Peer Review Article | Open Access | Published 9th January 2005
Developments in nanotechnology and nanomaterials in pharmaceutical science
David Carey, Advanced Technology Institute, University of Surrey, Guildford, UK | EJPPS | 1003 (2005) | Cite this article https://doi.org/10.37521/ejpps.10403| Click to download pdf
Summary
An understanding of the diverse nature of nanotechnology and the advantageous properties of nanomaterials· is gaining an appreciation across a whole range of disciplines. In this paper the role of nanotechnology as an enabling technology, and the characteristics of nanomaterials in general and to drug delivery systems in particular, will be presented. A brief discussion of some of the more common tools of nanotechnology leads to a description of nanopatterning using dip-pen nanolithography. Some of the important properties and applications of a range of 'bottom up' produced nanomaterials including functionalised carbon nanotubes and dendrimers are highlighted as well as the use of quantum dots which can be used as fluorescence biomedical tags.
Key words: Nanotechnology; nanomaterials; drug deli very systems
1. Introduction
What is nanotechnology?
Nanotechnology is concerned with materials science and engineering and its understanding and application at or around the nanometre level. In practice this is related to the ability to manipulate individual atoms or molecules in order to construct structures with dimensions of up to 100nm. Other definitions extend this upper size limit to 300nm, however, to date there is no single accepted definition of a nanomaterial.
Indeed, it could be argued that a single definition of the upper size limits of nanomaterials is unnecessary, provided the nanomaterial possesses superior properties. In the case of nanotechnology, the definition that has been given in a recent UK government report:
'Nanotechnology is about new ways of making things . It promises more for less: smaller, cheaper, lighter and
faster devices with greater functionality, using less raw material and consuming less energy'.
can be considered as the most practical .¹ Realising technology that functions at a nanometer scale can either be done using a ' top down' approach by building upon the expertise in microtechnology and reducing the dimensions accordingly. The alternative, ' bottom up ', approach exploits self-assembly of atoms, clusters of atoms and molecules . While the former follows naturally from established microsystems and microfabrication technology, the latter is associated with the realms of chemistry , biology and pharmaceutical chemistry as shown in Figure 1. Both approaches ultimately lead to nanoengineering, though the routes chosen will often
determine the products that are produced.
It is worth emphasising at an early stage that nanotechnology should be considered a broad term reflecting
techniques and processes that transcend the usual discipline boundaries. It is this interdisciplinarity that is one of the key aspects for the future success of nanotechnology. This interdisciplinary nature of nanotechnology is also reflected in the areas in which significant advances may emerge .
Specifically, in the UK the areas of drug delivery systems, electronics, tissue engineering, nanomaterials and sensors - especially at the functionalised bio-physics inte1face as well as instrumentation have been identified to be of particular importance ¹. These areas of research mirror the five areas of nanotechnology that have been prioritised in Japan after recommendations from the Council for Science and Technology Policy .² The five areas are:
( 1) nanodevices and materials for the next generation of communication systems,
(2) materials for environment and energy saving,
(3) nanobiology for medical-care technology and biomaterials,
(4) generic technologies such as fabrication and
(5) novel materials.
In the US the National Nanotechnology Initiative also strives to develop new products but particular emphasis is placed on fundamental research and development as well as training .³
Recent attempts to quantify the market value of nanotechnology have suggested that nanotechnology as a
whole is estimated to be $11 trillion by 2010 with the market associated with nanomaterials growing to $900 million in 2005 and $11 billion by 2010⁴. Furthermore , in a recent survey⁴ of companies producing nanomaterials it was reported that the primary market for nanomaterials in the medical/pharmaceutical sector is 30%. This was closely followed by chemical and advanced materials (29%), information and telecomunications (20%), energy ( 10%), automotive (5%) and aerospace (2%). Finally, it should be pointed out that, as noted in reference 1, nanotechnology can also be described as a disruptive technology - in which new products and ideas replace rather than increment what is currently available. The example of the compact disc as a common data storage device has made magnetic recording tape largely redundant. In this paper we wish to describe some existing and emerging aspects of nanotechnology, both in the form of instrumentation as well as materials and applications. The paper concentrates on the bottom up approach to nanomaterials with specific examples related to drug delivery.
Developments in nanotechnology instrumentation
1 Scanning probe microscopy
One of the key areas that has facilitated the development of nanotechnology has been the rapid development of instruments capable of observation and manipulation at the nanometer level. The two most common are the scanning tunnelling microscope (STM) and the atomic force microscope (AFM) as shown in Figures 2a and 2b. These are (the main) two instruments in a suite of instruments generally referred to as scanning probe microscopy (SPM). Other variations include instruments which probe electric, magnetic, electrical capacitance and electrochemical properties of various materials. The STM probes the electronic properties of the sample by applying a voltage difference between the sample and a sharpened metal tip.
