WO2005075663A2

WO2005075663A2 - Nanosized composite material containing a biological compound and process for the preparation

Info

Publication number: WO2005075663A2
Application number: PCT/IE2005/000009
Authority: WO
Inventors: Andrew Ian Minett; Simon Edward Moulton; Werner J. Blau; Gordon George Wallace
Original assignee: The Provost, Fellows And Scholars Of The College Of The Holy And Undivided Trinity Of Queen Elizabeth, Near Dublin; University Of Wollongong
Priority date: 2004-02-05
Filing date: 2005-02-07
Publication date: 2005-08-18
Also published as: IE20050054A1; WO2005075663A3

Abstract

A process for the preparation of a composite material comprises the steps of adding a biological compound to an as -produced material preparation to form a solution; mixing the solution to form a dispersion; allowing unwanted material to settle out of the dispersion and removing a composite material. The material may be isolated from the composite material dispersion by heating the dispersion to a temperature less than the combustion/decomposition temperature of the nanosized material.

Description

"Nanostructure material and process for the preparation thereof"

Introduction

Carbon nanotubes (CNT) have attracted a large amount of interest since their discovery in 1991 by Iijima [1J due to their exceptional electronic and mechanical properties [2]. Since their discovery, CNT have been studied for their application in a wide range of areas. Their exceptional electronic properties suggest that CNT will have the ability to mediate electron transfer reactions with electroactive species in solution when used as the electrode material 13]. To this end CNT have been applied to areas of research such as; hydrogen storage [4-6], photovoltaic devices [7, 8], actuators [9-11], energy storage [12-14] and field -emitting flat panel displays [15]. Interest in studying the biological application of solid-state devices based on nano- materials has increased significantly in recent years [16-19]. The potential use of CNT in biorelated areas has prompted many researchers to investigate the functionalisation of CNT with biological macromolecules such as proteins and oligosaccharides [20-25]. Combining the unique electronic properties of carbon nanotubes with remarkable biomolecular recognition capabilities could lead to miniature biological electronics and optical devices including probes and sensors [26].

Methods have been previously disclosed for purifying carbon nanotubes including purification by treatment with strong chemical oxidants, by burning of unpurified samples and using surfactants or polymers. One such method is described in US5,560,898. Chemical oxidants do separate the nanotubes from the impure soot but tend to break chemical bonds in the nanotubes, especially at the tips. Methods involving burning tend to produce higher purity samples but the yields are very poor (typically 1 to 2% yield of carbon nanotubes). Purification using surfactants is more efficient but involves high power ultrasonic bath treatment which is known to remove the nanotube end caps and break the nanotubes into shorter lengths. Various methods of producing CNT have been refined including arc discharging, laser ablation and chemical vapour deposition (CVD). The arc discharge technique remains the most popular for the fabrication of well-graphitised, high quality nanotube material [27]. While these methods produce carbon nanotubes, they also produce other non-nanotube material in the form of graphitic particles (GPs) such as carbon onions and turbostratic graphite which can be generally described as graphitic soot or carbon soot. A means of purifying this graphitic soot was first published by Coleman et al [28, 29]. Using conjugated polymer poly(m-phenylene-co-2,5- dioctyloxy-/?-phenylenevinylee) (P PV) the non-nanotube carbon particles were filtered out leaving the CNT suspended in the PmPV polymer solution. The high wettability between the polymer and the CNT, due to the coiling nature of the polymer, was attributed to this 'filtering' effect [30]. This purification method is effective but the process of making PmPV is time-consuming and expensive.

It has been demonstrated for multi-walled nanotubes (MWNTs) that the wrapping of polymer ropes around the nanotube lattice occurs in a well-ordered periodic fashion [30, 31]. This wrapping has been attributed to the van der Waals attraction between the 6-carbon phenyl ring of the polymer and the hexagonal lattice structure of the nanotube [32]. In this way, nanotubes are incorporated into the polymer matrix and the solution can be used to fabricate thin films for electronic or mechanical application [33].

Dieckmann et al [34] used non-specific binding of a -helix amphiphilic peptide to assist in dispersing CNT in aqueous media. The hydrophobic region of the peptide interacted with the aromatic surface of the CNT, while the hydrophilic face promoted self-assembly through peptide-peptide interactions. The problems involved with this method include limitations on the weight percent (% w/w) of carbon nanotubes that the method disperses in solution. Secondly, the method does not differentiate between nanotubes, or the contaminants in the nanotube soot. Amorphous carbon and metal particles remain in the dispersion. The invention is directed towards providing a nanostructure material and a process for the preparation thereof which will at least assist in overcoming some of the problems with known preparation and purification techniques. The composite dispersions and purified nanotubes of the invention have potential application in a variety of fields.

