EP2536395A1 - Preparation of functionalized nano/micro-structures by self-assembly of biomolecules - Google Patents

Preparation of functionalized nano/micro-structures by self-assembly of biomolecules

Info

Publication number
EP2536395A1
EP2536395A1 EP11707116A EP11707116A EP2536395A1 EP 2536395 A1 EP2536395 A1 EP 2536395A1 EP 11707116 A EP11707116 A EP 11707116A EP 11707116 A EP11707116 A EP 11707116A EP 2536395 A1 EP2536395 A1 EP 2536395A1
Authority
EP
European Patent Office
Prior art keywords
micro
nano
structures
self
hydrophobic
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP11707116A
Other languages
German (de)
French (fr)
Inventor
Niclas Solin
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Individual
Original Assignee
Individual
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Individual filed Critical Individual
Publication of EP2536395A1 publication Critical patent/EP2536395A1/en
Withdrawn legal-status Critical Current

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/141—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers
    • A61K9/143—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers with inorganic compounds
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/22—Hormones
    • A61K38/28—Insulins
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B82—NANOTECHNOLOGY
    • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y40/00—Manufacture or treatment of nanostructures

Definitions

  • the present invention relates generally to preparation of nano/micro- structures by use of self-assembly of a composite material formed from bio-molecules and guest molecules.
  • Typical desired properties include, for example, light emission or conduction of electricity.
  • many molecules used to infer such conducting or emissive properties contain a large proportion of hydrocarbons (which are hydrophobic), thus rendering them insoluble in aqueous media. This means that such molecules have an orthogonal solubility with respect to the bio-molecules, which typically are water soluble.
  • self-assembly processes of bio-molecules primarily occur in aqueous solvent; a solvent in which thus the functionalization agent has an intrinsically low solubility.
  • Another approach is to functionalize the self assembled nano/micro- structure; either after the self-assembly process is completed, or during the self- assembly process (US200803875 1 ) .
  • the resulting water- soluble guest molecule must have a high affinity for the self assembled nano/micro- structure, as otherwise the molecule will not interact with the structure and instead prefer to stay dissolved in the solvent rather than binding to the nano/micro- structure.
  • the present invention provides a solution to the problem by the method of preparing a composite material comprising bio-molecules and
  • An object of the present application is to provide means and methods that meet functional needs. This objective is in a first aspect achieved by self-assembly of a composite material.
  • the composite material is formed by grinding of a bio- molecular material, which can undergo self-assembly, with a guest molecule. It is known that co-grinding of a hydrophobic material together with a hydrophilic material render a composite material, but the material is often water soluble
  • the bio-molecule has high solubility in aqueous solvent, and the guest molecule has low solubility in aqueous solvent.
  • the composite material is then dissolved in aqueous solvent, and the dissolved composite material is exposed to conditions that promote the self-assembly of the bio-molecule into nano/micro- structures.
  • the present invention relates to methods for the
  • a composite material consisting of bio-molecules that have been mixed with guest molecules.
  • the method of mixing the bio-molecule and the guest molecule is in general terms to adsorb the guest molecule onto the bio-polymer, resulting in a composite material. This adsorption may be achieved by grinding of the two components in the solid state, but any other method (e.g. jet milling) resulting in a similar type of composite material may be used in place of grinding.
  • the composite material can be dissolved in aqueous solvent, and the dissolved composite material is exposed to conditions that promote the self-assembly of the bio-molecule into micro/nano-structures.
  • Examples of such self-assembled micro/nano-structures are amyloid-like fibrils, which can be formed by self-assembly of proteins.
  • the attached guest molecule is carried along into the final self-assembled structure.
  • it is desirable that the guest molecule has a low solubility in the aqueous solvent used for the self assembly reaction, as this will force the guest molecule to stay attached to the self-assembling component during the self- assembly process.
  • the functionalization material preferably consists of guest molecules which are hydrophobic, as such molecules are insoluble, or have a low solubility in aqueous media.
  • guest molecules which are hydrophobic, as such molecules are insoluble, or have a low solubility in aqueous media.
  • An object of the present invention is to provide nano/micro-structures, consisting of a material formed by a self-assembly process of a water soluble composite material, consisting of a bio-molecular material and a hydrophobic material, where said self-assembly process occurs in aqueous solvent and said composite material is formed by grinding of a water soluble bio-molecular material capable of undergoing self-assembly, where said bio-molecular material consists of: i) proteins; or ii) nucleic acids; or iii) peptide chains, and said grinding is done in the presence of hydrophobic materials consisting of: i) hydrophobic molecules containing unsaturated or saturated hydrocarbon units (including fullerenes or other types of carbon structures; or ii) hydrophobic nano-particles; or iii) hydrophobic inorganic molecules.
  • the bio-molecule is insulin.
  • said hydrophobic material consists of dye molecules, whereby the self-assembly process results in nano/micro- structures capable of being luminescent.
  • the dye molecule is lr(ppy) 3 or I r(piq) 3 or both.
  • the hydrophobic material has a capacity of conducting electricity whereby the self-assembly process results in nano/micro-structures capable of conducting electricity.
  • the hydrophobic material has magnetic properties, whereby the self-assembly process results in nano/micro- structures having magnetic properties.
  • the hydrophobic material consists of drug molecules, whereby the self-assembly process results in nano/micro- structures comprising drug molecules.
  • the employed hydrophobic materials are a combination of different hydrophobic materials, whereby the self-assembly process results in nano/micro-structures having a combination of two or more properties described above.
  • the bio-molecule is a protein or oligopeptide or polypeptide, all must be capable of undergoing self- assembly into amyloid-like structures, resulting in the formation of amyloid-like nano/micro-structures with properties as described above.
  • Another purpose of the present application is to provide a method for the formation of nano/micro structures comprising a bio-molecule and a hydrophobic material comprising the steps of: a) providing water soluble bio-molecules capable of undergoing self assembly processes consisting of i) proteins; or ii) nucleic acids; or iii) peptide chains b) providing hydrophobic material consisting of i) hydrophobic molecules containing unsaturated or saturated hydrocarbon units (including fullerenes or other types of carbon structures); or ii) hydrophobic nano-particles; or iii) hydrophobic inorganic molecules c) grinding the water soluble bio molecules according to a) in the presence of the hydrophobic material according to b) in the solid state thereby obtaining a water soluble composite material capable of undergoing self assembly processes in an aqueous solution into nano/micro structures having the hydrophobic material incorporated.