The tip is not in contact with the sample but a quantum mechanical tunnelling current appears which depends on the electronic properties of the sample (Figure 2a). The AFM comes in two main varieties; one in which a tip is in direct contact with the sample - known as contact mode and the other in which the amplitude of the vibrating cantilever is monitored as the sample is scanned in the xy plane - noncontact or tapping mode Figure 2b. Force-distant curve can also provide information on the mechanical and elastic properties of the sample under investigation.
In both of the two methods described, the surface topography of the sample can be probed; however, for
biological or soft samples the non-contact mode is usually preferred . In addition to surface topography, the variation of the cantilever's phase is able to prove additional information in terms of hydrophobic or elastic nature of the surface. However, quantitative information from phase imaging is difficult due to the relative large
variation of the cantilever (tens of nm) and the fact that in this range the cantilever experiences both attractive and repulsive interactions. One way around this is to use torsional AFM cantilevers rather than a vibrational cantilever. It is also possible to coat an AFM cantilever with a protein which can probe specific protein absorption on the substrate.
2 Dip-pen nanolithography
The AFM tip can also be employed in transporting molecules using capillary action Figure 2c and has given rise to dip-pen nanolithography (DPN). DPN is a directwrite soft serial lithography technique that can be used to create nanostructures without the need for resist or stamps. Using the AFM tip it is possible to produce high resolution features with widths of 15 nm. DPN is another way to pattern functionalised molecules at specific sites within a particular nanostructure.
(i) The original DP experiment consisted of writing l-octadecanethiol molecules with a 30 nm line width resolution on a gold thin film.⁵ The width of the line produced depends on tip scan speed , temperature and humidity.
(ii) Protein nanoarrays have also been produced using DPN and it was reported that that the nanoarrays exhibited almost no detectable non-specific binding of proteins to their passivated sections.⁶
(iii) Finally, modified oligonucleotides and modified DNA have both been patterned on silica and gold.
These two studies demonstrated the direct assembly of individual oligonucleotide-modified particles on a surface and the deposition of multiple DNA sequences in a single array.⁷
It should be noted that since DPN is a serial technique it complements parallel methods such as micro-and nanocontact imprinting. This latter technique is the embossing of a patterned mold in a heated resist. Two further areas of active research consist of the attachment of protein and carbon nanotubes to AFM tips for added selectivity.
3 Developments in biophotonics
A final area where nanotechnological tools are emerging, which is particularly important for drug deliver systems, is the use of biophotonics at the nanoscale. Conventional biophotonics uses light as a probe of the bulk properties and reactions of cells under investigation . A molecular understanding of drug-receptor interaction (e.g. drug induced receptor trafficking) and, consequent signalling sequelae, are important for understanding the molecular basis of drug action and rationale for future drug design and deli very. Conventional biophotonics microscopy of cells or tissues uses epifluorescence from biomolecular probes, such as green fluorescent protein (GFP). As such, probes are used as diagnostic devices to detect target
biomolecules (DNA, proteins , etc .), to assess binding of drugs to receptors for drug development, or to monitor in real time the activity and dynamics of biomolecules in living systems. A variant of the well known fluorescence microscopy is total internal reflection fluorescence (TIRF)microscopy, in which the excitation beam is internally reflected from a glass-liquid interface. The evanescent field decays exponentially over a few hundred nanometres from the interface, so that selective excitation of fluorophores close to the interface occurs.⁸ For a cell adhering to the glass, selective excitation of individual membrane-sited fluorophores may be undertaken. High resolution imaging of single fluorophores in or adjacent to a cell 's plasma membrane can therefore be performed , in order to examine membrane and near-membrane signalling events, for both fundamental and clinical applications.
A second aspect in modern biophotonics concerns the dynamics of single molecules which can also be probed using GFP; however, there are new classes of labels using nanometer sized semiconductor quantum dots (QDs).
Quantum dots are nanoparticles which luminesce under light, with the size of the dots controlling its colour, e.g. certain 2nm quantum dots luminesce green, while 5nm quantum dots luminesce red. In contrast to conventional fluorophores such as GFP, QDs exhibit high-intensity emission over a stable narrow spectral range and do not suffer significant photobleaching, and can be excited by a common light source. They may be smaller than GFP leading to reduced steric hindrance, a property that could be useful when used as a probe.
