Gold nanoparticles in analytical chemistry

Skeete,Department of Chemistry, State University of New York at hamton, Binghamton, NY, USA Bing-Leonor Soares,REQUIMTE, Departamento de Quı´mica e Bioquı´mica, Faculdade deCieˆncias, Un

Joseph A CarusoUniversity of Cincinnati, Cincinnati, OH, USA

Hendrik EmonsJoint Research Centre, Geel, Belgium

Gary HieftjeIndiana University, Bloomington, IN, USA

Kiyokatsu JinnoToyohashi University of Technology, Toyohashi, Japan

Uwe KarstUniversity of Mu¨nster, Mu¨nster, Germany

Gyro¨gy Marko-VargaAstraZeneca, Lund, Sweden

Janusz PawliszynUniversity of Waterloo, Waterloo, Ont., Canada

Susan Richardson

US Environmental Protection Agency, Athens, GA, USA

Gold Nanoparticles in Analytical ChemistryComprehensive Analytical Chemistry

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Contributors to Volume 66

Marı´a Jesu´s Almendral Parra, Departamento de Quı´mica Analı´tica, Nutricio´n yBromatologı´a, University of Salamanca, Plaza de la Merced s/n, Salamanca, SpainVincenzo Amendola, Department of Chemical Sciences, University of Padova,Padova, Italy

Pedro Baptista,CIGMH, Departamento de Cieˆncias da Vida, Faculdade de Cieˆncias eTecnologia, Universidade Nova de Lisboa, Caparica, Portugal

Encarnacio´n Caballero-Dı´az, Department of Analytical Chemistry, University ofCo´rdoba, Co´rdoba, Spain

Shaowei Chen,Department of Chemistry and Biochemistry, University of California,Santa Cruz, CA, USA

Han-Wen Cheng, Department of Chemistry, State University of New York atBinghamton, Binghamton, NY, USA

Jose´ M Costa-Ferna´ndez, Department of Physical and Analytical Chemistry,University of Oviedo, Oviedo, Spain

Patricia Crespo,Instituto de Magnetismo Aplicado and Dpto Fı´sica de Materiales,Universidad Complutense de Madrid, Madrid, Spain

Elizabeth R Crew, Department of Chemistry, State University of New York atBinghamton, Binghamton, NY, USA

Jorge Ruiz Encinar,Department of Physical and Analytical Chemistry, University ofOviedo, Oviedo, Spain

Alfredo de la Escosura-Mun˜iz, Institut Catala de Nanociencia i Nanotecnologia(ICN2), Bellaterra (Barcelona), Spain

Sara Figueiredo,CIGMH, Departamento de Cieˆncias da Vida, Faculdade de Cieˆncias

e Tecnologia, Universidade Nova de Lisboa, Caparica, Portugal

Ricardo Franco,REQUIMTE, Departamento de Quı´mica, Faculdade de Cieˆncias eTecnologia, Universidade Nova de Lisboa, Caparica, Portugal

Miguel Angel Garcı´a,Instituto de Magnetismo Aplicado and Dpto Fı´sica de riales, Universidad Complutense de Madrid, Madrid, Spain

Mate-Ineˆs Gomes, REQUIMTE, Departamento de Quı´mica e Bioquı´mica, Faculdade deCieˆncias, Universidade do Porto, Porto, Portugal; Instituto de Medicina Molecular,Faculdade de Medicina da Universidade de Lisboa, Lisboa, Portugal

Yan Guo,School of Environmental Science and Engineering, Nanjing University ofInformation Science and Technology, Nanjing, Jiangsu, P R China; Department ofChemistry and Biochemistry, University of California, Santa Cruz, CA, USA

xv

Antonio Hernando,Instituto de Magnetismo Aplicado and Dpto Fı´sica de Materiales,Universidad Complutense de Madrid, Madrid, Spain

Dominik Hu¨hn,Fachbereich Physik, Philipps Universita¨t Marburg, Marburg, GermanyChristine Kranz, Institute of Analytical and Bioanalytical Chemistry, University ofUlm, Albert-Einstein-Allee 11, 89081 Ulm, Germany

Bernhard Lendl,Institute of Chemical Technologies and Analytics, Vienna University

of Technology, Vienna, Austria

A´ ngela Inmaculada Lo´pez-Lorente, Department of Analytical Chemistry, University

of Co´rdoba, Co´rdoba, Spain

Jin Luo, Department of Chemistry, State University of New York at Binghamton,Binghamton, NY, USA

Moreno Meneghetti,Department of Chemical Sciences, University of Padova, Padova,Italy

Arben Merkoc¸i,Institut Catala de Nanociencia i Nanotecnologia (ICN2), Bellaterra(Barcelona), Spain; Institucio Catalana de Recerca i Estudis Avanc¸ats (ICREA),Barcelona, Spain

Boris Mizaikoff, Institute of Analytical and Bioanalytical Chemistry, University ofUlm, Albert-Einstein-Allee 11, 89081 Ulm, Germany

Sara Sa´nchez Paradinas, Departamento de Quı´mica Analı´tica, Nutricio´n y tologı´a, University of Salamanca, Plaza de la Merced s/n, Salamanca, Spain; Institutfu¨r Physikalische Chemie und Elektrochemie, Leibniz Universita¨t Hannover,Schneiderberg, Hannover, Germany

Broma-Wolfgang J Parak, Fachbereich Physik, Philipps Universita¨t Marburg, Marburg,Germany

Lucia Pasquato,Department of Chemical and Pharmaceutical Sciences, University ofTrieste, Trieste, Italy

Miguel Peixoto de Almeida,REQUIMTE, Departamento de Quı´mica e Bioquı´mica,Faculdade de Cieˆncias, Universidade do Porto, Porto, Portugal

Paolo Pengo, Department of Chemical and Pharmaceutical Sciences, University ofTrieste, Trieste, Italy

Eula´lia Pereira,REQUIMTE, Departamento de Quı´mica e Bioquı´mica, Faculdade deCieˆncias, Universidade do Porto, Porto, Portugal

Rosario Pereiro, Department of Physical and Analytical Chemistry, University ofOviedo, Oviedo, Spain

Josefina Pons, Inorganic Chemistry Unit, Chemistry Department, Science Faculty,Autonomous University of Barcelona, Barcelona, Spain

Georg Ramer,Institute of Chemical Technologies and Analytics, Vienna University ofTechnology, Vienna, Austria

Lourdes Rivas, Institut Catala de Nanociencia i Nanotecnologia (ICN2), Bellaterra(Barcelona), Spain; Inorganic Chemistry Unit, Chemistry Department, ScienceFaculty, Autonomous University of Barcelona, Barcelona, Spain

Alfredo Sanz-Medel,Department of Physical and Analytical Chemistry, University ofOviedo, Oviedo, Spain

Shiyao Shan,Department of Chemistry, State University of New York at Binghamton,Binghamton, NY, USA

Zakiya R Skeete,Department of Chemistry, State University of New York at hamton, Binghamton, NY, USA

Bing-Leonor Soares,REQUIMTE, Departamento de Quı´mica e Bioquı´mica, Faculdade deCieˆncias, Universidade do Porto, Porto, Portugal; REQUIMTE, Departamento deQuı´mica, Faculdade de Cieˆncias e Tecnologia, Universidade Nova de Lisboa,Caparica, Portugal

Mauro Stener,Department of Chemical and Pharmaceutical Sciences, University ofTrieste, Trieste, Italy

Laura Trapiella-Alfonso,Department of Physical and Analytical Chemistry, sity of Oviedo, Oviedo, Spain

Univer-Marek Trojanowicz, Department of Chemistry, University of Warsaw, Poland andLaboratory of Nuclear Analytical Methods, Institute of Nuclear Chemistry andTechnology, Warsaw, Poland

Miguel Valca´rcel, Department of Analytical Chemistry, University of Co´rdoba,Co´rdoba, Spain

Chuan-Jian Zhong, Department of Chemistry, State University of New York atBinghamton, Binghamton, NY, USA