  • the grinding in the method described above may be performed in the presence of additional additives such as solvents or salts, or the materials to be grinded are pre-treated by melting them together or one or more of the components are added as a solution, and the grinding is commenced after or during
  • the bio-molecule may be chosen from the group consisting of proteins or oligo-peptides or poly-peptides that are capable of undergoing self-assembly into amyloid-like structures resulting in the formation of amyloid-like nano/micro- structures with properties as mentioned above.
  • bio-molecule insulin In one special embodiment is the bio-molecule insulin.
  • the present invention relates to a variety of hydrophobic molecules or materials, such as hydrophobic molecules containing unsaturated or saturated hydrocarbon units (including fullerenes or other types of carbon structures); or ii) hydrophobic nano-particles; or iii) hydrophobic inorganic molecules.
  • hydrophobic molecules or material i.e., guest molecules have an orthogonal solubility relative to the bio-molecule. If the bio-molecule is water soluble, a guest molecule with low solubility in water is preferably used. Thus, if a self assembler is soluble in a certain solvent, the guest molecule should have a low solubility in that solvent.
  • the present invention is related to a wide variety of bio-molecules, and appropriate bio-molecules include, but are not limited to, peptides, nucleic acids and DNA, proteins, and any organic polymers or combination of these molecules.
  • bio-molecules include, but are not limited to, peptides, nucleic acids and DNA, proteins, and any organic polymers or combination of these molecules.
  • the key aspect of the bio-molecule used is that it must be soluble, and be able to undergo self-assembly processes, thereby resulting in the formation of structures in the nano/micro-dimension.
  • the bio-molecule must be able to form a water-soluble composite material with the guest molecule of interest.
  • other molecules or polymers than bio-molecules may be used in a similar fashion, as long as they have the capacity of undergoing self-assembly forming nano/micro- structures.
  • guest molecule means a molecule, which by intermolecular interactions or a covalent bond, forms a complex with a molecule, or ensemble of molecules, capable of undergoing self-assembly. It can also refer to a molecule interacting with a nano-microstructure formed by self assembly.
  • bio-molecule means any organic molecule that is produced by living organisms, or man-made molecules inspired by such bio-molecules, such as man-made peptides or nucleotides.
  • hydrophobic material means a material made up of molecules with low solubility in water.
  • nano/micro-structure is a structure that has dimensions in the nanometer-micrometer range.
  • self-assembly process means the spontaneous and reversible association of molecular species to form larger supramolecular entities according to the intrinsic information contained in the components.
  • amyloid-like structures means structures made up from molecules capable of forming amyloid structures that have been modified by the incorporation of other components such as guest molecules.
  • composite material means a material made up of two or more components.
  • the expression "functionalized” means that molecules having desired properties are added to a structure by the formation of a molecular complex between the molecules and the structure. The molecule thus becomes a part of the structure, thereby lending its properties to the structure. The properties of the molecule may, or may not, be altered by the addition to the structure.
  • dye molecules it is possible to prepare luminescent nano/micro- structures; by incorporating conducting or semi-conducting guest molecules it is possible to prepare conducting, or semiconducting nanostructures, respectively; by incorporating magnetic nanoparticles, or magnetic molecules, it is possible to prepare magnetic nanostructures; by incorporating redox-active materials, it is possible to prepare redox-active nanomaterials with tuned redox properties; by incorporating drug molecules, it is possible to prepare nano/micro-structure of potential use as drug delivery agents; by incorporating metal complexes with catalytic capacity, it is possible to prepare materials with tuned catalytic properties, either as prepared, or after further modification.
  • the range of application for the present invention is numerous.
  • One advantage of the present nano/micro structure is that it passes the brain-blood-barrier, a tremendous advantage for the design of drug delivery to the CNS.
  • the present invention is the function and/or predetermined release of the active molecule (guest molecule) initiated by a change in the environment, such as for example heat, pH or pressure changes.
  • a change in the environment such as for example heat, pH or pressure changes.
  • the release is initiated by low pH.
  • [0039] in yet another embodiment is the release of a drug or other compound (guest molecule) controlled, e.g., slower release, faster release or pulsative release of the guest molecule.
  • a drug or other compound guest molecule controlled, e.g., slower release, faster release or pulsative release of the guest molecule.
  • Another embodiment of the present invention is in applications were the "guest molecule" is protected against degradation or other threats that destroy the function, and therefore prolong its life time.
  • nano/micro-structures which combine two (or more) of the different types of guest molecule can be prepared, and nano/micro-structures with more than one selected property may thus be prepared.
  • nano/micro- structures which show synergistic effects from properties derived from two or more types of guest molecules may be prepared. Synergistic effects may also occur as a result of the guest molecule being incorporated into the nano/micro-structure, or as a result of interactions between the bio-molecule and the guest molecule.
  • the na no/micro-structures will posses different physical properties which may be of value in a range of applications.
  • Na no/micro-structures incorporating dye-molecule materials can be incorporated into light-emitting devices, such as OLEDs.
  • Nano/micro-structures including conducting guest materials can be incorporated into devices requiring conducting components in the nano/micro-scale.
  • Nano/micro-structures including magnetic guest materials can be incorporated into devices requiring magnetic components in the nano/micro-scale.
  • Such materials may also be of use as magnetic resonance imaging contrast agents, whereby the distribution of the nano/micro-structure in an organism may be followed.
  • the nano/micro-structure may also include hydrophobic drug-molecules, whereby the nano/micro-structures can be used as drug delivery agents.
  • the present invention thus solves the problem of functionalizing
  • nano/micro-structures formed by combining bio-molecular self assembly with guest molecules of low solubility in aqueous media. Furthermore, the operational simplicity and the suitability of the method for scale-up makes it a competitive choice for preparation of functionalized nano/micro-structures when compared to previously reported methods which rely on synthetic chemistry modifications of existing chemical substances.
  • the nano/micro-structures can be prepared from commercially available chemical substances which, in contrast to previously reported methods, can be used without any further synthetic