Statements of Invention

According to the invention there is provided a process for the preparation of a nanostructure composite material comprising the steps of; adding a biological compound to an as-produced nanostructure material preparation to form a solution; mixing the solution to form a dispersion; allowing unwanted material to settle out of the dispersion; and removing a nanostructure composite material.

In one embodiment of the invention the unwanted material is allowed to settle for at least 24 hours.

In another embodiment of the invention the unwanted material is allowed to settle for up to 5 days.

In one embodiment of the invention the nanostructure material is selected from any one or more of carbon nanotubes, non-carbon nanotubes, nanowires, nanorods and nanoparticles. Preferably the carbon nanotubes are selected from any one or more of single walled nanotubes (SWNT), double walled nanotubes (DWNT) and multi walled nanotubes (MWNT). In one embodiment of the invention the biological compound is a biopolymer. Preferably the biopolymer is any macromolecule comprising a protein, nucleic acid, polysaccharide or polyelectrolyte thereof.

In one embodiment of the invention the biopolymer may be selected from any one or more of glycosaminoglycan, enzyme, DNA, oligonucleotide, amino acids and chitin.

Preferably the glycosaminoglycan is selected from any one or more of chondroitin sulphate, heteropolysaccharides, polysaccharides and disaccharides.

Preferably the chitin is chitosan, preferably 1,4 alpha D-glucosamine polymer.

In one embodiment of the invention the macromolecule is of bacterial origin.

In one embodiment of the invention the biological compound is dissolved in water and the nanostructure material preparation is added to the solution.

In the case of DNA the biological compound is dissolved in water at a temperature of approximately 90°C.

In one embodiment of the invention the biological compound is present in a weight ratio of at least 1:1 with respect to the nanostructure material. The biological compound may be present in a weight ratio of at least 2:1, 3:1 or 4:1 with respect to the nanostructure material. In one embodiment of the invention the biological compound is present in a weight ratio of up to 10:1 with respect to the nanostructure material.

The invention also provides a process comprising the step of obtaining the nanostructure material from the nanostructure composite material dispersion. In one embodiment of the invention the nanostructure material is obtained from the composite material dispersion by heating the dispersion to a temperature less than the combustion/decomposition temperature of the nanotube content of the nanostructure material.

In one embodiment of the invention the purity of the isolated nanostructure material is greater than 20%, preferably greater than 30%, preferably greater than 40%, most preferably greater than 50%.

In one embodiment of the invention the isolated nanostructure material has an impurity level of less than 4%.

The invention provides a process for the preparation of a nanostructure material comprising the steps of; adding a biological compound to an as-produced nanostructure material preparation to form a solution; mixing the solution to form a dispersion; allowing unwanted material to settle out of the dispersion; removing a nanostructure composite material; and obtaining a nanostructure material from the nanostructure composite material dispersion.

The invention provides a process for the purification of a nanostructure material comprising the steps of adding a biological compound to an as-produced nanostructure material preparation to form a solution; mixing the solution to form a dispersion; allowing unwanted material to settle out of the dispersion; removing a nanostructure composite material; and obtaining a purified nanostructure material from the nanostructure composite material dispersion.

The invention also provides a nanostructure composite material whenever prepared by a process of the invention.

The invention also further provides a nanostructure material whenever prepared by a process of the invention.

The invention further provides carbon nanotubes whenever prepared by a process of the invention.

The invention also provides a composite comprising a nanostructure material; and a biological compound,

wherein the biological compound interacts with the nanostructure material.

The invention further provides a composite comprising a nanostructure material; and a biological compound,

wherein the biological compound is separable from the nanostructure material to provide a purified nanostructure material.

The invention also provides a composition comprising a biological compound and a nanostructure material. The invention further provides a nanostructure material substantially as hereinbefore described with reference to examples and accompanying drawings.

The invention also provides use of a composite as hereinbefore described in the manufacture of conductive and high strength composites, energy storage and energy conversion devices, sensors, field emission displays and radiation sources, hydrogen storage media, photovoltaic devices, actuators and devices containing nanometer- sized entities/components/structures, probes and interconnects, fuel cells, dry delivery, biomedical applications including tissue engineering, optics including nonlinear optics, biocompatible composites, fibres, mats or papers, biocompatible coatings, devices or sensors.

The invention further provides use of a purified nanostructure material as hereinbefore described in the manufacture of conductive and high strength composites, energy storage and energy conversion devices, sensors, field emission displays and radiation sources, hydrogen storage media, photovoltaic devices, actuators and devices containing nanometer-sized entities/components/structures, probes and interconnects, fuel cells, dry delivery, biomedical applications including tissue engineering, optics including non-linear optics, biocompatible composites, fibres, mats or papers, biocompatible coatings, devices or sensors.

