraoctylammonium bromide coated gold NPs exchange faster and more easily with
the dithiol-containing incoming molecules because thiols form a stronger bond in the go ld surface than
the bromide molecules [102, 131]. Thus, using a low ratio of cross -linker/gold NP molecules
(approximately 20) we should be able to observe preferential and controlled place exchanging at the
gold NP surface.
Moreover alternative linking chemistries should be further studied to identify any potential advantages
they may present over the thiol-bond coupling used in this work. For instance biological molecules
represent ideal candidates for use as pole functionalization molecules [132] due to the specificity of
their chemistry that would allow precise control over the chain composition.
Finally, an alternative to form structures following a particle by particle trend, focused on the control of
chemical and inter-particle reactivity at the single NP level, is to simultaneously arrange several particles
into self-assembled materials. This approach yielded a bigger scope of new structures at the nanometer
scale with a multitude of potential applications such as optical coatings and gas sensors [102].
82 of 94
7. References
1.
Feynman, R.P., Plenty of room at the bottom. J. Microelectromechanical Syst, 1992. 1: p. 60.
2.
Qun, H.W., J. G., Monofunctional gold nanoparticles: synthesis and applications. Journal of
Nanoparticle Research, 2006. 9(6): p. 1013-1025.
3.
Rejesky, D., Cpsc fy2010 agenda and priorities. , in Consumer Product, T. Report, Editor. 2009,
Safety Comission.
4.
Diegoli, S.M., A. L.; Begum, S.; Jones, I. P.; Lead, J. R.; Preece, J. A., Interaction between
manufactured gold nanoparticles and naturally occurring organic macromolecules. Science,
2008. 402(1): p. 51-61.
5.
Colvin, V.L., The Potential Environmental Impact of Engineered Nanomateri als. Nature
Biotechnology, 2003. 21(10): p. 1166-70.
6.
Kanellos, M. Carbon nanotubes enter Tour of France. CNET news.com 07/07/2006.
7.
Astruc, D., DM-C. , Gold nanoparticles: Assembly, supramolecular chemistry, quantumsizerelated properties, and applications toward biology, catalysis, and nanotechnology. Chemical
Reviews, 2004. 104(1): p. 293-346.
8.
Link, S.E.-S., M. A. , Spectral properties and relaxation dynamics of surface plasmon electronic
oscillations in gold and silver nanodots and nanorods. Journal of Physical Chemistry B 1999.
103(40): p. 8410-26.
9.
Andres, R.P.e.a., ‘‘Coulomb staircase’’ at room temperature in a self-assembled molecular
nanostructure. Science, 1996. 272(5266): p. 1323-1325.
10.
Love, J.C.E., L. A.; Kriebel, J. K.; Nuzzo, R. G.; Whitesides, G. M., Self-Assembled Monolayers of
Thiolates on Metals as a form of Nanotechnology. Chemmical Reviews, 2005. 105(4): p. 11031170.
11.
Templeton, A.C.W., W.P.; Murray, R. W. , Monolayer-Protected Cluster Molecules. Accounts of
Chemical Research, 2000. 33(1): p. 27-36.
12.
Mirkin, C.A., The beginning of a small revolution. Small, 2005. 1(1): p. 14-16.
13.
Graigner, D.W.C., D. G.,, Nanobiomaterials and Nanoanalysis: Opportunities for Improving the
Science to Benefit Biomedical Technologies. Advanced Materials, 2008. 20(5): p. 867-877.
14.
Ratner, M.R., D. , Nanotechnology: a gentle introduction to the next bigidea, ed. I. BooksCraft.
2003, Indianopolis: Bernard Goodwin.
15.
Yang, Y.Y., Y.; Wang, W.; Li, J.;, Precise Size Control of Hydrophobic Gold Nanoparticles using
Cooperative Effect of Refluxing Ripening and Seeding Growth. Nanotechnology, 2008. 19(17): p.
175603.
16.
Alivisatos, A., Semiconductor clusters, nanocrystals, and quantum dots. . Science, 1996.
271(5251): p. 933-37.
83 of 94
17.
