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WO1999036357A1 - Matieres mesoporeuses et leurs procedes de fabrication - Google Patents

Matieres mesoporeuses et leurs procedes de fabrication Download PDF

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Publication number
WO1999036357A1
WO1999036357A1 PCT/US1999/001116 US9901116W WO9936357A1 WO 1999036357 A1 WO1999036357 A1 WO 1999036357A1 US 9901116 W US9901116 W US 9901116W WO 9936357 A1 WO9936357 A1 WO 9936357A1
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WIPO (PCT)
Prior art keywords
pore
mesoporous
pore forming
materials
sol
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PCT/US1999/001116
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English (en)
Inventor
Yen Wei
Danliang Jin
Tianzhong Ding
Jigeng Xu
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Drexel University
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Publication date
Application filed by Drexel University filed Critical Drexel University
Priority to AU24600/99A priority Critical patent/AU2460099A/en
Publication of WO1999036357A1 publication Critical patent/WO1999036357A1/fr
Priority to US09/598,717 priority patent/US6696258B1/en

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    • 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
    • C—CHEMISTRY; METALLURGY
    • C01—INORGANIC CHEMISTRY
    • C01B—NON-METALLIC ELEMENTS; COMPOUNDS THEREOF; METALLOIDS OR COMPOUNDS THEREOF NOT COVERED BY SUBCLASS C01C
    • C01B33/00—Silicon; Compounds thereof
    • C01B33/113—Silicon oxides; Hydrates thereof
    • C01B33/12—Silica; Hydrates thereof, e.g. lepidoic silicic acid