Throughout the description the term dispersion and suspension are interchangeable. A dispersion is defined as the spatial property of being scattered about over an area or volume. A suspension is defined as a mixture in which fine particles are suspended in a fluid where they are supported by buoyancy. The mixing of carbon nanotubes and a biomolecule and a liquid is used to form an inhomogeneous 'mixture\ Sonication of this mixture leads to the formation of a stable dispersion or suspension, the term 'dispersion' or 'suspension' relating to the mixing of the entities. The term solution may also be used interchangeably with dispersion and suspension. A solution is defined as a homogeneous mixture of two or more substances, frequently but not necessarily in a liquid form.

The term 'unwanted' material includes any material such as non-suspended material which may contaminate the desired nanostructure composite material. Such undesired material includes graphitic particles, amorphous material, metal particles and other contaminating material.

Brief description of the drawings

The invention will be more clearly understood from the following description thereof given by way of example only with reference to the accompanying drawings, in which: -

Fig. 1 is a schematic representation of the percentage of nanotube content (CNT) and graphitic particle (GP) content in the resultant suspension content over increasing time. Fig. 2 shows the structures of chitosan (a), chondroitin sulfate sodium salt (b), and D-glucose (c);

Fig. 3 is a graph showing a typical EPR derivative spectra taken for the DNA-MWNT soot system. (A) DNA, (B) solute from the DNA-MWNT soot sample, (C) sediment from the DNA-MWNT soot sample, (D) MWNT soot. Note that the DNA has no EPR signal and that the MWNT soot is comprised of components from (B) and (C), showing phase separation of the MWNT and graphitic particles by the DNA. B(G) on the X-axis indicates the magnetic field strength; Fig. 4 shows the Raman spectra of multi-walled nanotubes, sediment and dispersion from a DNA-MWNT sample of Fig. 3 using a 632.8 nm laser and 300-line grating;

Fig. 5 is a graph showing the percentage of multi-wall carbon nanotubes retained in the solute dispersion, as a function of the DNA:NT ratio, in the range of 2:1 up to 10:1 as measured from EPR. The value of NT soot purity is also given as reference;

Fig. 6 is a graph showing the percentage of multi-wall carbon nanotubes versus nanotube diameter for the soot, solute and sediment before and after dispersion with DNA. The nanotube diameters were observed from TEM images. It can be observed that there are preferentially retained nanotube diameters. Those nanotube diameters larger than 30nm can be observed in the nanotube soot and in the sediment, whereas, there is only one nanotube diameters (37-39nm range) observed in the dispersion solute;

Fig. 7 is a graph showing the results of Thermogravimetric analysis (TGA) of DNA, multi-wall nanotube soot and the DNA:NT composite film, prepared from the DNA, multi-wall nanotube dispersion;

Fig. 8 is a graph showing the results of TGA analysis of chondroitin sulfate, multi-wall nanotube soot and the chondroitin sulfate:NT composite film; Fig. 9 is a graph showing the results of TGA analysis of chitosan, multi-wall nanotube soot and the chitosan:NT composite film;

Fig. 10 is a TEM image of (a) untreated MWNT soot (b) purified MWNTs using chitosan dispersant. Inset in (b) highlights (dashed lines) the thin biomolecule coating; and Fig. 11 is a TEM image of (a) purified MWNTs using DNA and; (b) purified MWNTs using chondroitin sulfate.

Detailed description

We have found an improved process for the purification and dispersion of carbon soot using a biological compound or biomolecule (naturally occurring, non man- made or non-synthetic substance). We have surprisingly found that long chain biomolecules may be used to 'filter' the carbon soot, resulting in the non-nanotube graphitic particles falling out of dispersion to form sediment while leaving the nanotubes well dispersed. The nanotubes may then be easily separated out of the dispersion.

The filtering process may be monitored by electron paramagnetic resonance (EPR) and Raman spectroscopy, allowing the determination of nanotube content in the dispersion. Differential scanning calorimetry (DSC) can also be used to investigate the effect of nanotube interaction due to the crystalline nature of the biomolecule. Transmission electron microscopy (TEM) was also used to investigate the wettability of the nanotubes by the biological solution to ascertain whether debundling of the nanotubes occurs in the dispersion.

While the effectiveness of dispersing and purifying carbon soot using a synthetic organic polymer has been studied extensively, the use of biomolecules or biopolymers such as proteins and DNA has not.