Daniel M, A.D., Gold nanoparticles: Assembly, supramolecular chemistry, quantumsize-related
properties, and applications toward biology, catalysis, and nanotechnology. Chemical Reviews,
2004. 104(1): p. 293-346.
18.
Adamson, A.W.G., A. P. , Physical Chemistry of Surfaces. 6th ed. Catalytic Chemistry, ed. J.W.
Sons. 1997, New York.
19.
Yee, C.K.J., R.; Ulman, A.; White, H.; King, A.; Rafailovich, M.; Sokolov, J., , Novel One-phase
Synthesis of Thiol-Functionalized Gold, Palladium and Iridium Nanoparticles using Superhydride.
Langmuir, 1999. 15(10): p. 3486-3491.
20.
Brust, M.W., M.; Bethell, D.; Schiffrin, D. J.; Whyman, R., Synthesis of thiol-derivatised GoldNanoparticles in a Two-phase Liquid-Liquid System. Journal of Chemical Society, 1994: p. 801-2.
21.
Niemeyer, C.M., Nanoparticles, proteins, and nucleic acids: Biotechnology meets materials
science Angewandte Chemie-International Edition 2001. 40(22): p. 4128-58.
22.
Feldheim, D.L., Keating, C. D., , Self-assembly of single electron transistors and related devi ces
Chemical Society Reviews 1998. 27(1): p. 1-12.
23.
Hostetler, M.J.G., S. J.; Stokes, J. J.; Murray, R. W., Monolayers in three dimensions: Synthesis
and electrochemistry of omega-functionalized alkanethiolate-stabilized gold cluster compounds.
Journal of the American Chemical Society 1996. 118(17): p. 4212-3.
24.
Link, S.E.-S., M. A., Size and temperature dependence of the plasmon absorption of colloidal
gold nanoparticles. . Journal of Physical Chemistry B 1999. 103(21): p. 4212-7.
25.
Collier, C.P.S., R. J.; Shiang, J. J.; Henrichs, S. E.; Heath, J. R., Reversible tunning of silver
quantum-dot monolayers through the metal -insulator transistor. Science, 1999. 277(5334): p.
1978-81.
26.
Malinsky, M.D.K., K. L.; Schatz, G. C.; Van Duyne, R. P.,, Chain length dependence and sensing
capabilities of the localized surface plasmon resonance of silver nanoparticles chemically
modified with alkanethiol self-assembled monolayers Journal of the American Chemical Society
2001. 123(7): p. 1471-82.
27.
Thomas, K.G.K., P. V.,, Chromophore-functionalized gold nanoparticles Accounts of Chemical
Research, 2003. 36(12): p. 888-98.
28.
Hayat, M.A., Colloidal Gold - Principles, Methods, and Applications, ed. I. Academic Press. 1989,
San Diego: Harcourt Brace Jovanovich, Publishers.
29.
Freestone, I., Meeks, N., Sax, M., Higgitt, C., The Lycurgus Cup - A Roman Nanotechnology. Gold
Bulletin, 2007. 40(4): p. 270-277.
30.
Antonii, F., Panacea Aurea-Auro Potabile. Bibliopolio Frobeniano, 1618.
31.
Kunckels, J., Nuetliche Observationes oder Anmerkungen von Auro und Argento Potabili.
Schutzens, 1676.
32.
Klickstein, H.S.L., H. M., A Source Book in Chemistry, 1400 - 1900, ed. O.U. Press. 1968.
33.
Helcher, H.H., Aurum Potabile oder Gold Tinstur. J. Herbord Klossen, 1718.
84 of 94
34.
Thompson, D., Michael Faraday's Recognition of Ruby Gold: the Birth of Modern
Nanotechnology. Gold Bulletin, 2007. 40(4): p. 267-269.
35.
Faraday, M., The Bakerian Lectura: Experimental Relations of Gold (and Other Metals) to Light.
Philosophical Transactions Royal Society of London 1857. 147: p. 145-181.
36.
Schmid, G., Clusters and Colloids - From Theory to Applications, ed. VCH. 1994, Weinheim.
37.
Glogowski, E.H., J.; Russel, T. P.; Emrick, T. , Mixed monolayer coverage on gold nanoparticles
for interfacial stabilization of immiscible fluids. Chemical Communications, 2005. 32: p. 405052.