  • a further aspect of the invention provides the possibility to prepare a variety of nano/micro-structures which will have properties derived from the guest molecule, and the incorporation of these nano/micro-structures into devices of various sorts. Moreover, as the guest molecules are not attached to the nano-micro structure by covalent bonds they may under appropriate conditions be released to the surroundings. The ⁇ -microstructure may therefore function as a container for molecules to be released to the surroundings.
  • Figure 1 show chemical structures of iridium complexes used for functionalization of na no/micro-structures made from self-assembly of insulin.
  • the iridium-complexes mentioned in the descriptions are tris[2- phenylpyridinato-C , ]iridiurn(lll), abbreviated as (lr(ppy) 3 ) or (1 ), and tris[l - phenylisoquinoline-C , ]iridiurn(lll) abbreviated as (lr(piq) 3 ) or (2).
  • Figure 2 shows AFM and fluorescence microscope characterization of nano/micro-structures formed by self-assembly of a composite material obtained by grinding insulin with complexes lr(ppy) 3 (1 ) or I r(piq) 3 (2). Fibrils formed from insulin grinded with a) complex lr(ppy) 3 (1 ) and b) complex I r(piq) 3 (2). c), d) Fluorescence microscope images of fibrils formed from insulin grinded with c) complex lr(ppy) 3 (1 ) and d) complex lr(piq) 3 (2).
  • Figure 3 shows examples of nano/micro-structures formed by self-assembly of a composite material obtained by grinding insulin with complexes lr(ppy) 3 (1 ) or lr(piq) 3 (2).
  • the images are fluorescence microscope images of a), b) spherulites and c), d) aggregated amyloid-like fibrils.
  • Figure 4 shows emission data from lr(ppy) 3 (1 ) and I r(piq) 3 (2) as free complexes in CHCI 3 solution, and in samples exposed to fibrillation conditions.
  • the top curve complex was exited at 380 nm; bottom curve complex was excited at 425 nm.
  • Figure 5 shows FT-Raman spectra recorded for powders of insulin, ground insulin, and the composite materials obtained by grinding insulin with metal-complex lr(ppy) 3 (1 ) or lr(piq) 3 (2).
  • Figure 6 shows Circular Dichroism spectra of solutions in 25 mM HCI incubated for 0 and 8 hours. Spectra are shown for insulin; ground insulin; and insulin ground with either of complex lr(ppy) 3 (1 ) or I r(piq) 3 (2).
  • Figure 7 shows MALDI-TOF spectra obtained on the composite material formed by grinding insulin with either of complex lr(ppy) 3 (1 ) or I r(piq) 3 (2).
  • the present invention relates to methods of preparing functionalized nano/micro-structures based on self-assembly of a composite material consisting of functionalized bio-molecules.
  • the bio-molecule material (the self assembler) and the guest molecule material (the functionalizing agent) are grinded together in the solid state.
  • additives such as solvent, or other materials, may be added to facilitate the grinding.
  • the materials may be pretreated with a variety of methods, such as melting the two components before grinding, or adding the guest molecule as a solution in organic solvent, and grinding the resulting material after, or during, the evaporation of the organic solvent.
  • the grinding may be performed with any kind of milling equipment, including simply a mortar and pestle. The grinding process results in a water soluble composite material formed by the bio-molecule and the guest molecule.
  • the grinding process may result in chemical changes, for example the formation or cleavage of covalent bonds, as long as the resulting composite material is still capable of undergoing self assembly processes.
  • the resulting composite material is dissolved into an appropriate solvent, or mixture of solvents, and is then exposed to conditions that promote self-assembly of the bio- molecule.
  • the resulting solution, dispersion, or precipitation of functionalized nano/micro-structures can be immobilized on a variety of solid supports, including, but not limited to silicon wafers, glass (e. g. glass slides, glass beads, glass wafers etc., silicon rubber, polystyrene, polyethylene, teflon, silica gel beads, gold, indium tin oxide (ITO coated materials, e.
  • Transfer of the functionalized micro-nanostructures to the solid support can be achieved by using, but not limited to, dip coating, spin- coating, microcontact printing, screen printing, ink jet technologies, spraying, dispensing and microfluidic printing by the use of soft lithography. Solvents for the functionalized micro-nanostructures of the present invention during the
  • immobilization to the solid support can be, but are not limited to, water, buffered water solutions, methanol, ethanol, chloroform and combinations thereof.
  • iridium-complexes mentioned in the experimental descriptions below are tris[2-phenylpyridinato-C , ]iridium(lll), abbreviated as (lr(ppy) 3 ) or (1 ), and tris[l -phenylisoquinoline-C , ]iridiurn(lll) abbreviated as (lr(piq) 3 ) or (2) (see figure 1 for structural drawings of the complexes)
  • the solution was then diluted with 25mM HCI to final concentrations of 4.2-4.6mg/ml of insulin and 0.07-0.08mg/ml of complex lr(ppy) 3 (1 ) or I r(piq) 3 (2).
  • the samples were then heated at 65°C for 40-72 hours in order to induce fibril formation.
  • the sample was prepared by drop casting the fibrillated bovine insulin/lr-complex onto Si/Si0 2 substrates.
  • AFM characterization of nano/microstructures formed by self-assembly of a composite material formed by grinding of insulin and iridium-complexes ( Figure 2).
  • Atomic force microscopy (AFM) imaging was performed on samples of the self-assembled bovine insulin/lr-complex prepared as given in example 1 . Images were recorded by standard procedures in tapping mode on a SFM-Nanoscope III, Digital Instruments, with a J scanner. Cantilevers for tapping mode were obtained from NT-MDT.
  • amyloid-like fibrils were deposited onto Si/Si02 substrates by molecular combing : 1 ⁇ of a drop of the fibrillated bovine insulin/lr-complex solution was deposited onto a Si/Si0 2 substrate, incubated for 1 minute and then gently blown off the substrate using a nitrogen gas flow.
  • the photoluminescence was measured in CHCI 3 solutions for complexes lr(ppy) 3 ( 1 ) or Ir (piq) 3 (2), and in 25 mM HCI solutions of the self-assembled composite material, prepared as given in example 1 .
  • the spectra were recorded with an ISA Jobin-Yvon Spex FluoroMax2 fluorimeter at excitation wavelength of 380nm and 425 nm for lr(ppy) 3 ( 1 ) and lr(piq) 3 (2), respectively.
  • a composite material was prepared by grinding insulin and iridium complexes. The resulting material was dissolved in 25mM HCI, and aliquots were withdrawn, and analyzed by CD. The rest of the sample was heated at 65°C / and after 8 hours the samples were again examined by CD. Samples prepared from native insulin, and ground native insulin was analyzed in a similar way. The far- Ultraviolet (UV) circular dichroism (CD) spectra ( ⁇ 0.1 25mg/ml protein
  • AAALDI-TOF MS measurements were performed with a Voyager-DE instrument equipped with a 337nm nitrogen laser, a-cyano-4-hydroxycinnamic acid (CHCA) was used as matrix.
  • CHCA a-cyano-4-hydroxycinnamic acid
  • a solution of the composite material (insulin and iridium-complexes) in 25mM HCI was prepared, and was then filtered through a 0.2 ⁇ PVDF filter.
  • the sample was then diluted with water, and this solution was mixed 1 : 1 with a saturated solution of CHCA in 50:50 acetonitrile/water (0.1 % TFA).
  • 0.5 ⁇ of the resulting solution was co-evaporated on a target plate, and the spectra were recorded in a positive reflector mode.
  • MS data were acquired using the instrument default calibration, without employing internal or external calibration.