We mixed carbon soot with long chain biomolecules. Upon sonication of the CNT- biomolecule solution, dark coloured dispersions formed. After three days the dispersions appeared homogenous and black in colour. The dispersion was easily separated from a fine layer of sediment at the bottom of the bottle by decantation. Typically the non-nanotube graphitic particles (GP) sediment out of the dispersion within 24hrs, however leaving the dispersion for up to 5 days has been shown to result in a higher ratio of CNT:GP in the dispersion. Fig. 1 is a schematic representation of the percentage dispersion content over increasing time.

The sediment comprises the non-nanotube graphitic particles while the dispersion comprises pristine / purified CNT. The dispersions are stable over time (>12 months), with no further precipitation observed.

The binding of the biomolecules to the nanotubes appears to displace unwanted material from the nanotube surfaces, keeping the majority of the nanotubes in the biomolecule solution whilst removing the unwanted materials. This is similar to that described for PmPV [29]. The increased amount of MWNT in the preparations of the present invention may be attributed to the larger size of the biomolecules used compared to the size of PmPV.

The purification process is efficient, non-destructive and superior to other known purification techniques. The process of the invention does not cause the nanotubes to break down. Other purification steps used such as chemical oxidation, extended ultrasound, burning and using surfactants are multi-step processes involving acids and other hazardous chemicals which lead to nanotube breakdown.

In the present invention the biomolecules interact with the carbon nanotubes. The term 'Mapping' may be used to describe the interaction which is taken to cover all means of interaction between the nanotubes and the biomolecules.

The preferred properties of the biomolecules of the invention include molecules or compounds or polymers including those that have flexible, long chain structures. These would include ring structures that could map onto the nanotube lattice. They are amphiphilic in that they comprise a polar, water-soluble group attached to a non- polar, water-insoluble hydrocarbon chain. Organisms containing DNA, chondroitin sulphate or similar would be bacteria, enzymes, microorganisms. This would include proteins and the amino acids making up the proteins. The biomolecules of the invention may include biopolymers.

Throughout the specification a biopolymer is taken to comprise any macromolecule which may be found within a living thing. Macromolecules include for example proteins (essentially long chains of amino acids), nucleic acids (such as DNA or

RNA), polysaccharides (long chains of simple sugars) or polyelectrolytes thereof. A macromolecule is a living organism that is formed by linking together several smaller molecules, such as a protein from amino acids or DNA from nucleotides. Subsets of biopolymers include glycosaminoglycans, enzymes, DNA, oligonucleotides, amino acids and chitin.

The macromolecule may be of bacterial origin.

Chitin is taken to include a tough, protective, semitransparent substance, primarily a nitrogen-containing polysaccharide, forming the principal component of arthropod exoskeletons and the cell walls of certain fungi. Chitin is the second most abundant natural polymer in the world after cellulose. Upon deacylation, it yields the novel biomaterial chitosan. Chitosan is biocompatible, antibacterial, environmentally friendly. The chitosan may be a polymer of 1-4 alpha D glucosamine.

A polymer is taken to comprise a naturally occurring or synthetic compound consisting of large molecules made up of a linked series of repeated simple monomers.

Glycosaminoglycan is taken to comprise a macromolecule found on the surface of eukaryotic cells which is thought to play a role in the cells recognition of other cells or of a substrate. It consists of a network of long, branched chains made up of repeating units of disaccharides which contain amino groups sugars, at least one of which has a negatively charged side group (carboxylate or sulphate). Common glycosaminoglycans include hyaluronate (D glucuronic acid N acetyl D glucosamine: MWt up to 10 million) for example hyaluronic acid, chondroitin sulphate (D glucuronic acid N acetyl D galactosamine 4 or 6 sulphate), dermatan sulphate (D glucuronic acid or L iduronic acid N acetyl D galactosamine), keratan sulphate (D galactose N acetyl D glucosamine sulphate) and heparan sulphate (D glucuronic acid or L iduronic acid N acetyl D glucosamine).

Chondroitin sulfate is taken to comprise one of several classes of sulfated glycosaminoglycans that is a major constituent in various connective tissues, especially in the ground substance of blood vessels, bone, and cartilage.

Glycosaminoglycan side chains (with the exception of hyaluronate) are covalently attached to a core protein at about every 12 amino acid residues to produce a proteoglycan, these proteoglycans are then noncovalently attached by link proteins to hyaluronate, forming an enormous hydrated space filling polymer found in extracellular matrix. The extent of sulphation is variable and the structure allows tremendous diversity.

The larger the biomolecule the better the dispersion and the stability of the dispersion. There seems to be a correlation between the molecular weight and the dispersive ability and stability of the biomolecule dispersion.

For example the structure of the sugar/saccharide allows glucose (Fig. 2(c)) to interact with carbon nanotubes. Glucose contains a six-membered ring which π-π stack with the available electron cloud on the nanotube lattice. However it appears that glucose as a molecule is physically too small to disperse the much larger carbon nanotubes. The glucose appears to 'coat' the nanotubes, however, it is not capable of 'holding' the nanotubes in a stable dispersion. The invention will be more clearly understood from the following examples.