Synthesis of nanomaterials
Nanomaterials are materials whose properties of interest have at least one dimension that is in the range from l to 100nm . Whilst there is no hard and fast rule as to what constitutes nanomaterial, this definition is sufficient to include nanopowders and dendrimers as well as micronlong rolled sheets of carbon that have a diameter of up to 30nm called carbon nanotubes. Larger diameter structures are referred to as carbon nanofibers. We will discuss both dendrimers and carbon nanotubes, as two examples of nanomaterials, later. There are several ways to make use of produce nanomaterials that employ a bottom up approach to growth.
(i) Gas-phase synthesis, including chemical vapour deposition , involves the splitting up of source gas/es
into reactive species such as radicals and subsequent deposition on to substrates. This allows the growth of
thin films and by using ultra-high vacuum and organometallics in a technique called molecular beam
expitaxy, mono- and even sub-mono layer deposition is possible. This can be used for a range of materials
including oxides and carbon films. Several growth modes may occur which include a layer-by-layer
growth (Figure 3a) in which a complete wetting layer is formed at one extreme, to another growth mode called
the V-W mode in which no wetting layer is present and island growth immediately occurs (Figure 3b). The
third growth mode (called the S-K mode - Figure 3c) results in an initially wetting layer but the increased
strain in the layer results in the uppermost layers breaking up into islands. Control of the island diameter
and their diameter distribution are crucial factors. Furthermore it is difficult to control the location of the islands without using advanced (expensive and low throughput) techniques such as electron beam lithography. One advantage of plasma based methods is that whilst the plasma temperature can be in excess of 1000°C the temperature on the substrate can be held at or near to room temperature. This allows the use of organic or low melting point substrates and changes can be tailored to happen only at the surface.
(ii) Metal thin films can be formed via magnetron sputtering or thermal evaporation . For lower quality,
electrodeposition may also be used. In the case of sputtering or thermal evaporation, it is possible
though subsequent annealing to form metal island themselves. Figure 3d shows an AFM image of nickel islands produced by annealing a 8nm thick metal film. Very often, larger islands grow at the expense of smaller islands by a mass transport process which results in well spaced out, larger diameter islands.⁹ Smaller islands can be produced by etching, using a plasma etch such as ammonia. Again, the diameter and distribution are important factors but whilst scalability is quite good, there is little control of the location of the islands.
(iii) Wet chemical processes including sol-gel methods. A sol consists of a colloidal solution suspended in a
liquid and a suspension that keeps its shape is called a gel. Conventional colloids are microns in diameter so
nanometer based sols are an emerging area of materials science. Nanomaterials based on sol-gel technology require the formation of nanosized sols and the gelation of the sol to form a continuous network. There are four discrete aspects in the sol-gel process; hydrolysis through the addition of hydroxy groups, polymerization of the monomers to form particles, the growth of the particles and finally the agglomeration of the particles and the formation of the network . Factors such as solution pH , temperature and reagent determine size.
(iv) Finally, dendrimers are well defined, highly branched mono-disperse nanometer-sized macromolecules that
radiate from a central core. The synthesis of these macromolecules proceeds via a repetitive reaction sequence that leads to complete shells and produces a highly symmetric structure in which there is an ability to accurately control the surface terminating groups.
This ability to decorate and functionalise the dendrimer surface by adding suitable reactive groups allows for
charged, hydrophilic or hydrophobic surfaces.
For generic nanomaterial drug delivery schemes, the active material (drug) is dissolved or encapsulated or
adhered to the surface of the nanomaterial. For drug targetting the carrier should:
(i) be capable of circulation in the bloodstream,
(ii) be sufficiently small to be able to enter the targeted
cells and
(iii) be able to release the drug when required.
As nanomaterials two candidates have emerged - nanopolymers and dendrimers.
Nanopolymers have an advantage over liposome and antibody drug delivery. It is well known that liposomes can be found in organs where they are not required and the presence of common tumour-related receptors on healthy cells makes targeting of treatment difficult. Water soluble polymers, such as poly(ethylene glycol), allow the use of single molecule species which can circulate in the body for 24 hours. One disadvantage of this polymer approach is that the drug carrying fraction is commonly less than liposomes. The attraction of dendrimers in drug delivery is partly due to presence of the cavity. At the centre of the dendrimer is a nm-sized cavity which is capable of hosting a wide range of inclusions such as metal atoms, or organic molecules. In this way the structure of dendrimer molecules is reminiscent of a micelle with drugs being located at the core of the dendrimer and the genetic or protein material attached on the surface. Drugs , ligands and agents to increase solubility may also be attached to the surface and may lead to polyvalent reactions. Finally, it is worth noting that nanomaterials have also emerged as an important delivery mechanism for DNA and gene therapy.