Landscapes

  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Veterinary Medicine (AREA)
  • General Health & Medical Sciences (AREA)
  • Nanotechnology (AREA)
  • Public Health (AREA)
  • Animal Behavior & Ethology (AREA)
  • Epidemiology (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Diabetes (AREA)
  • Condensed Matter Physics & Semiconductors (AREA)
  • Manufacturing & Machinery (AREA)
  • Crystallography & Structural Chemistry (AREA)
  • Inorganic Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • General Physics & Mathematics (AREA)
  • Endocrinology (AREA)
  • Zoology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Immunology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Abstract

The invention relates to methods of preparing structures in the micro/nano- dimension by self-assembly of a composite material formed from bio-molecules and guest molecules. These nano/micro-structures have properties derived from the guest molecule, and their structure will be influenced by the bio-molecule.

Description

Preparation of functionalized nano/micro-structures by self-assembly of bio- molecules
Technical field
[0001 ] The present invention relates generally to preparation of nano/micro- structures by use of self-assembly of a composite material formed from bio-molecules and guest molecules.
Background art
[0002] The use of bottom up methods to assemble structures in the nano/micro- dimension has been researched for decades. Much research has focused on the design of man-made molecules capable of self assembly into such nano/micro- structures.
[0003] For most applications of the resulting nano/micro-structures, it is essential to somehow functionalize the na no/micro-structure with guest molecules, which possess desired physical properties for the desired application. Typical desired properties include, for example, light emission or conduction of electricity. However, many molecules used to infer such conducting or emissive properties contain a large proportion of hydrocarbons (which are hydrophobic), thus rendering them insoluble in aqueous media. This means that such molecules have an orthogonal solubility with respect to the bio-molecules, which typically are water soluble. Furthermore, self-assembly processes of bio-molecules primarily occur in aqueous solvent; a solvent in which thus the functionalization agent has an intrinsically low solubility.
[0004] To circumvent the solubility-problem, two approaches are commonly used in the art. One approach is to attach the guest molecule to the bio-polymer with a covalent bond by means of traditional chemical synthesis methods. The resulting material is then exposed to conditions that promote self-assembly of the bio- molecule, resulting in a self assembled nano/micro-structure, having properties rendered by the guest molecule. The drawback of this method is that it requires large amount of synthetic work to prepare the necessary building block to be used in the self assembly process.
[0005] Another approach is to functionalize the self assembled nano/micro- structure; either after the self-assembly process is completed, or during the self- assembly process (US200803875 1 ) . In this case it is necessary to render the guest molecule used for functionalization water soluble. Moreover, the resulting water- soluble guest molecule must have a high affinity for the self assembled nano/micro- structure, as otherwise the molecule will not interact with the structure and instead prefer to stay dissolved in the solvent rather than binding to the nano/micro- structure.
[0006] One commonly used approach in the art is to render the desired guest molecules water soluble by adding polar functional groups to the guest molecule.
[0007] By modifying the guest molecule one way or another as described above often reduce or change the inherent properties and also limits its range of applications.
[0008] Accordingly, it remains technical problems to adapt hydrophobic guest molecules to self assembling structures in order to fully exploit potential applications of this technology. The present invention provides a solution to the problem by the method of preparing a composite material comprising bio-molecules and
hydrophobic guest molecules that self-assembly into nano/micro-structures when dissolved in aqueous solvent, which can be fully applicable in numerous fields of utility. Summary of invention
[0009] An object of the present application is to provide means and methods that meet functional needs. This objective is in a first aspect achieved by self-assembly of a composite material. The composite material is formed by grinding of a bio- molecular material, which can undergo self-assembly, with a guest molecule. It is known that co-grinding of a hydrophobic material together with a hydrophilic material render a composite material, but the material is often water soluble
(GB701 81 4).
[001 0] Preferably the bio-molecule has high solubility in aqueous solvent, and the guest molecule has low solubility in aqueous solvent. The composite material is then dissolved in aqueous solvent, and the dissolved composite material is exposed to conditions that promote the self-assembly of the bio-molecule into nano/micro- structures.
[001 1 ] In general terms, the present invention relates to methods for the
preparation of structures in the nano/micro-dimension by using self-assembly of a composite material, consisting of bio-molecules that have been mixed with guest molecules. The method of mixing the bio-molecule and the guest molecule is in general terms to adsorb the guest molecule onto the bio-polymer, resulting in a composite material. This adsorption may be achieved by grinding of the two components in the solid state, but any other method (e.g. jet milling) resulting in a similar type of composite material may be used in place of grinding. The composite material can be dissolved in aqueous solvent, and the dissolved composite material is exposed to conditions that promote the self-assembly of the bio-molecule into micro/nano-structures. Examples of such self-assembled micro/nano-structures are amyloid-like fibrils, which can be formed by self-assembly of proteins. As a part of the self-assembly process, the attached guest molecule is carried along into the final self-assembled structure. In order to ensure the incorporation of the guest molecule into the final na no/micro-structure, it is desirable that the guest molecule has a low solubility in the aqueous solvent used for the self assembly reaction, as this will force the guest molecule to stay attached to the self-assembling component during the self- assembly process. Thus, for the present invention, the functionalization material preferably consists of guest molecules which are hydrophobic, as such molecules are insoluble, or have a low solubility in aqueous media. As an example of molecules exhibiting the above characteristics, tris[2-phenylpyridinato- C , ]iridium(lll) (lr(ppy)3) and tris[l -phenylisoquinoline-C , ]iridiurn(lll) (lr(piq)3) (see Figure 1 ) can be mentioned.
[001 2] An object of the present invention is to provide nano/micro-structures, consisting of a material formed by a self-assembly process of a water soluble composite material, consisting of a bio-molecular material and a hydrophobic material, where said self-assembly process occurs in aqueous solvent and said composite material is formed by grinding of a water soluble bio-molecular material capable of undergoing self-assembly, where said bio-molecular material consists of: i) proteins; or ii) nucleic acids; or iii) peptide chains, and said grinding is done in the presence of hydrophobic materials consisting of: i) hydrophobic molecules containing unsaturated or saturated hydrocarbon units (including fullerenes or other types of carbon structures; or ii) hydrophobic nano-particles; or iii) hydrophobic inorganic molecules.