The multi-walled nanotube (MWNT) soot used was prepared at Trinity College, Dublin, Ireland, using the arc discharge method [35]. The biomolecules used were Salmon Sperm DNA (Mol wt 2.0 x 10⁶ (which equates to about 3077 base pairs / from Sigma), Chondroitin Sulfate Sodium Salt (Sigma - Mol wt in 300,000), Chitosan (Korea - Mol wt 100k to 300k) and Glucose. The structures of chondroitin sulfate sodium salt (a), chitosan (b) and glucose (c) are shown in Fig. 2. All solutions were prepared in Milli-Q water.

Example 1

20 mg of the biomolecule was added to 5 ml water (0.4% w/v) and heated to 90°C to assist in dissolving. The solution was then added to 20 mg of the MWNT (0.4% w/v). The solutions were then sonicated using 0.5 sec pulses for 5min using a high power sonic tip (120W) at room temperature. The MWNT-biomolecule dispersions were left for 3 days to allow any impurities to sediment out. For each sample the resulting dispersion was separated from the sediment by decanting. The remaining sediment was placed in an oven at 60°C until completely dry.

Using a sedimentation process, quantitative measurements of the percentage of nanotubes and graphitic particles in the given biomolecule-nanotube suspensions may be determined. The transmission of laser pulses through the centre of the sample over time are monitored. Briefly, after sonicating using 0.5sec pulses for 5mins as described above the dispersion is put into a glass cell. Monitoring the laser transmission through this mixture over time results in a turbidity versus sedimentation time graph that would allow calculation of the solute and sediment content to be quantitatively obtained. The transmission may be transformed into turbidity using the Beer-Lambert Law

-17 Mo Where I/I₀ is the transmittance, T is the turbidity and / is the sample length. The turbidity is the sample concentration multiplied by an extinction co-efficient, where this co-efficient represents all absorption and scattering processes. [39]

The method for single wall nanotubes is the same as for multi-wall nanotubes.

Single walled nanotubes used were produced by CNI (Carbon Nanotechnologies Inc - Smalley group and company, Rice, Texas) and by Nanocyl (Namur, Belgium).

Electron and paramagnetic resonance (EPR) and Raman spectroscopy were used to measure the MWNT and graphitic impurities (GP) in the dispersion and sediment respectively. EPR measurements were made at room temperature using the procedure outlined by Murphy et al [27]. By correct normalisation [36], the EPR signals from the dispersion and sediment could be directly compared. Raman spectroscopy was performed on a Jobin Yvon Horiba HR800 using a 632.8 nm laser utilising a 300-line grating. EPR and Raman were performed on the raw MWNT soot, the decanted MWNT dispersion and the sediment. Depending on the sample, between 3.7 mg and 10.3 mg of the dispersion was drop cast on spin-free quartz plates (3 mm x 10 mm) to ensure that an appreciable EPR signal was obtained. Two samples of the sediment (approximately 5 mg each) were also placed into spin-free quartz tubes. Raman spectra were recorded by placing a small quantity (2 mg) of the raw MWNT soot and sediment onto a glass slide. Approximately 0.2 ml of the MWNT dispersion was also placed onto the glass slide.

Transmission electron microscopy (TEM) images were obtained on a Hitachi H7000

TEM at lOOkeV. Samples for TEM were cast on carbon coated holey copper grids (300 mesh) from the dispersion and allowed to dry in air over night. Differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA) was performed on a freestanding film made by drop casting approximately 0.5 ml of sample onto a Teflon disc (dia = 1.5 cm). Electron Paramagnetic Resonance (EPR) The EPR spectra obtained (Fig. 3) were intergraded and in all cases could be deconvoluted into two Lorentzian fits. From these fits the g value (representing position) associated with MWNTs and GP could be calculated. The g values in the samples are quoted in column 4 of Table 1. For each sample, components with g values very close to 2.021 and 2.011 were observed, which are attributed GP and MWNTs respectively [28, 29, 33, 37] .

Table 1

NOTE: (1) NSI = (Area) x (Total Mass of sample), since Area is a normalized value for lmg of sample. (2) Sed = Sediment; Sol = solution/dispersion All the relevant numerical values used or calculated in this work are given in Table 1. The treatment of the EPR data was the same as that outlined by Murphy et al [37], namely, for each of the solutes, the EPR signal obtained was initially normalized to represent 1 mg of that solute. This signal was then multiplied by the total mass of the solute to give the normalized signal intensity (NSI) that one would obtain from a measurement of the whole solute. Because the solute and sediment signals were normalized, the sum of their MWNT and GP components must represent 100% of the MWNT and GP available. Therefore, by direct comparison of the solute and sediment NSIs for MWNT and GP, the percentage of MWNT and GP held in dispersion (and in the sediment) were calculated using Equation 1 (or similar for GP).