In almost all of the above examples the interaction between the species and the substrate is important. In the
case of the example shown in Figure 3d, the controlling factors are the interaction between the nickel and the
underlying oxide substrate and how the interaction is affected by heating the sample to 500°C. In the case of
dip-pen nanolithography (Figure 2c), the interaction between the substrate, the molecule and the cantilever and
how that interaction changes with temperature and humidity are important. These types of interaction are a
common feature in a range of self-assembled 2- and 3- dimensional assemblies. One area in nanobiology where this plays an important role is in the formation of self assembled monolayers (SAMs) where the important interaction is between the substrate and a specific functionalised group associated with part of a molecule. One well known example is that of alkanethiols on gold , in which the sulphur containing group (-SH) exhibits a strong attraction to the gold. The hydrocarbon tail points away from the gold substrate and through careful chemistry it is possible to add different active groups to the other end of the molecule. Addition of a methyl group can make the surface hydrophobic, whereas addition of a phosphate group can make the surface hydrophilic.
Finally mechanical processes such as ball milling consist of producing finely nanosized powders by breaking down the structure. Methods to produce large amounts of nanomaterial are likely to be one of the driving forces in nanomaterial research in the next few years.
Carbon nanotubes as a nanomaterial
Carbon nanotubes (CNTs) are tubular structures with nanometer sized diameters but which may have lengths in excess of 10 microns (Figure 4). They can be viewed as consisting of the sheets that make up graphite - called graphene layers - which are rolled up and exhibit some remarkable properties. ¹⁰ Single wall nanotubes have been reported to have an electrical current carrying ability a thousand times better than copper and a thermal conductivity larger than diamond which is the best, if somewhat expensive, available substrate for heat
dissipation in high power devices. In addition to single wall tubes, multiwall tubes consisting of concentric
cylinders of tubes are possible. There are three key areas in the application of CNTs: the hollow core, a surface that can be functionalised and electronic and related properties of the tubes themselves representing high aspect structures. Very often the centre of the nanotube consists of an empty core and holds the possibility that CNTs could be used in hydrogen storage as well as encapsulating atoms - this is sometimes referred to as a carbon peapod . ¹¹ In terms of pharmaceutical industry it may be possible that CNTs may find application in drug delivery in which the drugs are encapsulated in nanomaterial such as dendrimers which themselves are
encapsulated in CNTs. Secondly. the wholly carbon surface naturally leads to the importance of functionalisation. In the area of nanobiology chemical functionalisation of nanotubes is emerging as an
important avenue to break-up nanotube bundles and improving solubility. One approach to improve nanotube
solubility is by side-wall covalent functionalisation, or using wrapping effects around the nanotube using
peptides, DNA and conjugated polymers ¹². Functionalisation is also important for sensors, such as
molecular or gas sensors. It has been shown that the resonant frequency of a copper resonator coated with
CNTs is adjusted in the presence of polar gases such as NH3, as well as non-polar gases such as He and Ar¹³
Larger shifts in the resonant frequency are observed for the polar molecules due to larger changes in the dielectric constant associated with the absorption of the molecules.
Another type of NH3 detector can be constructed based upon conductivity changes for multiwall CNTs exposed to ammonia. In both cases the strong affinity of NH3 to the tubes makes degassing of the detector an important issue.
The high aspect ratio of the tubes coupled with their good electrical conductivity lends them to emission of
electrons when the tubes are exposed to a high electric field . In this way a display based on field emission from CNTs can be produced. Samsung have demonstrated a 38 inch full colour display. ¹⁴ In addition since CNTs are excellent electron sources they are finding applications in lighting sources and microwave devices. In addition to their electrical properties, CNT embedded in polymers have been shown to improve the mechanical properties of polymer-nanotube composites and produce composite fibers over 100 m in length .¹⁵
Summary
In conclusion, nanotechnology represents a truly interdisciplinary area where tools and techniques acquired
in one area can be applied elsewhere. Nanotechnology as a term should not be considered the domain of one
particular discipline but an umbrella in which both existing and emerging areas can be advanced.
Nanomaterials may also present the most efficient way to utilize a whole range of properties which to date, cost and scalability have made prohibitive. In terms of pharmaceutical science and drug delivery systems,
controlled and targeted drug delivery represents one of the frontier areas of science. Most drugs perform better in nanoparticulate form and can be administered in smaller doses. They have many advantages over conventional dosage forms being more efficient and with fewer side effects. Drugs can be encapsulated in a variety of carriers and encapsulated drugs can be protected from degradation. The carrier can be a dendrimer or a carbon nanotubes. The surface of the carrier can also be functionalised for greater specificity .