[001 3] In one embodiment of the nano/micro-structure, the bio-molecule is insulin.
[001 4] In one embodiment of the nano/micro-structure, said hydrophobic material consists of dye molecules, whereby the self-assembly process results in nano/micro- structures capable of being luminescent.
[001 5] In another embodiment of the nano/micro-structure, the dye molecule is lr(ppy)3 or I r(piq)3 or both. [001 6] In one embodiment of the nano/micro-structure, the hydrophobic material has a capacity of conducting electricity whereby the self-assembly process results in nano/micro-structures capable of conducting electricity.
[001 7] In one embodiment of the nano/micro-structure, the hydrophobic material has magnetic properties, whereby the self-assembly process results in nano/micro- structures having magnetic properties.
[001 8] In one embodiment of the nano/micro-structure, the hydrophobic material consists of drug molecules, whereby the self-assembly process results in nano/micro- structures comprising drug molecules.
[001 9] In yet another embodiment of the nano/micro-structure, the employed hydrophobic materials are a combination of different hydrophobic materials, whereby the self-assembly process results in nano/micro-structures having a combination of two or more properties described above.
[0020] In one embodiment of the nano/micro-structure, the bio-molecule is a protein or oligopeptide or polypeptide, all must be capable of undergoing self- assembly into amyloid-like structures, resulting in the formation of amyloid-like nano/micro-structures with properties as described above.
[0021 ] Another purpose of the present application is to provide a method for the formation of nano/micro structures comprising a bio-molecule and a hydrophobic material comprising the steps of: a) providing water soluble bio-molecules capable of undergoing self assembly processes consisting of i) proteins; or ii) nucleic acids; or iii) peptide chains b) providing hydrophobic material consisting of i) hydrophobic molecules containing unsaturated or saturated hydrocarbon units (including fullerenes or other types of carbon structures); or ii) hydrophobic nano-particles; or iii) hydrophobic inorganic molecules c) grinding the water soluble bio molecules according to a) in the presence of the hydrophobic material according to b) in the solid state thereby obtaining a water soluble composite material capable of undergoing self assembly processes in an aqueous solution into nano/micro structures having the hydrophobic material incorporated.
[0022] The grinding in the method described above may be performed in the presence of additional additives such as solvents or salts, or the materials to be grinded are pre-treated by melting them together or one or more of the components are added as a solution, and the grinding is commenced after or during
evaporation of said solvent.
[0023] The bio-molecule may be chosen from the group consisting of proteins or oligo-peptides or poly-peptides that are capable of undergoing self-assembly into amyloid-like structures resulting in the formation of amyloid-like nano/micro- structures with properties as mentioned above.
[0024] In one special embodiment is the bio-molecule insulin.
[0025] The present invention relates to a variety of hydrophobic molecules or materials, such as hydrophobic molecules containing unsaturated or saturated hydrocarbon units (including fullerenes or other types of carbon structures); or ii) hydrophobic nano-particles; or iii) hydrophobic inorganic molecules. It is considered beneficial if the hydrophobic molecules or material, i.e., guest molecules have an orthogonal solubility relative to the bio-molecule. If the bio-molecule is water soluble, a guest molecule with low solubility in water is preferably used. Thus, if a self assembler is soluble in a certain solvent, the guest molecule should have a low solubility in that solvent. By the use of a guest molecule with certain properties it is possible to give the final self-assembled nano/micro-structure different properties.
[0026] The present invention is related to a wide variety of bio-molecules, and appropriate bio-molecules include, but are not limited to, peptides, nucleic acids and DNA, proteins, and any organic polymers or combination of these molecules. The key aspect of the bio-molecule used is that it must be soluble, and be able to undergo self-assembly processes, thereby resulting in the formation of structures in the nano/micro-dimension. Moreover, the bio-molecule must be able to form a water-soluble composite material with the guest molecule of interest. Furthermore, other molecules or polymers than bio-molecules may be used in a similar fashion, as long as they have the capacity of undergoing self-assembly forming nano/micro- structures.
[0027] The term guest molecule means a molecule, which by intermolecular interactions or a covalent bond, forms a complex with a molecule, or ensemble of molecules, capable of undergoing self-assembly. It can also refer to a molecule interacting with a nano-microstructure formed by self assembly.
[0028] The term bio-molecule means any organic molecule that is produced by living organisms, or man-made molecules inspired by such bio-molecules, such as man-made peptides or nucleotides.
[0029] The term hydrophobic material means a material made up of molecules with low solubility in water.
[0030] The term nano/micro-structure is a structure that has dimensions in the nanometer-micrometer range. [0031 ] The term self-assembly process means the spontaneous and reversible association of molecular species to form larger supramolecular entities according to the intrinsic information contained in the components.
[0032] The term amyloid-like structures means structures made up from molecules capable of forming amyloid structures that have been modified by the incorporation of other components such as guest molecules.
[0033] The term composite material means a material made up of two or more components.
[0034] The expression "functionalized" means that molecules having desired properties are added to a structure by the formation of a molecular complex between the molecules and the structure. The molecule thus becomes a part of the structure, thereby lending its properties to the structure. The properties of the molecule may, or may not, be altered by the addition to the structure.
[0035] By preparing a nano/micro-structure by self-assembly of a composite material, which has been prepared from a bio-molecule material and a guest molecule, it is possible to obtain a nano/micro-structure with selected physical properties, as a result of the physical properties of the guest molecule. Thus by incorporating dye molecules it is possible to prepare luminescent nano/micro- structures; by incorporating conducting or semi-conducting guest molecules it is possible to prepare conducting, or semiconducting nanostructures, respectively; by incorporating magnetic nanoparticles, or magnetic molecules, it is possible to prepare magnetic nanostructures; by incorporating redox-active materials, it is possible to prepare redox-active nanomaterials with tuned redox properties; by incorporating drug molecules, it is possible to prepare nano/micro-structure of potential use as drug delivery agents; by incorporating metal complexes with catalytic capacity, it is possible to prepare materials with tuned catalytic properties, either as prepared, or after further modification.
[0036] In addition, the range of application for the present invention is numerous. One can mention designed pharmaceuticals, designed for being delivered to a certain cell type, receptor or organ or as a research tool, or as a diagnostic tool.
[0037] One advantage of the present nano/micro structure is that it passes the brain-blood-barrier, a tremendous advantage for the design of drug delivery to the CNS.