Contributors to Volume 66 xvii

Series Editor’s Preface

“Nanotechnology has been defined as the technology of the twenty-firstcentury, and it is expected that the broad range of nanomaterials together withtheir applications on the global market will constantly increase in the comingyears.” This sentence was written two years ago in the preface to Volume 59 ofComprehensive Analytical Chemistry, Analysis and Risk of Nanomaterials inEnvironmental and Food Samples, edited by myself and Dr M Farre

It is then obvious that there is a need for the Comprehensive AnalyticalChemistry series to look for new books in the field of nanomaterials This taskwas relatively easy In one of my regular telephone conversations with Prof.Miguel Valca´rcel, an old friend and well-known expert in analytical chemistry,

he suggested editing a book on gold nanoparticles I accepted immediately.The book that you have in your hands contains 14 chapters The first fivecover general aspects such as an introduction to analytical nanoscience andnanotechnology, the synthesis, characterization, and toxicity of gold nano-particles In the second part, gold nanoparticles are considered as target ana-lytes, with emphasis on their characterization and determination, includingspectroscopic, mass spectrometric, and separation techniques Part threedescribes the use of gold nanoparticles as analytical tools They can beincorporated in electrodes, and used as (bio)chemical sensors as well as lateralflow biosensors

With the comprehensive information on this type of nanoparticles, thismultipurpose book with novel applications in biology, the environment, andfood is a useful addition to the series and will be of great benefit to the broadnanoscience and nanotechnology community

Finally I would like to thank both editors of this book, Miguel Valca´rceland A´ ngela I Lo´pez-Lorente, for the amount of work, time, and expertise thatthey devoted to it I would like to acknowledge as well the various well-knownauthors for their contributions in compiling such a world-class and timely bookthat will be of help to newcomers, PhD students, and those senior researcherswho consider nanotechnology as one of the emerging challenges in the years tocome

D Barcelo´IDAEA-CSIC, Barcelona, and ICRA, Girona

July 10, 2014

xix

Volume Editor’s Preface

Today we are immersed in a full expansion of Nanoscience and nology (N&N) Analytical Science is an integral part of N&N since reliableinformation about the nanoworld is crucial in order to make well-foundedscientific and technical decisions in this area Two key facets of AnalyticalNanoscience and Nanotechnology (AN&N) can be noted: on the one hand, theconsideration of nanoparticles and nanostructured materials as tools for theinnovation and improvement of (bio)chemical measurement processes, and, onthe other hand, their consideration as objects (analytes) The use of nano-materials as analytical tools is the more developed field, however, the balance

Nanotech-is bound to change over the next few years due to the growing significance ofthe characterization of nanomaterials and the development of new instrumentsbased on nanotechnological approaches

Among the wide variety of nanoparticles commonly used in AN&N,namely carbon nanostructures such as carbon nanotubes, carbon dots, gra-phene, fullerenes, nanodiamonds, etc., semiconductor nanoparticles (quantumdots), or metallic nanoparticles (i.e., silver, titanium oxide, or magneticnanoparticles), this book focuses on nanoparticles of a specific nature: gold Inthis sense, the book is unique as it presents a systematic review on the differentaspects of gold nanoparticles in analytical chemistry Without doubt, goldnanoparticles are among the most relevant nanoparticles, having analyticalconnotations at a similar level to carbon nanotubes

The aim of this book is to bring gold nanoparticles closer to the readerinterested in AN&N, providing a comprehensive overview Although the focus

is on gold nanoparticles, many of the general conclusions can be extrapolated

to other nanoparticles Those professionals working not only in AN&N butalso in different fields involving the use of gold nanoparticles, such as catal-ysis, biological and medical applications, can also benefit from the book sincemany of the exceptional properties of gold nanoparticles can be applied fordifferent purposes

The 14 chapters are classified into three sections First, basic aspects ofgold nanoparticles such as their synthesis, physicochemical properties, orderivatization procedures are described in order to envisage their potential.The second part of the book reviews the techniques employed for both thecharacterization and determination of gold nanoparticles The last part isdevoted to the improvement of analytical processes by using gold nano-particles as tools in electrochemistry, spectroscopy, or biosensors

xxi

The editors wish to express their gratitude to those who have helped tobring this book to completion We would like to thank to all the authors fortheir contributions and the exhaustive revisions they have performed We alsolike to thank the cooperation of Elsevier and, for his technical support, Jose´Manuel Membrives.

Miguel Valca´rcelA´ngela I Lo´pez-Lorente

July 2014

Chapter 1

Analytical Nanoscience

and Nanotechnology

A´ngela Inmaculada Lo´pez-Lorente and Miguel Valca´rcel*

Department of Analytical Chemistry, University of Co´rdoba, Co´rdoba, Spain

*Corresponding author: E-mail: qa1meobj@uco.es

3.3 Improvement ofChromatographic andElectrophoretic

3.4 Improvement of Detection

4 Analysis of Nanoparticlesand Nanostructured Material 234.1 Information from the

4.2 Determination andCharacterization of

4.3 Microscopic Techniques 254.4 Separation Techniques 26Gold Nanoparticles in Analytical Chemistry http://dx.doi.org/10.1016/B978-0-444-63285-2.00001-8

Nanosciencehas multiple complementary definitions, such as “the science

of the synthesis, analysis and manipulation of materials at atomic, molecular,and macromolecular scales where physico-chemical properties may differsignificantly from those at a larger particulate scale,” [1] or, simply: “thescience based on the diverse structures of materials which have dimensions of

a billionth part of the meter”[2]

On the other hand, Nanotechnology “deals with the design, tion, production and application of structures, devices and systems by con-trolling the shape and size at the nanometer scale”[1]

characteriza-A substantial aspect of nanoscience and nanotechnology (N&N) is itsmultidisciplinary as well as transversal and convergent character Physicists,chemists, and engineers are the scientists and professionals more directlyinvolved, but their convergence with other areas such as information tech-nology and communication, biotechnology, and materials science, in a firstapproach, and medicine, pharmacy, agrifood, and diverse types of industriessuch as textile or energetic, in another, has to be pointed out

Analytical science cannot be left out of N&N[3]and, in fact, it is even present

in many definitions of N&N since reliable information about the nanoworld iscrucial to make well founded scientific and technical decisions in this area.Words belonging to the analytical discipline such as “analysis” or “character-ization” and others shared with other disciplines such as “use” or “employment”summarize the two key facets of the relationship between analytical chemistryand nanoscience and nanotechnology, namely, (1) the consideration of nano-particles and nanostructured materials as objects (analytes) or (2) tools for theinnovation and improvement of the (bio) chemical measurements processes.The major application areas of nanotechnology can be classified into fourgroups[3], namely, (1) nanobiotechnology and nanomedicine, (2) nanomaterials,(3) nanoelectronics, and (4) nanosensors/nanodevices, nanotechnologicalinstrumentation, and nanometrology The last area is directly related to analyticalscience, which also plays an essential role in the other three, for example, dealingwith the monitoring of production processes or both the characterization and use

of end products

1.2 Classifications

There are several emerging possibilities when introducing nanoscience andnanotechnology in the analytical scope Therefore, a multiple classificationbased on four complementary criteria has been created, which is shownschematically inFigure 1and is described in the following text

The first criterion (Figure 1(1)) considers the type of material analyzed,which can be conventional (macro or micro in size) or nanomaterials In thefirst case, nanoparticles can be involved in the analytical process, conferring to

it nanotechnological character An example is the use of quantum dots tionalized with antibodies, which can be injected in organisms in order todetect carcinogenic processes[4] In the second possibility, the target is theown nanoworld, which coincides with the consideration of nanomaterials asanalytes For example, the determination of nanomaterials such as goldnanoparticles [5] or carbon nanotubes [6e8] from environmental and bio-logical matrices[9]

func-The second criterion (Figure 1(2)) relies on the analytical consideration ofnanoparticles and nanostructured materials as objects (analytes) or toolsinvolved in the analytical process The extraction of chemical information

Nanoworld

Detection/quantification

of nanomaterials

TARGET OF THE ANALYSIS

ANALYTICAL NANOSCIENCE AND NANOTECHNOLOGY

NANOPARTICLES AND NANOSTRUCTURED MATERIAL

Macroworld Microworld

Nanomaterials as

ANALYTES

Nanometric analytical systems

Analytical nanosystems

Nanotechnological analytical systems

EXPLOITATION OF NANOMATTER PROPERTIES

of the nanosize; and (4) exploitation of the nanomatter properties.