Acknowledgement
The author is grateful to the EPSRC for funding under a Advanced Research Fellowship.
Table 5 Dispersion rates per minute, MCPs, when wearing undergarments and cleanroom garments
Figure 11 Dispersion rates of MCPs and particles from 3 people wearing cleanroom undergarments and cleanroom garments
8. Discussion and conclusions
Information is provided in this article about tests used to determine the contamination control properties of fabrics used to manufacture garments worn in pharmaceutical and healthcare cleanrooms. These tests can be used when garments are first selected for use in the cleanroom but in this article, they were used to investigate the deterioration of new garments. Tests were carried out on a previously unstudied fabric when new, and after 10, 20, 50, and 70 decontamination cycles. These cycles included washing, drying, sterilisation by gamma radiation. It is probable that sterilisation by autoclaving would have given different results, but this was not investigated.
The appearance of the fabric was observed as the number of decontamination cycles increased from new to 70 cycles and there was a clear loss in the fabric colour, as shown in Figure 4. In addition, the fabric was noticeably thinner after 70 cycles and it was difficult to put on garments without tearing the fabric. This change in the fabric was thought to indicate deterioration in the contamination control properties of the fabric and was investigated.
The weave of the fabric was observed by a scanning electron microscope and images are shown in Figure 8 of the new fabric and after 50 decontamination cycles. After 50 cycles, the fabric was shown to maintain a tight and consistent weave, with no indication of material breakdown, including the integral carbon encapsulated grid, and there was little or no difference from the new fabric. However, although no image is included in this article, it was found that after 70 cycles, there was a break-up of the carbon encapsulated grid.
As reported in the graph in Figure 5, the equivalent pore diameter of the new fabric was 11.3 µm and a reasonably consistent profile of pore size was maintained throughout the increasing number of decontamination cycles. After 70 cycles the pore size was the same as the new fabric. Pore size is a key parameter used to predict the ability of a woven fabric to provide effective barrier and containment control, and the smaller the pore size the more effective the control. It was expected that the particle removal efficiency would maintain a similar consistent profile through decontamination cycles. However, this was only partly confirmed by the results given in the table in Figure 6, as the overall drop in particle removal efficiency after 50 cycles was found to be 13.9%, and after 70 cycles it was 26.1%.
Tests were also carried out on the release of particles from the fabric (dry linting propensity). The results are given in the graph in Figure 7 and they showed an increase in particles after 10 contamination cycles but all subsequent results up to 70 cycles were less than those recorded for the new fabric.
When taking into account all of the above information, it is considered that a limit of 50 decontamination cycles should be placed on the use of the fabric studied before replacement. To confirm this, and to study the dispersion rates of MCPs from garments worn by personnel and made from this fabric, a dispersal chamber was used. Only one set of results was obtained from the three personnel who participated but the dispersal profiles from each individual were consistent. The control of dispersion of particles is related to the pore size of the garment’s fabric and, therefore, the larger the particle, the more effective the fabric will be. This was confirmed by the reductions of particles ≥0.5 µm, ≥5 µm, and MCPs (average size typically 12 µm ¹¹ ¹²), which gave average reductions compared to cleanroom undergarments of 35, 151 and 195-fold, respectively. It was also found that the average dispersion rate of MCPs from the three personnel when wearing garments that had gone through 50 decontamination cycles was 10/minute (0.2/s). This was close to the lower dispersion rate from personnel wearing garments made from different fabrics that had gone through 50 decontamination cycles and reported by Ljungqvist and Reinmuller ⁴ to have satisfactory emission rates of 9.8/s, 1.9/s, 0.1/s and 0.2/s.
Using the information from the contamination control tests of the fabric studied and the dispersion rate in the chamber, it appears that the control of the dispersion of MCPs by the fabric and garments was satisfactory up to 50 decontamination cycles but not 70. However, it has been shown that different fabrics will give different rates of change of their contamination control performances over time ², and it is also likely that the type of decontamination cycle will affect the rate of deterioration of fabrics. It may, therefore, be considered appropriate to investigate garments when first introduced into a cleanroom, and over time, by the use of tests described in this article to determine how many decontamination cycles can be used before garments lose their contamination control effectiveness.
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Author Information
Corresponding Author: David Carey, Lecturer and ESPRC Advanced Research Fellow
Naonelectronics Centre, Advanced Technology Institute,
School of Electronics and Physical Sciences,
University of Surrey,
Guildford
GU2 7XH
England
Email: david.carey@surrey.ac.uk