[0038] In one embodiment of the present invention is the function and/or predetermined release of the active molecule (guest molecule) initiated by a change in the environment, such as for example heat, pH or pressure changes. For example, if the purpose is to release a drug molecule in the gut, the release is initiated by low pH.
[0039] In yet another embodiment is the release of a drug or other compound (guest molecule) controlled, e.g., slower release, faster release or pulsative release of the guest molecule.
[0040] Another embodiment of the present invention is in applications were the "guest molecule" is protected against degradation or other threats that destroy the function, and therefore prolong its life time.
[0041 ] Moreover, nano/micro-structures which combine two (or more) of the different types of guest molecule can be prepared, and nano/micro-structures with more than one selected property may thus be prepared. Moreover, nano/micro- structures which show synergistic effects from properties derived from two or more types of guest molecules may be prepared. Synergistic effects may also occur as a result of the guest molecule being incorporated into the nano/micro-structure, or as a result of interactions between the bio-molecule and the guest molecule. Depending on the choice of guest molecule material, the na no/micro-structures will posses different physical properties which may be of value in a range of applications.
[0042] Na no/micro-structures incorporating dye-molecule materials can be incorporated into light-emitting devices, such as OLEDs. Nano/micro-structures including conducting guest materials can be incorporated into devices requiring conducting components in the nano/micro-scale. Nano/micro-structures including magnetic guest materials can be incorporated into devices requiring magnetic components in the nano/micro-scale. Such materials may also be of use as magnetic resonance imaging contrast agents, whereby the distribution of the nano/micro-structure in an organism may be followed. The nano/micro-structure may also include hydrophobic drug-molecules, whereby the nano/micro-structures can be used as drug delivery agents.
[0043] The present invention thus solves the problem of functionalizing
nano/micro-structures formed by combining bio-molecular self assembly with guest molecules of low solubility in aqueous media. Furthermore, the operational simplicity and the suitability of the method for scale-up makes it a competitive choice for preparation of functionalized nano/micro-structures when compared to previously reported methods which rely on synthetic chemistry modifications of existing chemical substances. In the present method, the nano/micro-structures can be prepared from commercially available chemical substances which, in contrast to previously reported methods, can be used without any further synthetic
modifications. A further aspect of the invention provides the possibility to prepare a variety of nano/micro-structures which will have properties derived from the guest molecule, and the incorporation of these nano/micro-structures into devices of various sorts. Moreover, as the guest molecules are not attached to the nano-micro structure by covalent bonds they may under appropriate conditions be released to the surroundings. The ηαηο-microstructure may therefore function as a container for molecules to be released to the surroundings.
[0044] It is to be understood that both the foregoing general description and the following detailed description are exemplary, and are intended to provide further explanation of the invention as claimed.
Brief description of drawings
[0045] The invention is now described, by way of example, with reference to the accompanying drawings, in which:
[0046] Figure 1 show chemical structures of iridium complexes used for functionalization of na no/micro-structures made from self-assembly of insulin.
[0047] The iridium-complexes mentioned in the descriptions are tris[2- phenylpyridinato-C , ]iridiurn(lll), abbreviated as (lr(ppy)3) or (1 ), and tris[l - phenylisoquinoline-C , ]iridiurn(lll) abbreviated as (lr(piq)3) or (2).
[0048] Figure 2 shows AFM and fluorescence microscope characterization of nano/micro-structures formed by self-assembly of a composite material obtained by grinding insulin with complexes lr(ppy)3 (1 ) or I r(piq)3 (2). Fibrils formed from insulin grinded with a) complex lr(ppy)3 (1 ) and b) complex I r(piq)3 (2). c), d) Fluorescence microscope images of fibrils formed from insulin grinded with c) complex lr(ppy)3 (1 ) and d) complex lr(piq)3 (2).
[0049] Figure 3 shows examples of nano/micro-structures formed by self-assembly of a composite material obtained by grinding insulin with complexes lr(ppy)3 (1 ) or lr(piq)3 (2). The images are fluorescence microscope images of a), b) spherulites and c), d) aggregated amyloid-like fibrils. [0050] Figure 4 shows emission data from lr(ppy)3 (1 ) and I r(piq)3 (2) as free complexes in CHCI3 solution, and in samples exposed to fibrillation conditions. The top curve complex was exited at 380 nm; bottom curve complex was excited at 425 nm.
[0051 ] Figure 5 shows FT-Raman spectra recorded for powders of insulin, ground insulin, and the composite materials obtained by grinding insulin with metal-complex lr(ppy)3 (1 ) or lr(piq)3 (2).
[0052] Figure 6 shows Circular Dichroism spectra of solutions in 25 mM HCI incubated for 0 and 8 hours. Spectra are shown for insulin; ground insulin; and insulin ground with either of complex lr(ppy)3 (1 ) or I r(piq)3 (2).
[0053] Figure 7 shows MALDI-TOF spectra obtained on the composite material formed by grinding insulin with either of complex lr(ppy)3 (1 ) or I r(piq)3 (2).
Description of embodiments
[0054] The detailed description of the invention that follows will deal separately with the guest molecules, bio-polymers, and methods of preparation of the
composite materials by mixing the guest molecules and bio-polymers. The invention is finally exemplified with a number of experiments demonstrating the utility thereof.
Methods of preparation of composite materials, and self assembly of the composite materials
[0055] As already indicated, the present invention relates to methods of preparing functionalized nano/micro-structures based on self-assembly of a composite material consisting of functionalized bio-molecules.
[0056] In order to prepare the composite material, which is the precursor for self- assembly, the bio-molecule material (the self assembler) and the guest molecule material (the functionalizing agent) are grinded together in the solid state. If desired, additives such as solvent, or other materials, may be added to facilitate the grinding. Moreover, the materials may be pretreated with a variety of methods, such as melting the two components before grinding, or adding the guest molecule as a solution in organic solvent, and grinding the resulting material after, or during, the evaporation of the organic solvent. The grinding may be performed with any kind of milling equipment, including simply a mortar and pestle. The grinding process results in a water soluble composite material formed by the bio-molecule and the guest molecule. The grinding process may result in chemical changes, for example the formation or cleavage of covalent bonds, as long as the resulting composite material is still capable of undergoing self assembly processes. The resulting composite material is dissolved into an appropriate solvent, or mixture of solvents, and is then exposed to conditions that promote self-assembly of the bio- molecule. The resulting solution, dispersion, or precipitation of functionalized nano/micro-structures can be immobilized on a variety of solid supports, including, but not limited to silicon wafers, glass (e. g. glass slides, glass beads, glass wafers etc., silicon rubber, polystyrene, polyethylene, teflon, silica gel beads, gold, indium tin oxide (ITO coated materials, e. g. glass or plastics), filter paper (e. g. nylon, cellulose and nitrocellulose), standard copy paper or variants and separation media or other chromatographic media. Transfer of the functionalized micro-nanostructures to the solid support can be achieved by using, but not limited to, dip coating, spin- coating, microcontact printing, screen printing, ink jet technologies, spraying, dispensing and microfluidic printing by the use of soft lithography. Solvents for the functionalized micro-nanostructures of the present invention during the