Analytical Nanoscience and Nanotechnology Chapter j 1 5

from the structured nanomaterials (composition, chirality, reactivity, etc.) is anindispensable complement to the physical characterization, which is morewell-known (dimensions, topography, etc.) [10] On the other hand, nano-materials can be used as analytical tools in order to develop new analyticalprocesses or to improve existing ones (i.e., development of optical sensors,development of stationary and pseudostationary phases in chromatography andcapillary electrophoresis, mechanical sensors, etc.).

Criteria 3 and 4 (Figure 1(3) and (4)) are based on exploitation in theanalytical scope of the exceptional properties of nanomaterials, inexploiting the nanosize, or both This leads to the definition of three types

of analytical systems related to nanoscience and nanotechnology: technological analytical systems, nanometric analytical systems, andanalytical nanosystems [11] Nanotechnological analytical systems exploitthe exceptional physico-chemical properties of nanomaterials (althoughthey are in micro/macro analytical systems) accounting for the most currentuses of analytical nanoscience Nanometric analytical systems, which arebased exclusively on the nanosize of the devices involved, are exemplified

nano-by nanochip liquid chromatography systems[12] exploiting the advantages

of working with flow rates as low as a few nanolitres per minute, ananopipette [13], or levitated nanodrops as analytical containers [14].Finally, analytical nanosystems successfully integrate the previous twotypes of systems by exploiting both the nanosize and nanomaterialsproperties (e.g., individual carbon nanotubes for use as electrodes [15],supramolecular systems that selectively recognize an analyte [16], and theso-called lab-on-a-particle [17])

1.3 Synthesis of Nanoparticles

Nanomaterials can exist in the environment from a natural source, such asorganic colloids, magnetite, aerosols, iron oxides, etc The nanotechnologicalrevolution is posed in a change of paradigm in the fabrication of products Twoapproaches can be used to raise nanosize, namely, (1) “top-down” strategies,based on methodologies which achieve nanosize materials from macro-materials (nanoparticles are directly generated from bulk materials via thegeneration of isolated atoms usually involving physical methods such asmilling or attrition, repeated quenching and photolithography [18]) and (2)

“bottom-up” strategies, based on the creation of complex nanostructures fromatomic or molecular functional elements They comprise molecular compo-nents as starting materials linked with chemical reactions, nucleation, andgrowth processes to promote the formation of clusters Numerous kinds ofnanoparticles have been produced by liquid-phase synthesis, using techniquessuch as co-precipitation of sparingly soluble products by addition, exchange,and reduction reactions, oxidation, hydrolysis [19], solegel processing [20],microemulsions[21], etc The latter approach is generally considered to be far

more promising due to the higher level of control offered.Figure 2shows ascheme of the different strategies (“top-down” and “bottom-up”) used toachieve the nanoscale.

1.4 Types of Nanoparticles

According to the IUPAC Glossary, a nanoparticle is a microscopic particlewhose size is measured in nanometers, often restricted to so-called nanosizedparticles (NPs; < 100 nm in aerodynamic diameter), also called ultrafineparticles [22] With the expected increase in the applications of nanotech-nology, more and more products will be manufactured containing componentswhich will fit the commonly used definition of the nanoscale, as having a size

between approximately one and 100 nm These wide varieties of structures have been classified in multiple ways in the literature.Figure 3shows the most relevant types of nanostructures in analytical nano-science and nanotechnology, classified according to four nonexclusive criteria.Nanoparticles can be classified as natural, anthropogenic (incidental), orengineered in origin[9] (Figure 3(1)).

nano-From a practical point of view, it is important to know the homogeneity ofthe nanostructured materials both for scientific studies as well as for industrialapplications Homogeneity can be referred to in terms of chemical composi-tion or dimensionality (Figure 3(2)) Identical nanoparticles are those with thesame chemical composition and dimensions On the contrary, nanoparticleswith the same chemical composition but different dimensions usually presentdifferent properties

Concerning the nature or chemical composition of nanostructures, thosecan be classified (Figure 3(3)) as inorganic (e.g., noble metal nanoparticles,quantum dots, etc.), organic (fullerenes, carbon nanotubes, dendrimers,molecular imprinted polymers, etc.) or mixed (gold nanoparticles modifiedwith calixarenes, carbon nanotubes functionalized with ferrocene, etc.) In thiscontext, there is a growing interest in the development of hybrid nanoparticles,which can be defined as well-organized nanomaterials consisting of two ormore types of individual nanocomponents[23]

Nanoscale in ZERO dimensions

Nanoscale in ONE dimension

Nanoscale in TWO dimensions

Nanoscale in THREE dimensions

Inorganic Organic

FIGURE 3 Classification of nanostructures according to their origin (1), homogeneity (2), nature (3), and dimensionality (4).

The last classification of nanomaterials is based on dimensionality criteria(Figure 3(4)) As shown in the figure, two classifications may be done, takinginto account both the strict dimensions (in the nanoscale) of the nanostructurethat give rise to those exceptional properties and the dimensions of thematerial where nanostructures are present.

The Royal Society of Chemistry and the Royal Academy of Engineeringclassified nanostructures in function of the number of dimensions in thenanoscale (below 100 nm)[24], distinguishing three types of nanostructures:(1) nanoscale in one dimension, such as surfaces with nanometric thickness(e.g., graphene sheets); (2) nanoscale in two dimensions, such as carbonnanotubes, inorganic nanotubes, nanowires, etc.; (3) nanoscale in threedimensions, which includes metallic nanoparticles and their oxides, quantumdots, fullerenes, and dendrimers Classification of nanoscale at zero dimensioncan also be added, such as materials composed by dispersed nanoparticles.Other authors[25]have classified nanostructures depending on the number

of dimensions which exceed 100 nm, being above the nanoscale, structures being thus categorized as 0D, 1D, 2D, or 3D A 0D nanostructure is

nano-a mnano-aterinano-al with nano-all its dimensions comprised in the nnano-anometric scnano-ale (e.g.,metallic nanoparticles, quantum dots, etc.) Carbon nanotubes are an example

of 1D nanostructures, which have one dimension of micro/macrometric size,such as nanowires or nanorods 2D nanostructures have two dimensions abovenanoscale while one of them is below 100 nm That is the case of surfacenanocoatings or thin films of molecular monolayers Finally, 3D nano-structures are those whose three dimensions escape from the nanoscale, but thematerial is comprised by a set of nanoparticles forming a block of micro/macrometric size (e.g., nanoporous materials, powders)

We will focus on engineered nanoparticles, which can be classified cording to their nature as organic, inorganic, or hybrid

ac-1.4.1 Organic Nanoparticles

1.4.1.1 Carbon Nanomaterials

Graphitic forms include 0D fullerene, 1D CNT, and 3D graphite, and the 2Dcase comes to the graphene, a single layer of carbon atoms formed in ahoneycomb lattice

Graphene is an open, flat, two dimensional structure composed of carbonatoms organized in a network of hexagons attached to each other This ispossibly a result of sp2hybridization of the carbon atoms present in the sheet

It has a large specific surface area and can be easily modified with functionalgroups, especially via graphene oxide Graphene quantum dots (GQDs), a newkind of quantum dots, have emerged and ignited tremendous research interest.GQDs are defined as graphene sheets with lateral dimensions less than 100 nm

in single, double, and few (3 to<10) layers GQDs show low cytotoxicity,excellent solubility, chemical inertia, stable photoluminescence, and better

Analytical Nanoscience and Nanotechnology Chapter j 1 9

surface grafting Therefore, they are promising in optoelectronic devices,sensors, bioimaging, etc.[26].