38.
Foos, E.E.S., A. W.; Twigg, M. E.; Ancona, M. G, Thiol-Terminated Di-, Tri-,and Tetraethylene
Oxide Functionalized Gold Nanoparticles: A Water-Soluble, Charge-Neutral Cluster. . Chemistry
of Materials, 2002. 14(5): p. 2401-2408.
39.
Kanaras, A.G.K., F. S.; Schaumburg, K.; Kiely, C. J.; Brust, M. , Thiolalkylated tetraethylene glycol:
a New Ligand for Water Soluble Monolayer Protected Gold Clusters. . Chemical
Communications, 2002. 20: p. 2294-2295.
40.
Yonezawa, T.S., M.; Kunitake, T. , Practical Preparation of Size-Controlled Gold Nanoparticles in
Water. Chemistry Letters 1997. 26(7): p. 619-620.
41.
Warner, M.G.R., S. M.; Hutchison, J. E., Small, Water-Soluble, Ligand-Stabilized Gold
Nanoparticles Synthesized by Interfacial Ligand Exchange Reactions. . Chem. Mater. , 2000.
12(11): p. 3316-3320.
42.
Rowe, M.P.P., K. E.; Kim, K.; Kurdak, Ç.; Zellers, E. T.; Matzger, A. J., , Single-Phase Synthesis of
Functionalized Gold Nanoparticles. Chemistry of Materials, 2004. 16(18): p. 3513-17.
43.
Zheng, N.F., J.; Stucky, G. D., On e-Step One-Phase Synthesis of Monodisperse Noble-Metallic
Nanoparticles and their Colloidal Cristals, . Journal of the American Chemical Society, 2006.
128(20): p. 6550-1.
44.
Mie, G., Beiträge sur Optik trüber Medien, speziell killoidaler Metallösungen. Annalen der
Physik, Vierte Folge, 1908. 25(3): p. 376-445.
45.
Barnes, W.L.D., A.; Ebbesen, T. W. , Surface plasmon subwavelength optics. Nature, 2003.
424(6950): p. 824-830.
46.
Du, G.X.M., T.; Suzuki, M.; Saito, S.; Fukuda, H.; Takahashi, M. , Evidence of localized surface
plasmon enhanced magneto-optical effect in nanodisk array Applied Physics Letters 2010. 96(8):
p. 081915.
47.
Liz-Marzán, L.M., Tailoring Surface Plasmons through the Morphology and Assembly of Metal
Nanoparticles. Langmuir, 2006. 22(1): p. 32-41.
48.
Link, S.W., Z. L.; El -Sayed, M. A., Alloy formation of Gold-silver Nanoparticles and the
Dependence of the Plasmon Absorption on their Composition. Journal of Physical Chemistry B,
1999. 103(18): p. 3529-33.
49.
Schasfoort, R.B.M.T., A. J. , Handbook of Surface Plasmon Resonance, ed. T.R.S.o. Chemistry.
2008, Cambridge: RSC publishing.
85 of 94
50.
Mrksich, M.W., G. , Using self-assembled monolayers to understand the interactions of manmade surfaces with proteins and cells. Annual Review of Biophysics and Biomolecular Structure,
1996. 25: p. 55-78.
51.
Jackson, A.M.H., Y; Silva, P. J.; Stellacci, F From Homoligand- to Mixed-Ligand- MonolayerProtected Metal Nanoparticles: A Scanning Tunneling Microscopy Investigation Journal of the
American Chemical Society, 2006. 128(34): p. 11135-49.
52.
Jackson, A.M.M., J. W.; Stellacci, F. , Spontaneous assembly of subnanometre ordered domains
in the ligand shell of monolayer-protected nanoparticles Nat Matter, 2004. 3(5): p. 330-336.
53.
Akthakul, A.H., A. I.; Stellacci, F.; Mayes, A. M. , Size Fractionation of Metal Nanoparticles by
Membrane Filtration. Adv. Mater, 2005. 17(5): p. 532-535.
54.