immobilization to the solid support can be, but are not limited to, water, buffered water solutions, methanol, ethanol, chloroform and combinations thereof.
Supporting polymers of other kinds can also be added in this step. [0057] It will be apparent to those skilled in the art that various modifications and variations can be made to the structure of the present invention without departing from the scope or spirit of the invention. In view of the foregoing, it is intended that the present invention cover modifications and variations of this invention provided they fall within the scope of the following claims or their equivalents.
Examples
[0058] The iridium-complexes mentioned in the experimental descriptions below are tris[2-phenylpyridinato-C , ]iridium(lll), abbreviated as (lr(ppy)3) or (1 ), and tris[l -phenylisoquinoline-C , ]iridiurn(lll) abbreviated as (lr(piq)3) or (2) (see figure 1 for structural drawings of the complexes)
Example 1
Preparation of a functional ized nano/ micro-structure formed by self-assembly of a composite material formed by grinding of insulin and iridium-complexes.
[0059] In a typical procedure, 50mg bovine insulin was ground with 1 .Omg of Iridium-complex lr(ppy)3 (1 ) or I r(piq)3 (2) (see figure 1 for structures). The grinding was carried out in intervals for a total of 1 0 minutes. This resulted in a green or red composite material, which could be dissolved in 2M guanidine hydrochloride giving a green or red solution, which was dialyzed at +4°C against 3 rounds of 25mM HCI. After dialysis the solution was filtered through a 0.2μιτι PVDF filter. The solution was then diluted with 25mM HCI to final concentrations of 4.2-4.6mg/ml of insulin and 0.07-0.08mg/ml of complex lr(ppy)3 (1 ) or I r(piq)3 (2). The samples were then heated at 65°C for 40-72 hours in order to induce fibril formation.
Example 2
Attempt at extraction of the iridium-complex with chloroform, and comparison of the filtration behavior of solutions of the composite material before and after heating. [0060] 50mg bovine insulin was ground with 1 .Omg of the Iridium complex lr(piq)3 (2) as described in example 1 . A solution of the resulting composite material in 25mM HCI was then prepared as described in example 1 . The red colored solution was filtered through a 0.2μιτι PVDF filter. The resulting red solution was split in two parts. For one of the two solutions an attempt was made to extract the solution with chloroform. The color of the aqueous phase remained red, and the organic phase remained colorless. The other solution was heated at 65°C for 72 hours, after which an attempt was made to filter the solution through a 0.2μιτι PVDF filter. Upon filtration, the solution passing the filter became colorless, and all of the red colored material was retained in the filter.
Example 3
Fluorescence microscope characterization of nano/micro-structures formed by self- assembly of a composite material formed by grinding of insulin and iridium- complexes (Figure 2 and Figure 3).
[0061 ] The fluorescence microscope images of the self-assembled bovine insulin/lr-complex, prepared as in example 1 , were recorded with an
epifluorescence microscope (Zeiss Axiovert inverted microscope A200 Mot) equipped with a CCD camera (Axiocam HR) using a 40 x objective and a
405/30nm filter for the green emitting complex lr(ppy)3 (1 ), and 470/40nm filter for the red emitting complex I r(piq)3 (2), respectively. For the observation, the sample was prepared by drop casting the fibrillated bovine insulin/lr-complex onto Si/Si02 substrates.
Example 4
AFM characterization of nano/microstructures formed by self-assembly of a composite material formed by grinding of insulin and iridium-complexes (Figure 2). [0062] Atomic force microscopy (AFM) imaging was performed on samples of the self-assembled bovine insulin/lr-complex prepared as given in example 1 . Images were recorded by standard procedures in tapping mode on a SFM-Nanoscope III, Digital Instruments, with a J scanner. Cantilevers for tapping mode were obtained from NT-MDT. For the AFM observation, the amyloid-like fibrils were deposited onto Si/Si02 substrates by molecular combing : 1 Ομί of a drop of the fibrillated bovine insulin/lr-complex solution was deposited onto a Si/Si02 substrate, incubated for 1 minute and then gently blown off the substrate using a nitrogen gas flow.
Example 5
Characterization of luminescent properties of nano/micro-structures formed by self- assembly of a composite material formed by grinding of insulin and iridium- complexes (Figure 4).
[0063] The photoluminescence was measured in CHCI3 solutions for complexes lr(ppy)3 ( 1 ) or Ir (piq)3 (2), and in 25 mM HCI solutions of the self-assembled composite material, prepared as given in example 1 . The spectra were recorded with an ISA Jobin-Yvon Spex FluoroMax2 fluorimeter at excitation wavelength of 380nm and 425 nm for lr(ppy)3 ( 1 ) and lr(piq)3 (2), respectively.
Example 6
Characterization by Raman spectroscopy of the composite material formed by grinding of insulin and iridium complexes (Figure 5).
[0064] The composite material was prepared by grinding of insulin and iridium complexes lr(ppy)3 and lr(piq)3. The grinding was done as given in example 1 . The samples were recorded on the resulting powders of the materials, which were pressed into the cavity of aluminium disks. Samples were also prepared from just the metal complexes lr(ppy)3 and lr(piq)3, insulin, and ground insulin. Raman spectra were recorded using a Bruker FRA 1 06 Raman spectrometer (laser λ= 1 064nm), and the spectra from 1 024 scans were recorded at a power setting of ~500mW and a resolution of 4cm-1.
Example 7
Characterization by circular dichroism of the composite material formed by grinding of insulin and iridium complexes, and the nano/micro-structures formed by self assembly of the composite material (Figure 6).
[0065] A composite material was prepared by grinding insulin and iridium complexes. The resulting material was dissolved in 25mM HCI, and aliquots were withdrawn, and analyzed by CD. The rest of the sample was heated at 65°C/ and after 8 hours the samples were again examined by CD. Samples prepared from native insulin, and ground native insulin was analyzed in a similar way. The far- Ultraviolet (UV) circular dichroism (CD) spectra (~0.1 25mg/ml protein
concentration; diluted with Milli-Q water) were recorded using Chirascan CD Spectrometer and 2mm path length quartz cuvette.
Example 8
Characterization by MALDI-TOF MS of the composite material formed by grinding of insulin and iridium complexes (Figure 7).
[0066] AAALDI-TOF MS measurements were performed with a Voyager-DE instrument equipped with a 337nm nitrogen laser, a-cyano-4-hydroxycinnamic acid (CHCA) was used as matrix. A solution of the composite material (insulin and iridium-complexes) in 25mM HCI was prepared, and was then filtered through a 0.2μιτι PVDF filter. The sample was then diluted with water, and this solution was mixed 1 : 1 with a saturated solution of CHCA in 50:50 acetonitrile/water (0.1 % TFA). 0.5μί of the resulting solution was co-evaporated on a target plate, and the spectra were recorded in a positive reflector mode. MS data were acquired using the instrument default calibration, without employing internal or external calibration.