Another two examples of 0D carbon-based fluorescent nanomaterials arediamond nanocrystals (DNs) and carbon dots (CDs), which have also drawnmuch attention in recent years In general, DNs consist of about 98% carbonwith residual hydrogen, oxygen, and nitrogen; they possess a sp3hybridizedcore and have small amounts of graphitic carbon on the surface LuminescentCDs comprise discrete, quasispherical carbon nanoparticles with sizes below

10 nm

Fullerenes are closed-cage carbon molecules containing pentagonal andhexagonal rings They comprise a wide range of isomers and homologousseries, from the most studied C60and C70to the so-called higher fullerenes like

C240, C540, and C720 They possess relatively high electron affinity, phobic surface, and high surface/volume ratio

hydro-Carbon nanotubes have received special attention ever since their discovery

by Iijima in 1991 [27] They are tubular in shape and consist entirely ofcovalently bonded carbon atoms They can be described as hollow graphiticnanomaterials comprising one (single-walled carbon nanotubes, SWNTs), two(double-walled carbon nanotubes, DWNTs) or multiple (multiwalled carbonnanotubes, MWNTs) layers of graphene sheets They possess nonpolar bondsand high aspect ratios, which make them insoluble in water and facilitate theiraggregation

Carbon nano-onions (CNOs) are nanoparticles with a diameter between 20and 100 nm composed of several layers of concentric fullerenes and constitutethe spherical version of MWNTs They were described by Ugarte[28]in 1992,although few applications have been reported

Moreover, carbon nanofibers (CNFs) are solid carbon fibers with lengths inthe order of a few microns and diameters below 100 nm CNFs differ fromCNTs in the absence of a hollow cavity, and the diameters of CNFs aregenerally higher than those of the corresponding CNTs[29]

Finally, carbon nanocones were first synthesized by vapor condensation ofcarbon atoms on a graphite substrate[30] The disclination of each structurecorresponds to the presence of a given number of pentagons in the seed fromwhich it grew: disks (no pentagons), five types of cones (one to five penta-gons), and open tubes (six pentagons) The unique electronic distribution,which is provided by these pentagonal rings to the carbon nanocones, results inenhanced local density at the cone apex One major class of cone structures aresingle-walled carbon nanohorns (SWNHs) with the narrowest opening anglewith five pentagonal rings in their apex (Figure 4)

1.4.1.2 Other Organic Nanomaterials

Organic polymeric nanoparticles are prepared from polymers and consideredpotential drug delivery devices Dendrimers are one kind of polymeric nano-particles constructed by the successive addition of layers of branching groups

Molecularly imprinted polymers (MIPs) are a specific class of selectivesorbents that have built-in functionality to achieve the complementaryrecognition of a given chemical compound[31] Although MIP particles can

be nanometric in size, they usually fall in the micrometric range

In addition, an increasing number of diverse nanomaterials are emergingfor biopharmaceutical applications, such as liposomes, nanomicelles, nano-vesicles nanoemulsions, etc.[32]

Quantum dots (QDs) are semiconductor nanocrystals with all threedimensions falling in the 1e10 nm size range In many respects, these lumi-nescent nanocrystals constitute a transitional stage between bulk semi-conductors and single atoms The QD core is made up of elements from theIIeVI (e.g., CdSe, CdTe, CdS, and ZnSe), IIIeV (e.g., InP and InAs), orIVeVI (e.g., PbSe) group [33] They have aroused widespread interest byvirtue of their exceptional optical, electronic, electrochemical, photophysical,redox, and catalytic properties

Nanoparticulate metal oxides are widely used, such as TiO2, Al2O3,ZrO2, MnO and CeO2, as well as nanoparticles of iron oxides (FeOx).Attention should be paid to the increasingly used magnetic nanoparticles,which have been synthesized with a number of different compositions andphases, including iron oxides, such as Fe3O4and g-Fe2O3, pure metals, such

as Fe and Co, and spinel-type ferromagnets, as well as alloys [34] over, silica nanoparticles (SiO2) are characterized by presenting high surfaceareas and exhibit intrinsic surface reactivity, which allows chemical modi-fications[10]

More-FIGURE 4 Main types of carbon-based nanomaterials Adapted with permission from Ref [29]

Analytical Nanoscience and Nanotechnology Chapter j 1 11

Nano-size zeolite, clays, and ceramics are other nanoparticles that havebeen proposed for various applications[35](Figure 5).

1.4.3 Hybrid Nanoparticles

Hybrid nanoparticles can be defined as well-organized nanomaterials sisting of two or more types of individual nanocomponents [23] Thosenanocompounds can be bound via organic/inorganic molecular bridges ordirectly attached to one another Apart from the special type of bonding be-tween the nanocompounds, the exceptional properties of hybrid nanoparticlesare due to the highly organized arrangement of the nanoconstituents.Nanoparticles possess excellent properties that can be boosted or supple-mented by combining two or more types of materials into a hybrid nano-composite In general, hybrid nanoparticles can be classified into two differenttypes according to the combined properties[23], namely, (1) properties of theisolated nanoparticles are different but complementary, and (2) properties ofthe isolated nanoparticles are of the same nature, but their combinationproduces important synergistic effects

con-1.5 Properties of Nanoparticles

The “nanoscale” has introduced a new scenario where impressive changes inphysico-chemical principles, laws, and properties are observed as regardsmacro- and micro-materials

Two especially significant differences have been described: the surface/volume ratios and chemical reactivities of nanostructured matter in compari-son to macro- and micro-scale matter In addition, quantum effects areenhanced by effect of the configuration of molecular orbitals (similar to that ofatomic orbitals) and, as a consequence, chemical, optical, electrical, thermal,and magnetic characteristics are unique on the nanoscale Such changes haveshaped the impact of nanotechnology in a number of scientific, technical, andindustrial fields

images of model inorganic nanoparticles, namely,

Ag, Au, TiO 2 , Fe 3 O 4 , CeO 2 , and NPs Adapted with

permission from Ref [36]

2 INTRODUCTION TO ANALYTICAL NANOSCIENCE

AND NANOTECHNOLOGY

2.1 Facets of Analytical Nanoscience and Nanotechnology

Generally, nanomaterials in the analytical nanoscience and nanotechnology(AN&N) scope are considered as analytical objects or analytical tools.Analytical nanoscience and nanotechnology currently provide one of the mostpromising avenues for developments in analytical science, derived from theirtwo main fields of action, namely the analysis of nanostructured materials andtheir use as tangible tools [37] The use of nanotechnological tools inanalytical methods can improve the analytical properties and enables thedevelopment of new types of analysis On the other hand, analysis of thenanoworld is an issue of analytical chemistry

A third classification, as an interface between the two previous, can beintroduced, which is the use of nanomaterials in analytical processes for thecharacterization and/or determination of other nanomaterials, seeFigure 6.Examples of this facet are, for instance, the use of silver or gold nanoparticles

as SERS substrates for the determination of other nanomaterials (e.g., carbonnanotubes[6]), or the use of membranes composed by multiwalled carbonnanotubes for the preconcentration and determination of single-walledcarbon nanotubes[7]

2.2 Types of Analytical Systems

The use of nanomaterials as analytical tools is one of the facets of analyticalnanoscience and nanotechnology Nanotechnology-based analytical processes

2 3

Use of nanomaterials in analytical processes of characterization and determination of nanoparticles

FIGURE 6 Different facets of nanoparticles in analytical nanoscience and nanotechnology (AN&N) scope.