Georganopoulou, D.G.C., L.; Nam, J. M.; Thaxton, C. S.; Mufson, E. J.; Klein, W. L.; Mirkin, C. A.,
Nanoparticle-based Detection in Cerebral Spinal Fluid of a Soluble Pathogenic Biomarker for
Alsheimer’s Disease Proceedings of the National Academy of Sciences U.S.A. , 2005. 102(7): p.
2273-2276.
55.
Wu, Z.Z., B.; Yan, B. , Regulation of Enzyme Activity through Interactions with Nanoparticles.
International Journal of Molecular Sciences, 2009. 10(10): p. 4198-4209.
56.
Andres, R.P.B., J. D.; Henderson, J. I.; Janes, D. B.; Kolagunta, V. R.; Kubiak, C. P.; Mahoney, W.
J.; Osifchin, R. G. , Self-Assembly of a two-dimensional superlattice of Molecularly linked Metal
Clusters. Science, 1996. 273(5282): p. 1690-1693.
57.
Ingram, R.S.M., R. W., Electroactive Three-Dimensional Monolayers: Anthraquinone ωFunctionalized Alkanethiolate-Stabilized Gold Clusters. . Langmuir, 1998. 14(15): p. 4115-4121.
58.
Ulman, A., Formation and Structure of Self-assembled monolayers. Chemical Reviews, 1996.
96(4): p. 1533-54.
59.
Schwartz, D.K., Mechanisms and Kinetics of Self-Assembled Monolayer Formation Annual
Review of Physical Chemistry, 2001. 52: p. 107-137.
60.
Vos, J.G.F., R. J.; Keyes, T. E. , Formation and Characterization of Modified Surfaces, in
Interfacial Supramolecular Assemblies, L. John Wiley & Sons, Editor. 2003, Wiley-Interscience:
West Sussex, England. p. 88-94.
61.
Hasan, M.B., D.; Brust, M., The Fate of Sulfur-Bound Hydrogen on Formation of Self-Assembled
Thiol Monolayers on Gold: 1H NMR Spectroscopic Evidence from Solutions of Gold Clusters.
Journal of the American Chemical Society, 2002. 124(7): p. 1132-3.
62.
Bareman, J.P.K., M. L. , Collective tilt behavior in dense, substrate-supported monolayers of
long-chain molecules: a molecular dynamics study. J. Phis. Chem., 1990. 94(13): p. 5202-5.
63.
Vericat, C.V., M. E.; Salvarezza, R. C., Self-assembled monolayers of alkanethiols on Au(111):
Surface structures, defects and dynamics. Journal of Physical Chemistry B, 2005. 7(18): p. 325868.
64.
Prime, K.W., G.;, Self-Assembled Organic Monolayers - Model Systems for studying adsorption
of proteins at surfaces. Science, 1991. 252(5009): p. 1164-7.
86 of 94
65.
Zanchet D, H.B., Ugarte D. , Structure population in thiol -passivated gold nanoparticles. J. Phys
Chem B. , 2000. 104(47): p. 11013-11018.
66.
Gutierrez, W.C.A., J.; Perez-Alvarez, M.; Marin-Almazo M.; Yacaman, J. M. , On the structure and
formation of self-assembled lattices of gold nanoparticles. J Clust Sci. , 1998. 9(4): p. 529-545.
67.
Hostetler, M.J.T., A. C.; Murray, R. W.; , Dynamics of Place-Exchange Reactions on MonolayerProtected Gold Cluster Molecules. Langmuir, 1999. 15(11): p. 3782-89.
68.
Georgakilas, V.G., D.; Tzitzios, V.; Pasquato, L.; Guildi, D. M.; Prato, M., Decorating Carobon
Nanotubes with Metal or Semiconductor Nanoparticles. Journal of Materials Chemistry, 2007.
17(26): p. 2679-94.
69.
Stranick, S.A., S.; Parikh, A.; Wood, M. , Nanometer-scale phase separation in mixed
composition self-assembled monolayers. Nanotechnology, 1996. 7(4): p. 438-442.
70.
Folkers, J.L., P.; Whitesides , G. , Self-assembled monolayers of alkanethiols on gold:
comparisons of monolayers containing mixtures of short - and long-chain constituents with
methyl and hydroxymethyl terminal groups. . Langmuir, 1992. 8(5): p. 1330-1341.