NSI NT I SOL

PNT = 100 (1) NSI NT I SOL + NSI NT I SED

Using the NSI values listed in Table 1 and equation 1 it was possible to calculate the amount of MWNTs (NT_S0|) and GP (GP_soι) in the solution of each dispersion. The results are presented in Table 2 as the percent of MWNT and GP in solution/dispersion determined by EPR at room temperature. The results were obtained using equation 1 and the NSI data in Table 1. It appears that similar to PmPV [29], the binding of the biomolecules to the nanotubes on a 1:1 mass ratio displaces the unwanted material from the nanotube surfaces, keeping the majority of the nanotubes in the biomolecule solution/dispersion whilst removing the unwanted materials. The increased amount of MWNT may be attributed to the larger size of the biomolecules used.

Table 2.

Biomolecule NT_sol (%) GP_SOι (%) DNA 25 373 Chondroitin 26 3.5 Chitosan 56 3.4 Others [29, 38] ascribe MWNT-PmPV interactions to be a result of the polymer wrapping itself around the MWNTs, therefore dispersing the tubes. This wrapping is credited to the helical nature of the PmPV polymer. Wrapping of the tubes also allows the polymer to surround layers of nanotubes, permitting sufficiently close intermolecular proximity for π-π stacking of the 6-member ring of the polymer and the hexagonal features of the tubes to occur. This interaction also aids in the dispersion process.

The same mechanisms appear to be occurring with the biomolecules, however since the biomolecules used have a greater molecular mass they are able to hold more nanotubes and graphitic particles. As the molecular weight of the biomolecule increases so too does the %NT_so|, further indication that the size of the dispersing molecules plays an important role.

In the case of the MWNT-glucose solution it was immediately clear that both the nanotubes and graphitic particles precipitated completely. Due to complete precipitation it was not possible to decant any dispersion and therefore no EPR and Raman spectra were obtained of the MWNT-glucose hybrid. The precipitation of both MWNTs and GP from the glucose solution indicates that they are not dispersible in this biomolecule solution.

The fact that the low molecular weight glucose was not able to disperse any of the MWNT soot, even though it contains 6-member rings, suggests that polymer mapping is the dominant interaction involved in MWNT dispersing process. This may also indicate that biomolecules comprising a 6-membered ring need to have a certain molecular weight in order to form a suspension of CNT.

Raman Spectroscopy

Raman spectroscopy was used to confirm the results obtained by EPR. Raman spectroscopy was obtained using a 632.8 nm laser and a 300-line grating. Coleman et al [29] used Raman spectroscopy to investigate the purity of extracted graphitic material. A typical Raman spectrum of MWNT soot is shown in Fig. 4 where DNA is the biomolecule used. The excitation wavelength was 632.8 nm. The broad band at 1332 cm^"1 (D line) is typical of unpurified nanotube samples and arises from amorphous graphite carbon particles and defect sites on nanotubes. The band at 1582 cm^"1 (G line) is assigned to the E_2g mode of the Multi-wall nanotubes, similar to that found for highly organized pyrolitic graphite. The proportion of MWNT to amorphous material is given by the ratio of the intensities of the 1582 cm^"1 relative to the 1332 cm^"1. Fig. 4 shows a greater proportion of MWNT to GP in the solution/dispersion spectra than for the sediment and soot spectra. This indicates that most of the GP precipitated out of the dispersion to form the sediment resulting in a solution rich in MWNTs. These results were also observed for the chondroitin sulfate and chitosan samples.

It must be noted that while the relative intensities can be compared, Raman spectroscopy, in this instance, is not quantitative. However, it is possible to compare the efficiency of each biomolecule dispersion to retain nanotubes by first determining the ratio of MWNT to GP in both the sediment and solution/dispersion of each sample (Equation 2 - a is either Sed or Sol).

NT a Ratio_α = (2) ^l GP _a

Assuming that the sum of the NTRatiθs₀ι and NTRatio_Sed accounts for all of the MWNTs present in the sample it is possible to determine what percentage of the MWNTs are present in the solution (Equation 3).

The percentage of MWNT present in the biomolecule solutions is shown in Table 3. Raman spectroscopy has indicated that the DNA and Chondroitin biomolecules are equally efficient in retaining MWNT while the Chitosan is slightly less efficient. Multiple Raman spectra were recorded for each sediment and solution/dispersion with their respective 1332 nm and 1582 nm band intensities recorded. The NT_so| (%) value shown in Table 3 is the average of these spectra and associated calculations. The error provides an indication of the reproducibility of the spectra intensities.