Analytical Nanoscience and Nanotechnology Chapter j 1 13

can exploit both the nano size and exceptional properties of structured matter In this sense, as it was previously introduced (seeFigure 1and relatedtext), analytical systems based on nanoscience and nanotechnology can beclassified into three types[11]:

nano-1 Nanometric analytical systemsare based on nanosize and/or nanofluidics,for example, nanochip liquid chromatography However, some authorshave placed them outside the scope of nanotechnology

2 Nanotechnological analytical systems exploit the exceptional chemical properties of nanomaterials

physico-3 Analytical nanosystems integrate the previous two systems, for example,individual carbon nanotubes for use as electrodes (Figure 7)

2.2.1 Nanometric Analytical Systems

As previously defined, nanometric analytical systems are analytical systemsthat take advantage of the fact that some technical characteristics or elements

of the analytical process have nanometric size of flow Thus, their foundation

is not based on the new physico-chemical scenario imposed by nanomaterialsand their exceptional physical and chemical properties They can be consid-ered as a trend of miniaturization

They cannot be completely considered inside the scope of nanotechnology,although because of the undoubted advantages they offer, many authorsinclude them Some representative examples of this kind of systems arereported as follows

Some nanometric analytical systems are based on nanometric volumesemployed Such is the case of the use of levitated nanodrops[14] This strategyhas been employed for online monitoring of chemical reactions in

Interfaces

ANALYTICAL NANOSYSTEMS (ideals)

Characteriscs

exploited in the

analycal context

NANOMETRIC ANALYTICAL SYSTEMS

NANOTECHNOLOGICAL ANALYTICAL SYSTEMS

Nanometric size

Exceponal properes of nanomaterials

ultrasonically levitated, nanoliter-size droplets by Raman spectroscopy Aflow-through microdispenser connected to an automated flow injection systemwas used to dose picoliter droplets into the node of an ultrasonic trap Asequence of reagents can be injected via the microdispenser into the levitateddroplet Thus, chemical reactions can be carried out and monitored online Thedroplet suspended in the air enables the removal of interferences from therecipient and a rapid evaporation of the solvent This has eased the directanalysis with Raman microscopy[14]as well as the development of interfaces

in direct couplings in which sample solvent is a problem, such as in assisted laser desorption/ionization time of fight mass spectrometry(MALDI-TOF-MS)[38]

matrix-The chromatographic systems called “Nano-LC” are based on the use ofliquid mobile phases with a flow rate of a few nanolitres per minute[39], incontrast with conventional ones in the range of microliters per minute Thatimplies a new design for the chromatography instrument, but it offers ad-vantages such as ease of use, both the delay time for the gradient to arrive atthe head of the column and the dead volume between the separation columnand the ESI tip are minimized, resulting in a shorter running time and reducedband broadening

The so-called “nanopipettes” [13] are integrated, carbon-based pipetteswith nanoscale dimensions that can probe cells with minimal intrusion, injectfluids into the cells, and concurrently carry out electrical measurements Theyallow the direct transference of nanovolumes in the range of nano/picolitersbetween micro/nanometric compartments This transference is based on theelectro-osmotic phenomena Their applications in the field of biotechnologyare very promising, as well as in high-throughput analysis, key in the field ofthe pharmacological industry to perform multisynthetic experiments in thedevelopment of new drugs

2.2.2 Nanotechnological Analytical Systems

Nanotechnological analytical systems exploit the exceptional properties of thenanometric materials (nanoparticles and nanostructured materials) withanalytical purposes They posses micrometric dimensions, but once theirdimensions can be reduced into the nanoscale, they could be called “analyticalnanosystems.”

In these systems, there is a micro-element which acts as a bridge betweenthe nanocomponents and the signal transduction system through a simplephysical bond The active nanocomponent is responsible for the interactionand produces the analytical signal

One illustrative example is micro-electromechanical systems cantilevers”)[40,41], which are based on the immobilization of a biochemicalreceptor in a probe of nanometric dimensions in a micromechanical siliconsystem that is sensitive to the environment, including chemical compounds and

(“nano-Analytical Nanoscience and Nanotechnology Chapter j 1 15

biological entities which affect their mechanical characteristics in such amanner that the change can be measured in terms of electrical or opticalproperties[5] Although a micro-electromechanical system is micrometric insize, its sharp tip, which allows interaction forces at the atomic scale to besensed, is a nanometric device Their most remarkable advantages are theirreduced dimensions, portability (in some cases), and their high levels ofsensitivity and selectivity for the determination of a wide variety of analytes.Their scope is defined by the biochemical interaction analyte-receptor.Other examples of nanotechnological analytical systems are the field effecttransistors (FETs), which have been revitalized by the introduction of nano-structured materials[41] Such is the case of the use of semiconducting carbonnanotubes or a network of nanotubes placed between two electrodes ofmicrometric dimensions Carbon nanotubes are derivatized in order to achieve

a more specific interaction with the analyte When it is retained, a change inthe electronic properties of the carbon nanotubes is originated, which leads to

a variation in the potential between the electrodes, used as an analytical signal.Semiconducting carbon nanotubes have been employed for the determination

of a wide variety of gaseous analytes, such as NH3, CO, and CO2 amongothers because they exhibit larges changes in electric conductivity in theirpresence[42]

Finally, another example of this type of system is a nano-electrode, sisting of a single multiwalled carbon nanotube (2e3 mm in length and 30 nm

con-of diameter) bonded to the end con-of an etched tungsten tip[15], which acts as abridge This is a real nano-electrode since it has nanometric dimensions andexploits the exceptional properties of nanomatter, although it is coupled to amicrometric device, the tungsten tip The carbon nanotube surface can befunctionalized with a wide variety of (bio) chemical compounds in order toimprove the selectivity and the field of application It offers the advantage ofcompatibility with micro/nano-size samples (e.g., cells) This nano-electrodehas been used to determine dopamine and glutamate (immobilizing gluta-mate oxidase enzyme in CNT surface) in a physiological medium with aquantification limit of 100 mm

2.2.3 Analytical Nanosystems

These systems can be defined as instruments or devices that have a nanometricsize and are controlled by the physico-chemical laws of Nanoscience Inaddition to the nanometric size, the exceptional properties of nanomatter arealso exploited This is an ideal situation, since microcomponents are needed toconnect the nanoworld with the macroworld[3]

A nanochemical approach to a real analytical nanosystem is the so-called

“lab-on-a-particle” [17], which aims to emulate the micrometric scale velopments (“lab-on-a-chip”) This nanosystem consists on a supramolecularnanoarchitecture that incorporates chemical entities that can be employed as a

de-“gate” to allow controlled access to a specific site on the supramolecular

complex An example is mesoporous silica nanoparticles functionalized withswitchable molecules, whose inner pores can be used to entrap chemicalspecies The gate will open upon the application of physico-chemical externalimpulses, such as photochemical or electrochemical, and can release confinedguests or allow molecular species of the solution to be incorporated.

2.3 Evolution and Limit of Analytical Nanoscience

and Nanotechnology

Nanoscience and nanotechnology unarguably have promising prospects

A report of the US National Science Foundation [43] predicted that a newrevolution based on the bio-nano-info triangle will shortly surpass the presentevolution of the computer-info binomial

The scientific and industrial impact of nanoscience and nanotechnologyhas grown dramatically in recent years The exponential growth of the number

of papers on this topic published in the last 10 years in the SciFinder and thevast amount of economic resources invested in industrial technologicaldevelopments each year[44]constitute the best support for the brilliant presentand promising future of nanoscience and nanotechnology It should be pointedout that there are more than one million scientific papers published on thistopic (Figure 8)

The impact of nanotechnology leaves no shadow of doubt Hassan [45]emphasized that the most crucial aspect in this respect is the small things-bigchanges binomial Analytical chemistry should play a major role in the comingadvances in N&N, particularly in these aspects: (1) the environmental andtoxicological impacts of nanotechnology, (2) the need for homogeneous, pure,

FIGURE 8 Scientific and industrial evolution of nanoscience and nanotechnology: (left) number

of publications per year, (right) millions of euros involved in European nanotechnological industry.