71.
Smith, R.K.R., S. M.; Lewis, P. A.; Monnell, J. D.; Clegg, R. S.; Kelly, K. F.; Bumm, L. A.; Hutchison,
J. E.; Weiss, P. S., Phase separation within a binary self-assembled monolayer on Au{111} driven
by an amide-containing alkanethiol Journal of Physical Chemistry B, 2001. 105(6): p. 1119-1122.
72.
Centrone, A.H., Y.; Jackson, A. M.; Zerbi, G.; Stellacci, F. , Phase separation on mixedmonolayer-protected metal nanoparticles: a study by infrared spectroscopy and scanning
tunneling microscopy. . Small, 2007. 3(5): p. 814-817.
73.
Singh, C.G., P. K.; Horsch, M. A.; Jackson, A. M.; Larson, R. G.; Stellacci, F.; Glotzer, S. C.,
Entropy-Mediated Patterning of Surfactant-Coated Nanoparticles and Surfaces Phys Rev L ett .,
2007. 99(22): p. 226106.
74.
Casagrande, C.V., C.R. , Janus Beads - Realization and 1st Observation of Interfacial Properties.
Comptes Rendus de L'Academie des Sciences, 1988. 306: p. 1423-25.
75.
Carney, R.P.D., G.A.; Dubois, C.; Kim, H.; Kim, J.Y.; Singh, C.; Ghorai, P. K.; Tracy, J. B.; Stiles, R.
L.; Murray, R. W.; Glotzer, S. C.; Stellacci, F. , Size limitations for the formation of ordered
striped nanoparticles. Journal of the American Chemical Society, 2008. 130(3): p. 798-799.
76.
Centrone, A.P., E.; Sharma, M.; Myerson, J.W.; Jackson, A.M.; Marzari, N.; Stellacci, F., The role
of nanostructure in the wetting behavior of mixed-monolayer-protected metal nanoparticles.
Proc Natl Acad Sci USA, 2008. 105(29): p. 9886-9891.
77.
Hu, Y.U., O.; Dubois, C.; Stellacci, F. , Effect of Ligand Shell Structure on the Interaction between
Monolayer-Protected Gold Nanoparticles. Journal of Physical Chemistry C, 2008. 112(16): p.
6279-6284.
78.
Uzun, O.H., Y.; Verma, A.; Chen, S.; Centrone, A.; Stellacci, F., Water-Soluble Amphiphilic Gold
Nanoparticles with Structured Ligand Shells,. Journal of the Chemical Society, Chem. Commun. ,
2007. 2: p. 196-198.
87 of 94
79.
McFarland, A.D.V.D., R. P., Single Silver Nanoparticles as Real -Time Optical Sensors with
Zeptomole Sensibity. Nano Letters, 2003. 3(8): p. 1057-62.
80.
Wooley, K., New nanoparticle coating mimics dolphin skin prevents 'biofouling' of ship hulls, in
Innovations Report - Forum for Science, Industry and Business. 2002, Materials Sciences.
81.
Brown, L.O.H., J. E. , Convenient Preparation of Stable, Narrow-Dispersity, Gold Nanocrystals by
Ligand Exchange Reactions. Journal of the American Chemical Society, 1997. 119(50): p. 1238412385
82.
Dubois, L.H.N., R. G., The Synthesis, Structure, and Properties of Model Organic Surfaces. Annual
Reviews of Physical Chemistry 1992. 43: p. 437-463.
83.
Collard, D.M.F., M. A. , Use of electroactive thiols to study the formation and exchange of
alkanethiol monolayers on gold. Langmuir, 1991. 7(6): p. 1192-1197.
84.
Scott, J.R.B., L. S.; Everett, W. R.; Wilkins, C. L.; Fritsch,I. , Laser Desorption Fourier Transform
Mass Spectrometry Exchange Studies of Air-Oxidized Alkanethiol Self-Assembled Monolayers on
Gold. Analytical Chemistry, 1997. 69(14): p. 2636-2639.
85.
Schlenoff, J.B.L., M.; Ly, H. , Stability and Self-Exchange in Alkanethiol Monolayers. Journal of
the American Chemical Society, 1995. 117(50): p. 12528-12536.