Table 3.

# The values shown in these columns are from one spectrum only. They have been included to assist in the interpretation of the calculations.

Example 2 - determining biomolecule to nanotube ratio Dispersion of biomolecule and MWNT were prepared as described in Example 1 increasing the ratio of biomolecule to MWNT. The ratio of DNA to nanotube content was increased to 2:1, 3:1, 4:1 ,5:1 and 10:1. As shown in Fig. 5 up to 93% of available nanotubes could be retained in the solute. However, it was found that at ratios of 10:1 a decrease in suspended nanotubes was found.

For chitosan it was found that the viscosity of the chitosan dispersion became too high above a ratio of 2:1 (chitosan:NT). Increased amounts of nanotubes were found in the 2:1 ratio sample, but the viscosity of the 3:1 sample was so high that everything was kept in 'dispersion '. That is, no sediment was obtained even after 3 days of settling. The results were visualised using TEM microscopy (Figs 10 and 11), where the purification effect could easily be seen by the reduced amount of graphitic particles in the samples.

Statistics compiled from the TEM images of DNA based dispersions show that there are some nanotube diameters that are preferentially retained in the solute as shown in Fig. 6. The graph shows that for nanotube diameters above the 28-30nm range, only one nanotube diameter (37-39nm range) was found in the solute, whereas larger diameter nanotubes were found in the soot and corresponding sediment samples. The diameter may be a factor in the optimisation of the process.

Thermogravimetric analysis (TGA) was used to study the decomposition characteristics of the MWNT-biomolecule dispersion with respect to the biomolecule and original MWNT soot. The TGA results on the DNA, chrondroitin and chitosan biomolecules are shown in Figs 7 to 9.

In the case of a chitosan biomolecule composition we have found that the biomolecule suddenly completely burns off by 600°C (Fig. 9). Surprisingly however it was found in the traces for DNA and chrondroitin (Fig. 7 and 8 respectively) that even at 1000°C, up to 20% of the biomolecule and the biomolecule:NT composite still remain. This indicates a very strong interaction between the biomolecule and the nanostructured material. Other separation methods such as changing the pH to improve combustibility or sonication may be used to isolate the nanostructure material from the biomolecule.

The strength of the bond may correlate to the amount of salt associated with DNA and chrondroitin. Removal of the salt by dialysis with pure water may aid separation.

The solutions have also been stable for up to 12months without visible sedimentation occurring in the bottom of the stored sample flasks. Re-bundling of the nanotubes was also inhibited as observed using optical microscopy. Re-bundling of the nanotubes results in destabilisation of the dispersion/suspension resulting in the nanotubes dropping out of the dispersion and forming a sediment at the bottom of the vial.

The invention has been described with particular reference to arc discharge nanotubes. The invention however also works with nanotubes produced by other methods, including single wall nanotubes. However, because these other nanotubes are produced using metallic catalysts, EPR cannot be used to quantify the purification process as the presence of the metal swamps the entire EPR spectrum.

The properties of carbon nanotubes such as their extremely small size combined with a highly symmetrical structure results in remarkable quantum effects. For example the quantum wire feature of a single-wall nanotube. The electronic properties of nanotubes have been correlated with mechanical, chemical, thermal, biological and magnetic interactions. Due to their large surface areas and σ-π rehybridisation, nanotubes are very attractive in chemical and biological applications because of their strong sensitivity to chemical or environmental interactions. The combination of these properties and interactions may be associated with applications of carbon nanotubes in electrochemical, electromechanical, thermal electronic and electromagnetic sensors and actuators.

Combining the unique electronic properties of carbon nanotubes with remarkable biomolecular recognition capabilities has numerous applications such as miniaturised biological electronics and electrical and optical devices such as sensors. Another application for combining the properties of carbon nanotubes and biomolecules is in the biomedical and medical device fields. For example, utilising the mechanical strength of carbon nanotubes and at the same time introducing a biocompatible component (coating) to the carbon nanotubes would lead to such devices as biocompatible stents. The invention is not limited to the embodiments hereinbefore described which may be varied in detail.

References

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Claims

1. A process for the preparation of a nanostructure composite material comprising the steps of; adding a biological compound to an as-produced nanostructure material preparation to form a solution; mixing the solution to form a dispersion; allowing unwanted material to settle out of the dispersion; and removing a nanostructure composite material.

2. A process as claimed in claim 1 wherein the unwanted material is allowed to settle for at least 24 hours.

3. A process as claimed in claim 1 or 2 wherein the unwanted material is allowed to settle for up to 5 days.

4. A process as claimed in any of claims 1 to 3 wherein the nanostructure material is selected from any one or more of carbon nanotubes, non-carbon nanotubes, nanowires, nanorods and nanoparticles.