Analytical Nanoscience and Nanotechnology Chapter j 1 17

well-characterized nanoparticles, (3) the need to address nanometrology, and(4) nanomedicine, in the development of biomeasurement nanosystems andthe characterization and monitoring of nanostructured pharmaceuticals[3].Regarding analytical nanoscience and nanotechnology, the use of nano-materials as analytical tools is the most developed field Although more thanone half of reported applications relate to the use of nanoparticles, the balance

is bound to change over the next few years due to the growing significance ofthe characterization of nanomaterials and the development of new instrumentsbased on nanotechnological approaches (Figure 9)

Nowadays, the prefix “nano” is opening many doors, which leads to manyauthors misusing it to their advantage The prefix “nano” should be used inconnection to nanomaterials, nanostructures, and nanodevices that exploit notonly the size but also the exceptional properties of the nanoworld The samehappened in the past with other overexploited keywords such as “sensor.”Fortunately, this terminological abuse has vanished over time

2.4 Ethical and Social Implications

Nanotechnology has been deemed as a key emerging technology for fulfillingthe “grand challenges of our time” (Lund Declaration [46]) in areas such ashealth care, energy production, environmental protection, and potable waterprocurement However, it shows two contradictory connotations, namely, (1)the production of new materials with outstanding mechanical, optical, electric,and magnetic properties for a wide variety of uses, which is highly positive,and (2) its uncertain effects on human health and the environment, which ishighly negative[47]

There are two major deficiencies regarding the impact of nanotechnology.One is that the risks and ethical implications of nanotechnology in the pro-duction and industrial domains have not been considered The other is that,although the potential hazards of nanotechnological products has been

FIGURE 9 Present situation and trends in the two facets of analytical nanoscience and technology: nanomaterials as analytical tools and analysis of the nanoworld.

nano-described, their toxicity remains uncertain due to the lack of scientificconsensus in many cases[48].

Nano-ethics, which is the term used to designate ethics in the development

of nanosize objects, comprises three different fields of study (the first twoare closely related): ecotoxicity, nanotoxicity, and violation of privacy.Ecotoxicity is related to the potential environmental damage of nanomaterials,whereas nanotoxicity focuses on the effects of nanoparticles on health.Assessing the risks of nanomaterials is especially important in nanomedicine,where they are used to prevent, diagnose, and treat specific diseases[49] Theviolation of privacy field is associated with the production of nanosensors Thedevelopment of new communication technologies may result in an increasinginvasion of privacy and violation of personal rights[47]

Government’s concern about this issue is raised, as well as of variousprestigious scientific and technical organizations No doubt that this is apending issue of nanotechnology, which is the key for future consolidation.Recently, an integral approach to the social responsibility of nanoscience andnanotechnology [47] has been reported, which proposes a framework forconducting responsible research in nanoscience

3 USE OF NANOPARTICLES AS TOOLS IN ANALYTICAL PROCESSES

3.1 Objectives

One of the main goals of analytical chemistry is the development of newmethodologies which improve existing ones and meet new demands of (bio)chemical information posed by the present social and economic problems Thegrowing demand of (bio) chemical information requires the development ofnew tangible and intangible tools to support analytical processes Improve-ments in analytical processes can be measured in terms of analytical proper-ties: capital, basic, and productivity-related (seeFigure 10)

The new scene created by the growth of nanoscience and nanotechnologyhas had an impact in the field of analytical chemistry Nanotechnological toolshave been used in analytical methods by exploiting the excellent properties ofnanoparticles in order to improve well-established analytical methods or todevelop others for new analytes or matrices The use of nanoparticles shouldlead to improved selectivity, sensitivity, rapidity, miniaturizability, or porta-bility of the analytical system This facet of analytical nanotechnology is themost extensively explored[3]

The most widely used nanoparticles in analytical sciences include (1) silicananoparticles, (2) metallic nanoparticles (quantum dots, gold nanoparticles,etc.), (3) carbon nanoparticles (mainly fullerenes and carbon nanotubes), (4)organic polymer nanoparticles (e.g., molecular imprinted polymers), and (5)supramolecular aggregates (nanomicelles, nanovesicles), as depicted in

Analytical Nanoscience and Nanotechnology Chapter j 1 19

Figure 11 The explored nanoparticle property can be electrical, optical,thermal, magnetic, or chemical Frequently, however, two or more propertiesare explored at once.

An analytical process can be divided into several steps, as represented inFigure 11, namely, sample preparation (including sampling), chromatographic(LC, GC) and capillary electrophoretic techniques, detection and data handlingand treatment to offer the results as required The role of nanoparticles differsbetween the different steps of the analytical process Figure 11 shows theproportion of described analytical procedures in which nanoparticles areinvolved

3.2 Sample Treatment: Purification and Preconcentration

of Analytes

Sample preparation has also profited from the use of nanomaterials, albeit to alesser extent than detection The incorporation of nanoparticles in the sampletreatment step, in general, helps to simplify it For example, conventionalsorbents for solid-phase extraction and solid-phase micro-extraction have beenreplaced by nanomaterials Nanoparticles, according to their participation androle in the sample treatment step can be classified as follows[10]: (1) nano-particles acting as sorbent agents[50], where direct interaction between theanalyte and the nanoparticle takes place; (2) nanoparticles acting as an inert

Address new demands for information

B.4 A

NANOSCIENCE &

NANOTECHNOLOGY

Increase of analycal properes

Capital Basic Productivity-related

ANAL YTICAL CHEMISTR

Y

FIGURE 10 The multiple roles of nanotechnology in analytical science: (A) direct impact, and (B) indirect impact through their role as support for other scientific-technical advances.

FIGURE 11 The role of nanoparticles in different steps of the analytical process.

support, such as a silica nanoparticle functionalized with a complexationagent; (3) nanoparticles having special magnetic properties, which can eitherdirectly adsorb the analyte or can be functionalized with organic groups (theuse of a magnetic field can simplify the analytical procedure); and (4) nano-particles acting as ionization agents for the direct analysis of samples by ionsecondary mass spectrometry.

Carbon nanomaterials have become the focus of attention thanks to theirsingular p-p electron configuration, as well as metal oxides by virtue of theirhigh surface area In this sense, carbon nanotubes have been widely used assorbents for solid-phase extraction[51] Packed nanotubes tend to aggregate tosome extent, their inclusion as a nanoscience application requires thataggregation be avoided, and interactions between analytes and isolatednanoparticles be favored [52] Conical carbon nanoparticles, such as single-walled nanohorns (SWNHs) [53]or carbon nanocones/disks [54] have beenalso used for solid-phase (micro) extraction In addition to solid-phaseextraction, nanoparticles have been used in other sample treatments such asmembrane filtration, for example, with membranes composed of or modified

So far, nanoparticles have found extensive use in capillary electrophoresis(CE), packed capillary electrochromatography (CEC), open tubular CECformats, and microchip CE Several nanoparticles have been employed aspseudostationary phases in capillary electrophoresis [58], such as SWNHs,CNTs, and fullerenes [59] or graphene [60] A new electrophoretic modedesignated as micellar nanoparticle dispersion electrokinetic chromatography(MiNDEKC) has been developed which proved to be effective in the separa-tion of aromatic compounds [61]or even chiral compounds [62]

There are also some reports of the use of nanoparticles in LC, ion matography (IC), and GC Nanoparticles for GC are usually packed into atubular column As an alternative to packing, CNTs can be self-assembled into

chro-a gchro-as chromchro-atogrchro-aphic column [63] NPs have been also inserted into amonolithic column in LC[64]

3.4 Improvement of Detection Processes

Detection is the analytical step in which nanoparticles have been most widelyused by virtue of their ability to replace conventional materials, as well as theadvantages of electrochemical biosensors In this sense, biomolecules arestabilized by the nanoparticles, which increase active surfaces and facilitateelectron transfer In addition, the development of optical nanosensors has beenfostered by the exceptional optical properties of metallic nanoparticles derivedfrom their plasmon resonance, as well as photofluorescent properties in thecase of semiconducting nanoparticles (quantum dots)[65], carbon dots [66](CDs), or graphene quantum dots[67]