86.
Nishida, N.H., M.; Sasabe, H.; Knoll, W. , Formation and Exchange Processes of Alkanethiol SelfAssembled Monolayer on Au(111) Studied by Thermal Desorption Spectroscopy and Scanning
Tunneling Microscopy. Japanese Journal of Applied Physics, 1996. 36: p. 2379-2385.
87.
Ingram, R.S.H., M. J.; Murray, R. W. , Poly-hetero-ω-functionalized Alkanethiolate-stabilize Gold
Cluster Compounds. Journal of the American Chemical Society, 1997. 119(39): p. 9175-8.
88.
Westerlund, F.B., T. , Directed assembly of gold nanoparticles. Current Opinion in Colloid &
Interface Science 2009. 14(2): p. 126-134.
89.
Maier, S.A.B., M. L.; Kik, P. G.; Meltzer, S.; Requicha, A. A. G.; Koel, B. E.; Atwater, H. A.,
Plasmonics - a route to Nanoscale Optical Devices. Advanced Materials, 2001. 13(19): p. 1501-5.
90.
Thomas, K.G.B., S.; Ipe, B. I.; Joseph, S. T. S.; Kamat , P. V., Unidirectional Plasmon Coupling
through Longitudinal Self-assembly of Gold Nanorods. Journal of Physical Chemistry B, 2004.
108(35): p. 13066-68.
91.
DeVries, G.A.B., M.; Hu, Y.; Jackson, A. M.; Long, B.; Neltner, B. T.; Uzun, O.; Wunsch, B. H.;
Stellacci, F. , Divalent Metal Nanoparticles Science, 2007. 315(5810): p. 358-361.
92.
Rosi, N.L.T., C. S.; Mirkin, C. A., Control of Nanoparticles Assembly using DNA-modified Diatom
templates. Angewandte Chemie - International Edition, 2004. 43(41): p. 5500-3.
93.
Bachand, G.D.R., S. B.; Boal, A. K.; Gaudioso, J.; Liu, J.; Bunker, B. C. , Assembly and transport of
nanocrystal CdSe quantum dot nanocomposites using microtubules and kinesin motor proteins.
Nano Lett., 2004. 4(5): p. 817-21.
94.
Wang, G.L.M., R. W., Controlled assembly of Monolayer-Protected Gold Clusters by Dissolved
DNA. Nano Lett., 2004. 4(1): p. 95-101.
88 of 94
95.
Tang, Z.Y.K., N. A., One-Dimensional Assemblies of Nanoparticles: Preparations, Properties and
Promise. Adv. Mater. , 2005. 17(8): p. 951-62.
96.
Schreiber, F., Structure and growth of self-assembling monolayers Prog. in Surface Science,
2000. 65(5-8): p. 151-257.
97.
Luedtke, W.D.L., U., Structure and Thermodynamics of Self-Assembled Monolayers on Gold
Nanocrystallites Journal of Physical Chemistry B, 1998. 102(34): p. 6566-6572.
98.
Ghorai, P.K.G., S. C. , Molecular Dynamics Simulation Study of Self-Assembled Monolayers of
Alkanethiol Surfactants on Spherical Gold Nanoparticles. Journal of Physical Chemistry C, 2007.
111(43): p. 15857-62.
99.
Nelson, D.R., Toward a Tetravalent Chemistry of Colloids. Nano Letters, 2002. 2(10): p. 11251129.
100.
Brust, M.B., D.; Kiely, C.; Schiffrin, D. , Self-assembled Gold Nanoparticle Thin Films with
Nonmetallic Optical and Electronic Properties Langmuir, 1998. 14(19): p. 5425-5429.
101.
Brust, M.B., D.; Schiffrin, D. J.; Kiely, C. , Novel Gold-Dithiol Nano-networks with Non-metallic
Electronic Properties. J. Adv. Mater 1995. 7(9): p. 795-797.
102.
Hussain, I.W., Z.; Cooper, A. I.; Brust, M. , Formation of Spherical Nanostructures by the
Controlled Aggregation of Gold Colloids. Langmuir, 2006. 22(7): p. 2938-2941
103.