Claims

1 . A na no/micro-structure, consisting of a material formed by a self-assembly process of a water soluble composite material, consisting of a bio-molecular material and a hydrophobic material, where said self-assembly process occurs in aqueous solvent and said composite material is formed by grinding of a water soluble bio-molecular material capable of undergoing self-assembly, where said bio- molecular material consists of: i) proteins; or ii) nucleic acids; or iii) peptide chains, and said grinding is done in the presence of hydrophobic materials consisting of: i) hydrophobic molecules containing unsaturated or saturated hydrocarbon units (including fullerenes or other types of carbon structures); or ii) hydrophobic nano- particles; or iii) hydrophobic inorganic molecules.
2. The na no/micro-structure according to claim 1 , wherein said bio-molecule is insulin.
3. The na no/micro-structure according to claim 1 , wherein said hydrophobic material consists of dye molecules, whereby the self-assembly process results in nano/micro-structures capable of being luminescent.
4. The na no/micro-structure according to claim 3, wherein said dye molecule is lr(ppy)3 or I r(piq)3 or both.
5. The na no/micro-structure according to claim 1 or 2, wherein said hydrophobic material have a capacity of conducting electricity whereby the self- assembly process results in nano/micro-structures capable of conducting electricity.
6. The ηα no/micro-structure according to claim 1 or 2, wherein the hydrophobic material has magnetic properties, whereby the self-assembly process results in nano/micro-structures having magnetic properties.
7. The na no/micro-structure according to claim 1 or 2, wherein the hydrophobic material consists of drug molecules, whereby the self-assembly process results in nano/micro-structures comprising drug molecules.
8. The na no/micro-structure according to claim 1 or 2, wherein the employed hydrophobic materials is a combination of different hydrophobic materials, whereby the self-assembly process results in nano/micro-structures having a combination of two or more properties according to claims 2-7.
9. The na no/micro-structure according to claim 1 or 2, wherein said bio- molecule is a protein or oligopeptide or polypeptide, capable of undergoing self- assembly into amyloid-like structures, resulting in the formation of amyloid-like nano/micro-structures with properties according to claims 2-8.
1 0. A method for the formation of nano/micro structures comprising a bio- molecule and a hydrophobic material comprising the steps of: a) providing water soluble bio-molecules capable of undergoing self assembly processes consisting of i) proteins; or ii) nucleic acids; or iii) peptide chains b) providing hydrophobic material consisting of i) hydrophobic molecules containing unsaturated or saturated hydrocarbon units (including fullerenes or other types of carbon structures); or ii) hydrophobic nano-particles; or iii) hydrophobic inorganic molecules c) grinding the water soluble bio molecules according to a) in the presence of the hydrophobic material according to b) in the solid state thereby obtaining a water soluble composite material capable of undergoing self assembly processes in an aqueous solution into nano/micro structures having the hydrophobic material incorporated.
1 1 . The method according to claim 1 0, where the grinding is performed in the presence of additional additives such as solvents or salts, or the materials to be grinded are pre-treated by melting them together, or one or more of the components are added as a solution, and the grinding is commenced after or during
evaporation of said solvent.
1 2. The method according to claim 1 0 or 1 1 , where said bio-molecule is a protein or oligo-peptide or poly-peptide, capable of undergoing self-assembly into amyloid-like structures, resulting in the formation of amyloid-like nano/micro- structures with properties as claimed in claims 2-8.
1 3. The method according to claim 1 2, wherein said bio-molecule is insulin.
EP11707116A 2010-02-18 2011-02-17 Preparation of functionalized nano/micro-structures by self-assembly of biomolecules Withdrawn EP2536395A1 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
SE1000157 2010-02-18
PCT/EP2011/052368 WO2011101420A1 (en) 2010-02-18 2011-02-17 Preparation of functionalized nano/micro-structures by self-assembly of biomolecules

Publications (1)

Publication Number Publication Date
EP2536395A1 true EP2536395A1 (en) 2012-12-26

Family

ID=43875566

Family Applications (1)

Application Number Title Priority Date Filing Date
EP11707116A Withdrawn EP2536395A1 (en) 2010-02-18 2011-02-17 Preparation of functionalized nano/micro-structures by self-assembly of biomolecules

Country Status (2)

Country Link
EP (1) EP2536395A1 (en)
WO (1) WO2011101420A1 (en)

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
GB701814A (en) 1950-05-10 1954-01-06 Gen Aniline & Film Corp Water-dispersible dyestuff powders
EP1873869B1 (en) 2006-06-28 2013-07-10 Weidmüller Interface GmbH & Co. KG Electric module

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO2011101420A1 *

Also Published As

Publication number Publication date
WO2011101420A1 (en) 2011-08-25

Similar Documents

Publication Publication Date Title
Liu et al. Synthetic mimics of biotin/(strept) avidin
Lévy Peptide‐Capped Gold Nanoparticles: Towards Artificial Proteins
Gao et al. Quantum-dot nanocrystals for ultrasensitive biological labeling and multicolor optical encoding
Aili et al. Folding induced assembly of polypeptide decorated gold nanoparticles
Dorokhin et al. Reversible phase transfer of (CdSe/ZnS) quantum dots between organic and aqueous solutions
Chan et al. Nanocrystal biolabels with releasable fluorophores for immunoassays
US20070116627A1 (en) Carbon nanotube compositions and devices and methods of making thereof
Kainz et al. Combined covalent and noncovalent functionalization of nanomagnetic carbon surfaces with dendrimers and BODIPY fluorescent dye
US7507530B2 (en) Nanoparticle complexes having a defined number of ligands
González-Campo et al. Supramolecularly oriented immobilization of proteins using cucurbit [8] uril
Bhattacharya et al. Insights into the thermodynamics of polymer nanodot–human serum albumin association: a Spectroscopic and calorimetric approach
US7449445B2 (en) Conductive peptide nanofiber and method of manufacture of the same
CA2717719A1 (en) Nanoparticle conjugates
EP1534831A2 (en) Composition, method and use of bi-functional biomaterials
JP2007022873A (en) Water-dispersible protein/carbon nanotube composite and its production method and use
Pepe-Mooney et al. Characterization of Mesoscale Coiled-Coil Peptide–Porphyrin Complexes
Pepe-Mooney et al. Peptides as materials
Díaz et al. Sequence decoding of 1D to 2D self‐assembling cyclic peptides
JP2003329686A (en) Luminescent fine particle
Lim et al. Probing molecular mechanisms of M13 bacteriophage adhesion
Yin et al. Chameleonic dye adapts to various environments shining on macrocycles or peptide and polysaccharide aggregates
Zhao et al. Enhancing Optical Activities of Benzimidazole Derivatives through Coassembly for High-Efficiency Synthesis of Chiroptical Nanomaterials and Accurate ee% Detection of Natural Acids
Wagner et al. Switchable electrostatic interactions between gold nanoparticles and coiled coil peptides direct colloid assembly
Sun et al. Drug sensing protein crystals doped with luminescent lanthanide complexes
WO2011101420A1 (en) Preparation of functionalized nano/micro-structures by self-assembly of biomolecules

Legal Events

Date Code Title Description
PUAI Public reference made under article 153(3) epc to a published international application that has entered the european phase

Free format text: ORIGINAL CODE: 0009012

17P Request for examination filed

Effective date: 20120917

AK Designated contracting states

Kind code of ref document: A1

Designated state(s): AL AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HR HU IE IS IT LI LT LU LV MC MK MT NL NO PL PT RO RS SE SI SK SM TR

DAX Request for extension of the european patent (deleted)
STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: THE APPLICATION IS DEEMED TO BE WITHDRAWN

18D Application deemed to be withdrawn

Effective date: 20150829