Quantum dot-based sensors for chemical and biological detection havebeen widely developed [68] by virtue of the special optical and electronicproperties of the component QDs plus the possibility to functionalize themrelatively easily with a wide variety of biological as well as relevant moleculesfor other important applications[69]

Gold nanoparticles have been also widely used by virtue of their opticalproperties For example, mercury traces have been detected by ultra-violetevisible spectroscopy using gold nanoparticles functionalized withcomplementary DNA sequences, which are intensely colored and are present

as colloidal dispersion[70]

Carbon nanotubes and gold nanoparticles are the types of nanoparticlesmost widely used for developing electrochemical (bio) sensors [71].CNT-based sensors can be used to detect changes in their electronic propertiesresulting from the sorption of molecules on their surface The electronicproperties of CNTs encourage their use as electrodes to mediate electro-netransfer reactions with electroactive species in solution [72] Combininggold nanoparticles and carbon nanotubes was found to enhance some elec-trocatalytic properties of electrodes[73]

4 ANALYSIS OF NANOPARTICLES

AND NANOSTRUCTURED MATERIAL

4.1 Information from the Nanoworld

The development of analytical methodologies for extracting quality tion from nanoparticles or nanostructured compounds is of high interest due tothe poor knowledge available about their actual composition and character-istics, as well as their presence in environmental, biological, or agrifoodmatrices Developing effective analytical methods for the rapid, accuratecharacterization of nanoparticles is mandatory with a view to advancingnanotechnology[3]

informa-This topic has clear connotations of interface between physics andchemistry, as can be inferred from Figure 12, where the physical (size,properties, and topography) and chemical characteristics (composition,

Analytical Nanoscience and Nanotechnology Chapter j 1 23

chirality, reactivity, and type of bond) are reflected It should be pointed out inthis context that there is an increasing interest in biological characteristics(diffusion through membranes, toxicity, biotransformations, nanoparticlesinteraction with cells and microorganisms) derived from the chemical reac-tivity of the nanomatter Among the types of characterization that are needed,perhaps none is more challenging than chemical characterization[74].Extracting information from the nanoworld entails measuring dimensions

of only a few nanometers So-called nanometrology is, therefore, a hot topic.Although nanometrology is of special concern to physicists, analyticalchemists can make substantial contributions to its advancement

Several types of techniques are available to characterize NPs[75]and theirsurface chemistry[76] Herein, the different techniques have been classified asmicroscopic, spectroscopic, and separation

4.2 Determination and Characterization of Nanoparticles

As can be seen in Figure 13 characterization encompasses determination,which has analytes as targets Objects or systems are characterized, beingrelated with knowledge in the primary data-information (provided byanalytes)-knowledge hierarchy In the analytical-chemical context, informa-tion is related to qualitative and quantitative results, while knowledge is relatedwith reports Characterization can be defined as the identification and/orquantification of specific properties or characteristics of the sample or material[77] On the other hand, determination can be considered as the application ofthe complete analytical process for detecting, identifying, and quantifying ananalytical parameter[78]

FIGURE 12 Types of information about nanomatter according to the nature of its characteristics.

4.3 Microscopic Techniques

Microscopic techniques include optical, electron, and scanning probe scopy The most used techniques for the characterization of NPs are electronmicroscopies (EM), transmission electron microscopy (TEM), high-resolutionTEM, and scanning electron microscopy (SEM), and scanning probemicroscopies (SPM), namely, atomic force microscopy (AFM) and scanningtunneling microscopy (STM) Depending on the technique, resolutions down

micro-to the subnanometer range can be achieved Besides the imaging of particles, these methods enable the determination of the aggregation, disper-sion, size, structure, and shape of NPs In addition, SEM and AFM offerthree-dimensional images of nanoparticles

nano-However, microscopic techniques present shortcomings in sample ration as well as the statistical uncertainty due to the human subjectivityimplied when deciding which parts of the grid are photographed In order tomeasure an accurate size distribution of nanoparticles, it is necessary to countand measure thousands of particles in order to obtain reliable counting sta-tistics Regarding sample preparation, SEM and TEM have to operate in avacuum and, therefore, only dry, solid samples can be investigated Thetransfer of the sample from dispersion to dried state changes the size distri-bution, as well as the aggregation state of the nanoparticles, or leads to theprecipitation of salts Furthermore, electron microscopy is usually a destruc-tive method, meaning that the same sample cannot be analyzed twice or byanother method for validation Also, biological samples often need treatment,such as heavy metal staining, for improved contrast In this case, a scanningtransmission electron microscope (STEM) belonging to the group of TEMscan be of use

prepa-FIGURE 13 Differentiation between characterization and determination in the chemical context and their relationship with primary data-information-knowledge hierarchy.

analytical-Analytical Nanoscience and Nanotechnology Chapter j 1 25

In order to image NPs in more natural conditions, environmental scanningelectron microscopy [79] (ESEM) has been developed Although its samplechamber and detector cannot achieve atmospheric pressure, nanoparticles can

be visualized under almost natural conditions [80], with residual hydrationwater still on the particles This water layer also serves as a conductor on thesurface so the sample does not need to be conductively coated[81]

AFM is one of the most common nanometrology methods and hasnumerous applications [82] With this technique, it is possible to analyzesamples under moist conditions or even in liquids, which affords minimumperturbation Under liquid conditions, particles can eventually stick to thecantilever, which leads to imaging artifacts This smearing effect can beminimized by using a noncontact scanning mode where the tip is not touchingthe nanoparticles but only feels its forces

When combined with analytical methods, additional information can begained about the elemental composition of the sample Electron microscopymethods can be combined to perform energy dispersive X-ray spectroscopy(EDX or EDS) and electron energy loss spectroscopy (EELS) for the elementalanalysis of the nanoparticles, whereas selected area electron diffraction(SAED) gives information about the crystalline properties of NPs[83].X-ray microscopy (XRM) can provide spatial resolution imaging of aspecimen in the aqueous state without the need for sample preparation[84] Avariation of the XRM is the scanning transmission X-ray microscopy (STXM),which has been used, for example, to characterize metallic Fe particles forremediation purposes[85]

Moreover, near-field scanning optical microscopy (NSOM), with a lution between 50 and 100 nm, can be used to image aggregates of nano-particles [86] A modification of confocal microscopydconfocal laserscanning microscopy (CLSM)dis able to detect fluorescent samples and can

reso-be used to characterize colloids since it is able to image thick samples[80]

4.4 Separation Techniques

Separation techniques such as size-exclusion chromatography (SEC), dynamic chromatography (HDC), capillary electrophoresis (CE), dielec-trophoresis (DEP), and field-flow fractionation (FFF) can also be used tocharacterize NPs These methods are sensitive and enable further analysis ofthe sample

hydro-The most employed method is SEC[87], which, combined with detectiontechniques such as voltammetric, ICP-MS, DLS, and multiangle laser lightscattering (MALLS), also called static light scattering (SLS), can be applied tothe characterization of AuNPs, QDs, and SWNTs, for example A drawback ofSEC is that irreversible adsorption onto the stationary phase may occur.Separation and characterization of NPs with hydrodynamic chromatog-raphy is based on their hydrodynamic radius Liquid chromatography coupled

with voltammetric detection has been also employed for the separation ofmetal nanoparticles[88].