Sih, B.C.W., M. O. , Metal Nanoparticle-Conjugated Polymer Nanocomposites. Chemical
Communications, 2005: p. 3375-84.
104.
Perepichka, D.F.R., F. , Metal Nanoparticles: From “Artificial Atoms” to “Artificial Molecules”
Angewandte Chemie - International Edition, 2007. 46(32): p. 6006-6008.
105.
Sung, K.-M.M., D. W.; Peelle, B. R.; Zhang, S.; Jacobson, J. M. , Synthesis of Monofunctionalized
Gold Nanoparticles by Fmoc Solid-Phase Reactions. J. Am. Chem. Soc., 2004. 126(16): p. 506465.
106.
Worden, J.G.S., A. W.; Huo, Q. , Controlled Functionalization of Gold Nanoparticles through a
Solid Phase Synthesis Approach. Chemical Communications, 2004(5): p. 518-9.
107.
Huo, F.L.-J., A. K. R.; Mirkin, C. A. , Asymmetric functionalization of Nanoparticles based on
Thermally addressable DNA Interconnects. Advanced Materials, 2006. 18(17): p. 2304-6.
108.
Deng, Z.T., Y.; Lee, S. H.; Ribbe, A. E.; Mao, C. , DNA-Encoded Self-Assembly of Gold
Nanoparticles into One-Dimensional Arrays. Angewandte Chemie - International Edition, 2005.
44(23): p. 3582-85.
109.
Aldaye, F.A.S., H. F. , Sequential Self-Assembly of a DNA Hexagon as a Template for the
Organization of Gold Nanoparticles. Angew. Chem., 2006. 45(14): p. 2204-09.
110.
Hore, P.J., Nuclear Magnetic Resonance. Oxford Chemistry primers, ed. O.U. Press. 1995,
Oxford: Oxford University Press Inc.
111.
Hoppert, M., Microscopic Techniques in Bioltechnology, ed. W.-. VCH. 2003, Verlag.
112.
Ricci, D.B., P. C., Atomic Force Microscopy: Biomedical Methods and Applications. Methods in
Molecular Biology, ed. H. Press. Vol. 242. 2004, New Jersey.
89 of 94
113.
Vilarinho, P.M.R., Y; Kingon, A., Scanning Probe Microscopy: Characterization, Nanofabrication
and Device Application of Functional Materials. NATO Science Series, ed. K. academic. 2005,
Dordrecht.
114.
Kreibig, U.Q., M., Optical properties of aggregates ofsmall metal particles. Surface Science,
1986. 172(3): p. 557-577.
115.
Hyningl, D.L.V.Z., C. F., Formation Mechanisms and Aggregation Behaviour of Borohydride
Reduced Silver Particles. Langmuir, 1998. 14(24): p. 7034.
116.
Natan, M.J.B., K. R., Hydroxylamine Seeding of Colloidal Au Nanoparticles in Solution and on
Surfaces. Langmuir, 1998. 14(4): p. 726.
117.
Owen, T., Fundamentals of modern UV-visible spectroscopy, ed. Primer. 2000, Germany.
118.
Collier, C.P.S., R. J.; Shiang, J. J.; Henrichs, S. E.; Heath, J. R., Reversible tunning of silver
quantum-dot monolayers through the metal -insulator transistor. Science, 1997. 277(5334): p.
1978-81.
119.
Jacobsen, N.E., NMR Spectroscopy Explained - Simplified Theory, Applications and Examples for
Organic Chemistry and Structural Biology, ed. Wiley-Interscience. 2007, New Jersey: John Wiley
& Sons, Inc.
120.
Zhang, X.-F.Z., Z., Progress in Transmission Electron Microscopy 1 - Concepts and Techniques.
Springer Series in Surface Sciences, ed. G.G. Ertl, R.; Lüth,H.; Mills, D. L. 1999, New York:
Springer.
121.
Goodhew, P.J.H., F. J.; Beanland, R., Electron Microscopy and Analysis, ed. T.a. Francis. 2001,
London.
122.
Yao, N.W., Z. L., Handbook of Microscopy for Nanotechnology. 2005, New York: Kluwer
Academic Publishers
123.