5. A process as claimed in claim 4 wherein the carbon nanotubes are selected from any one or more of single walled nanotubes (SWNT), double walled nanotubes (DWNT) and multi walled nanotubes (MWNT).

6. A process as claimed in any preceding claim wherein the biological compound is a biopolymer.

7. A process as claimed in claim 6 wherein the biopolymer is any macromolecule comprising a protein, nucleic acid, polysaccharide or polyelectrolyte thereof.

8. A process as claimed in claim 6 or 7 wherein the biopolymer is selected from any one or more of glycosaminoglycan, enzyme, DNA, oligonucleotide, amino acids and chitin.

9. A process as claimed in claim 8 wherein the glycosaminoglycan is selected from any one or more of chondroitin sulphate, heteropolysaccharides, polysaccharides and disaccharides.

10. A process as claimed in claim 8 wherein the chitin is chitosan, preferably 1,4 alpha D glucosamine polymer.

11. A process as claimed in any of claims 7 to 10 wherein the macromolecule is of bacterial origin.

12. A process as claimed in any preceding claim wherein the biological compound is dissolved in water and the nanostructure material preparation is added to the solution.

13. A process as claimed in claim 12 wherein the biological compound is dissolved in water at a temperature of approximately 90°C.

14. A process as claimed in any preceding claim wherein the biological compound is present in a weight ratio of at least 1:1 with respect to the nanostructure material.

15. A process as claimed in any preceding claim wherein the biological compound is present in a weight ratio of at least 2:1 with respect to the nanostructure material.

16. A process as claimed in any preceding claim wherein the biological compound is present in a weight ratio of at least 3:1 with respect to the nanostructure material.

17. A process as claimed in any preceding claim wherein the biological compound is present in a weight ratio of at least 4:1 with respect to the nanostructure material.

18. A process as claimed in any preceding claim wherein the biological compound is present in a weight ratio of up to 10:1 with respect to the nanostructure material.

19. A process as claimed in any preceding claim comprising the step of obtaining the nanostructure material from the nanostructure composite material dispersion.

20. A process as claimed in claim 19 wherein the nanostructure material is obtained from the composite material dispersion by heating the dispersion to a temperature less than the combustion/decomposition temperature of the nanotube content of the nanostructure material.

21. A process as claimed in claim 19 or 20 wherein the purity of the isolated nanostructure material is greater than 20%.

22. A process as claimed in claims 19 to 21 wherein the purity of the isolated nanostructure material is greater than 30%.

23. A process as claimed in claims 19 to 22 wherein the purity of the isolated nanostructure material is greater than 40%.

24. A process as claimed in claims 19 to 23 wherein the purity of the isolated nanostructure material is greater than 50%.

25. A process as claimed in any of claims 19 to 24 wherein the isolated nanostructure material has an impurity level of less than 4%.

26. A nanostructure composite material whenever prepared by a process as claimed in any of claims 1 to 18.

27. A nanostructure material whenever prepared by a process as claimed in any of claims 19 to 25.

28. Carbon nanotubes whenever prepared by a process as claimed in any of claims 1 to 25.

29. A composite comprising a nanostructure material; and a biological compound, wherein the biological compound interacts with the nanostructure material.

30. A composite comprising a nanostructure material; and a biological compound, wherein the biological compound is separable from the nanostructure material to provide a purified nanostructure material.

31. A composition comprising a biological compound and a nanostructure material.

32. A composite substantially as hereinbefore described with reference to the examples and accompanying drawings.

33. A nanostructure material substantially as hereinbefore described with reference to the examples and accompanying drawings.

34. Use of a composite as claimed in any of claims 26 or 29 to 32 in the manufacture of conductive and high strength composites, energy storage and energy conversion devices, sensors, field emission displays and radiation sources, hydrogen storage media, photovoltaic devices, actuators and devices containing nanometer-sized entities/components/structures, probes and interconnects, fuel cells, dry delivery, biomedical applications including tissue engineering, optics including non-linear optics, biocompatible composites, fibres, mats or papers, biocompatible coatings, devices or sensors.

35. Use of a purified nanostructure material as claimed in any of claims 27 or 33 in the manufacture of conductive and high strength composites, energy storage and energy conversion devices, sensors, field emission displays and radiation sources, hydrogen storage media, photovoltaic devices, actuators and devices containing nanometer-sized entities/components/structures, probes and interconnects, fuel cells, dry delivery, biomedical applications including tissue engineering, optics including non-linear optics, biocompatible composites, fibres, mats or papers, biocompatible coatings, devices or sensors.