Capillary electrophoresis (CE) has emerged as a useful technique, withfewer surface effects than in other techniques (e.g., SEC) and with low con-sumption of sample and reagents, for the characterization of nanoparticles[89] CE has been used to separate a variety of differently sized materials,including gold and silver nanoparticles, CNTs, and QDs, among others[90].Different modalities have been employed such as isoelectric focusing (IEF) orgel electrophoresis

Dielectrophoresis (DEP) has also emerged as a powerful technique in order

to manipulate nanoparticles [90] Special emphasis is given to the trophoretic separation of CNTs, due to its usefulness in discriminating metallicones from semiconductor[91], since it is capable of separate species not only

dielec-by size but also dielec-by dielectric constant However, DEP is further from routineanalytical application than gel or capillary electrophoresis

Field-flow fractionation separates nanoparticles according to particle size

in terms of their diffusion coefficients in a very thin, open channel Separationrelies on the combination of an applied field and a longitudinal carrier flow Ithas been successfully used to analyze a wide range of NPs[92,93]includingSiO2, CNTs, QDs, gold, and silver

Other separation techniques used in this field are centrifugation [94],ligand-assisted extraction [95], solid-phase extraction [96], microextractionwith ionic liquids [5,8], microfiltration [97], dialysis [98], and cloud-pointextraction[99]of different nanomaterials (Table 1)

4.5 Spectroscopic Techniques

Spectroscopic methods also have a wide range of applications for the analysis ofnanomaterials Fluorescence-based techniques are used for the analysis

of nanoparticles with a strong native fluorescence, such as quantum dots (QDs),

or more recently, carbon dots (CDs) or graphene quantum dots, which can becharacterized by their absorption or fluorescence spectra The position ofemission bands can be correlated with particle size and is commonly used

to monitor size in the synthesis of QDs[100], for example In describing theUVeVis absorption spectra of metal NPs, the term surface plasmon is used,which describes the oscillating electron clouds present at the metal-solutioninterface Particle concentrations can be quantified from absorption measure-ments provided the optical constants for the nanoparticles are known[100,101].Vibrational spectroscopies, both Raman and infrared, are useful forthe analysis of nanoparticles, especially carbon nanoparticles The two tech-niques differ in the excitation source used, the way signals are monitored, andthe selection rules employed on vibrational modes At present, Raman spec-troscopy is the most important technique for the analysis of carbon nanotubes[102] because Raman measurements are simple, can be done at room

Analytical Nanoscience and Nanotechnology Chapter j 1 27

temperature and under ambient pressure, are quick, nondestructive, andnoninvasive[103], and it is possible to find sophisticated laboratory equipment

as well as less expensive, portable equipment The widespread acceptance ofRaman spectroscopy relies on its usefulness to provide information aboutvibrational properties that can be correlated with the structure and electronicproperties of the nanotubes Tip-enhanced Raman scattering (TERS) has beenalso used for carbon nanotubes [104], for example In the case of SWNTs,

TABLE 1 Specifications of Methods for Analysis and Characterization

of Nanoparticles

Method

Approximate Size Range (nm)

Limit of Detection a

Level of Sample Perturbation

a For comparison mass concentration limit of detection for 100 nm particles are estimated.

b (d.d.)= detection dependant

c (UV: ppm, Fluo/ICP-MS: ppb)

Adapted with permission from Ref [81].

NIR-fluorescence[105] is a technique widely used for their characterizationaccording to their chirality[106] as well as their quantitative analysis in anaquatic environment[107].

The average bulk chemical composition of a sample can be determined byusing spectroscopic techniques such as atomic absorption spectroscopy,inductively coupled plasma with atomic emission spectroscopy (ICP-AES),optical emission spectroscopy (ICP-OES) [108], and mass spectrometry(ICP-MS) [109,110], especially for the analysis of metallic nanoparticles.These techniques feature good limits of detection, providing compositions re-sults which are sample averages, and they also afford multi-element analysis.However, they are destructive and subject to matrix interferences[111] Massspectrometry techniques are gaining growing importance thanks to theircompatibility with any type of sample, their extremely high sensibility, and easycoupling with separation techniques to obtain real-time information[112].Light scattering is a very commonly used method to determineparticle size Among the scattering techniques, dynamic light scattering (DLS)

is widely used for sizing of NPs and determining their aggregation

in suspensions [113] The advantages of DLS are the rapid and simpleoperation, readily available equipment, and minimum perturbation ofthe sample The limitations are the interpretation and critical review of the dataobtained, especially for polydisperse systems[81] Turbidimetry and nephe-lometry have been also employed to measure particle concentration[114].Laser-based methods worth mentioning are the small angle X-rayscattering (SAXS), which is able to characterize mono- and poly-dispersesystems, and laser-induced breakdown spectroscopy (LIBS), which has avery low detection limit and is suitable for the size and concentration analysis

of colloids Nuclear magnetic resonance (NMR) spectroscopy [115] isused to determine the dynamics and three-dimensional structure of thesamples, whereas X-ray spectroscopy provides crystallographic informationwhich can be used for the characterization of NP surfaces and coatings[116]

4.6 Other Techniques

In addition to those techniques, electrochemical methods[117]of analysis ofnanoparticles have been also employed for their analysis such as voltammetry[118,119] Thermogravimetric analysis (TGA) has been also utilized, forexample, as a bulk characterization method for determining carbon nanotubequality after manufacturing[120]and/or after oxidation processes[121].Nowadays, there is a tendency to combine microscopic and spectroscopictechniques in order to obtain information about both the size and chemicalcomposition of materials Examples are the combination of AFM-ATR-IR[122], AFM-Raman[123], or AFM-SECM (scanning electrochemical micro-scopy)[124] These techniques may also be useful in the case of the analysis

of nanomaterials

Analytical Nanoscience and Nanotechnology Chapter j 1 29

4.7 Nanometrology

Nanotechnology constitutes a large industrial sector and is expected tocontinue to grow at a very fast rate Such a rapid development has raised theneed for accurate, precise control of the dimensions of nanomaterials Reliableand precise measurement in the nanoscale is a bottleneck for the full devel-opment of nanoscience and nanotechnology

Nanometrology can be defined as the science of measurement at thenanoscale level Nanometrological measurements include not only length orsize (shape, aspect ratio, and size distribution) but also chemical composition,nanoparticle concentrations, and optical, force, mass, electrical, and variousother types of properties[125]

The complexity of nanometric measurements arises from the smalldimensions of nano-objects, which are usually less than 100 nm in size.Measurements at this scale require a high precision (frequently about 0.1 nm)and, hence, effective methods of measurement Moreover, a feature of nano-materials is the exceptional properties they exhibit by virtue of their small size.However, efficiently exploiting such properties requires a sound knowledge oftheir nature and the ability to characterize the physics or chemistry of verysmall objects[125]

To measure is to compare, and for this purpose new standards and referencematerials are needed, both for instrumental and metrological calibration There

is a lack of reference nanomaterials, only a few size, but not mass or numberconcentration certified nanoparticle reference materials are available (e.g.,NIST reference materials 8011e8013 [126], ERM-FD100 and ERM-FD304[127], or BAM-N001 [128]) The development of chemical nanometrologyrequires extensive specific infrastructure, in which standards constitute itsAchilles heel

5 FINAL REMARKS

Analytical nanoscience and nanotechnology constitutes a subdiscipline that isnot diverted from general exponential growth of almost all branches of nano-science and nanotechnology in the second decade of the twenty-first century.Frequently, it is forgotten that the nanometric size is not the key of this growth,but the exceptional physico-chemical properties of nanostructured materialsthat have conferred them such scientific and technological importance

In fact, the rapid growth of nanotechnology may alter the natural sequence

of any scientific-technical evolution: research þ development þ innovation(Rþ D þ I) The intermediate step, which is indispensable, is being alarm-ingly reduced in nanotechnology, with the aim of rapidly introducing into themarket the nanotechnological products

Two key facets of analytical nanoscience and nanotechnology can bedefined, (1) the consideration of nanoparticles and nanostructured materials

as tools for the innovation and improvement of (bio) chemical measurementprocesses, which is the aspect most developed to date, and (2) theirconsideration as objects, with the aim of extracting physico-chemical in-formation from the nanoworld A third option exists as an interface betweenboth of them, which consists of nanoparticles being employed as tools inanalytical processes for the characterization/determination of othernanomaterials.

ACKNOWLEDGMENTS

The authors wish to thank Spain’s Ministry of Innovation and Science for funding Project CTQ2011-23790 and Junta de Andalucı´a for Project FQM4801.

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