Williams, D.B.C., C.B., Transmission Electron Microscopy - Basics. A Textbook for Materials
Science. 1996, New York: Plenum Press.
124.
Labande, A.R., J.; Astruc, D., Supramolecular Gold Nanoparticles for the Redox Recognition of
Oxoanions: Synthesis, Titrations, Stereoelectronic Effects, and Selectivity. Journal of the
American Chemical Society, 2002. 124(8): p. 1782-9.
125.
Liao, J.Z., Y.; Yu, W.; Xu, L.; Ge, C.; Liu, J.; Gu, N., Linear Aggregation of Gold Nanoparticles in
Ethanol. Colloids and surfaces. A, Physicochemical and engineering aspects, 2003. 223(1-3): p.
177-183.
126.
Galletto, P.B., P. F.; Girault, H. H.; Antoine, R.; Broyer, M., Size Dependence of the Surface
Plasmon Enhanced Second Harmonic Response of Gold Colloids:Towards a New Calibration
Method. Chemical Communications, 1999(7): p. 581-582.
127.
Ohara, P.C.H., J. R.; Gelbart, W. M., Self-Assembly of Submicrometer Rings of Particles from
Solutions of Nanoparticles. Angewandte Chemie - International Edition, 2003. 36(10): p. 10781080.
90 of 94
128.
Khanal, B.P.Z., E. R., Rings of Nanorods. Angewandte Chemie - International Edition, 2007.
46(13): p. 2195-2198.
129.
Sidhaye, D.S.P., B. L. V., Melting Characteristics of Superlattices of Alkanethiol-Capped Gold
Nanoparticles: The "Excluded" Story of Excess Thiol. Chemistry of Materials, 2010. 22(5): p.
1680-5.
130.
Shaffer, A.W.W., J. G.; Huo, Q., Comparison Study of the Solution Phase versus Solid Phase Place
Exchange Reactions in the Controlled Functionalization of Gold Nanoparticles. Langmuir, 2004.
20(19): p. 8434-8351.
131.
Shon, Y.-S.C., S.; Voundi, P., Stability of Tetraoctylammonium Bromide-Protected Gold
Nanoparticles: Effects of Anion Treatments. Colloids and surfaces A: Physicochemical and
Engineering aspects, 2009. 353(1-3): p. 12-17.
132.
Mann, S.S., W.; Li, M.; Connolly, S.; Fitzmaurice, D., Biologically Programmed Nanoparticle
Assembly. Advanced Materials, 1999. 12(2): p. 147-150.
91 of 94
Appendix
1:0 MH/OT
Figure A.1 1 H NMR spectrum obtained for 1:0 MH/OT in CD3 OD. The presence of a peak in the NMR spectrum
associated to the the hydrophilic ligand (MH at 3.4 ppm) shows the presence of free ligands in solution.
5:1 MH/OT
Figure A.2 1 H NMR spectrum obtained for 5:1 MH/OT in CD3 OD. The presence of a peak in the NMR spectrum
associated to the the hydrophilic ligand (MH at 3.4 ppm) shows the presence of free ligands in solution.
92 of 94
1:1 MH/OT
Figure A.3 1 H NMR spectrum obtained for 1:1 MH/OT in CD3 OD. The presence of a peak in the NMR spectrum
associated to the the hydrophilic ligand (MH at 3.4 ppm) shows the presence of free ligands in solution.
1:2 MH/OT
Figure A.4 1 H NMR spectrum obtained for 1:2 MH/OT in CD3 OD. The presence of a peak in the NMR spectrum
associated to the the hydrophilic ligand (MH at 3.4 ppm) shows the presence of free ligands in solution.
1:5 MH/OT
Figure A.5 1 H NMR spectrum obtained for 1:5 MH/OT in CD3 OD. The presence of a peak in the NMR spectrum
associated to the the hydrophilic ligand (MH at 3.4 ppm) shows the presence of free ligands in solution.
93 of 94
Figure A.6 UV/Vis spectra of 1:2Batch A MH/OT particles. Only a very slight decrease of the SPR band was observed
suggesting that the aggregation was taking more time
94 of 94