Definitions

  • Surfactants ionic and neutral have been the most commonly used pore forming materials for directing the formation of mesoporosity.
  • the ionic pathways are based on charge matching between the ionic surfactants and ionic inorganic precursors through electrostatic interaction.
  • Neutral surfactants are theorized to use hydrogen bonding between the surfactants and the precursors to direct formation of mesostructures.
  • mesoporous materials may also be prepared from interlay er crosslinking of a layered silicate through ion exchange reactions with organic cations.
  • MCM-41 mesoporous materials have an array of hexagonal arrangements of uniform mesopores of 15 to 100 A in diameter, which could be controlled by the hydrophobic alkylchain length of ionic surfactants or with the aid of auxiliary organic compounds as spacers.
  • the ionic templates are usually removed by high temperature calcination or ion exchange.
  • a family of hexagonal mesoporous silicas have been prepared.
  • the pore size may be adjusted by changing the hydrophobic tail length of the amines.
  • the template can be removed by solvent extraction.
  • the mesoporous materials have greater wall thicknesses (1.7-3.0 nm) due to the absence of electrostatic or charge-matching effects, and thus higher thermal stability than M41S materials.
  • the materials exhibit both complementary textural and framework-confined mesoporosity. The toxicity of amines also remains a concern if a large scale production is intended.
  • microporous and mesoporous materials are for the immobilization of biologically active agents within these materials. Immobilization of enzymes, in particular, has been a subject of extensive research efforts because of its immense technological potentials. Among the popular methods of immobilization is formation of chemical bonding between enzymes and a solid support, which often alters the enzymatic activity.
  • a variety of enzymes and other bioactive substances have been entrapped in inorganic oxides such as silica for biocatalysis and biosensor applications through conventional sol-gel processes.
  • silica matrices i.e. typical pore diameter ⁇ 15 A
  • Mesoporous materials are valuable to the life sciences because the larger pore size in comparison with microporous materials allows for a more suitable environment and better mass transfer for biologically active agents.
  • Much of the prior art involving the immobilization of biologically active agents in porous materials involves use of microporous materials, not mesoporous materials.
  • Biologically active agents previously have been bound to microporous materials, but the pore diameters result in steric hindrance and mass transfer limitations on the use of such materials in biological reactions.
  • Immobilization of enzymes, and other biologically active agents, by entrapment in a gel matrix is based on the occlusion of an enzyme within a constraining structure tight enough to prevent the relatively large protein molecules from diffusing into the surrounding media, while still allowing penetration of the relatively small substrate and product molecules in and out of the matrix. Due to the advantages in their generality, the methods which are widely used for entrapping biologically active agents, include adsorption on an inert support, encapsulation within a semipermeable membrane, covalent crosslinking of the protein molecules or coupling to a support.
  • the conventional sol-gel process for the formation of ceramic or glass materials consists of hydrolysis of a metal alkoxide precursor, typically tetramethylorthosilicate or tetraethylorthosilicate for forming silica, in the presence of an acid or, less often, a base catalyst, followed by the polycondensation of the inorganic intermediates and evaporation of solvents, giving a porous solid gel.
  • a metal alkoxide precursor typically tetramethylorthosilicate or tetraethylorthosilicate for forming silica
  • an acid or, less often, a base catalyst followed by the polycondensation of the inorganic intermediates and evaporation of solvents, giving a porous solid gel.
  • This type of air-dried xerogel typically possesses numerous pores or channels well below 15 A in diameter, depending on the synthesis conditions.
  • Such materials are often used as the host matrices for the encapsulation of various types of chemicals,
  • Mesoporous sol-gel materials may be an alternative host for the development of improved biogels.
  • mesoporous silicate and aluminosilicate, as well as other metal oxides with pore diameters in the range 20 to 100 A, and up to 300 A have been synthesized based on the template-directed hydrothermal reactions according to the varied ionic or nonionic surfactant templating pathways.
  • the template-based synthesis approaches to the mesostructured materials are difficult to adapt for direct immobilization of biologically active agents due to the severe reaction conditions necessary.
  • the substrate molecules will never have a chance to encounter the enzyme molecules entrapped in the core of a relatively large particulate because they have already been consumed and converted into the products by the enzyme molecules in the outer shell before they can reach the particle center. This will yield a lowered apparent activity when the calculation is based on the total amount of enzyme entrapped. It is known that volume diffusion proceeds in large pores exceeding 100 nm while Knudsen diffusion proceeds in narrow pores below 100 nm in which the mean free way of the molecules exceeds the pore diameter, and sol-gel materials having pore diameters in the range of micropores ( ⁇ 20 A) or mesopores (20-500 A) fall into this category. It has been expected and pointed out that diffusion limited reaction rates have been evident in microporous sol-gel materials and virtually an obstacle in the practical applications.
  • a mesoporous material for use in applications with biologically active agents that can be prepared using a pore forming material that is readily available, non-toxic and inexpensive, and is free from the undesirable effects upon biologically active agents such as denaturation or toxicity.
  • a simple, controllable synthesis method of such a mesoporous material and for immobilizing a biologically active agent within such a mesoporous material in which the biologically active agent can retain an acceptable or high degree of its native activity.
  • the invention includes a method for making a mesoporous material which comprises forming an aqueous solution having an organometallic compound, adding a solution comprising a pore forming material to form a sol-gel matrix by polycondensation, drying the sol gel matrix and removing the pore forming material from the dried sol-gel matrix to thereby form a mesoporous material.
  • the pore forming material is selected from the group consisting of monomeric polyols, polyacids, polyamines, carbohydrates, oligopeptides, oligonucleic acids, carbonyl functional organic compounds, and mixtures and derivatives thereof.
  • the pore forming material is a nonsurfactant, polar compound capable of forming hydrogen bonding.
  • a biologically active agent is added after adding the solution of the pore forming material.
  • the invention further includes a mesoporous material having pores and formed from a sol-gel matrix comprising a pore forming material and having a surface
  • the invention further includes a method of using a mesoporous material with a biologically active agent which comprises preparing a mesoporous material having pores from a sol-gel matrix which comprises an organometallic compound and a pore forming material selected from the group consisting of monomeric polyols, polyacids, polyamines, carbohydrates, oligopeptides, oligonucleic acids, carbonyl functional compounds, and mixtures and derivatives thereof, immobilizing a biologically active agent within the pores of the mesoporous material and introducing the immobilized active agent into a biological system.
  • the invention also includes a mesoporous material comprising pores having an average pore diameter of from about 30 A to about 60 A, wherein a plurality of the pores are interconnected within the mesoporous material and a biologically active agent immobilized within the pores.
  • Fig. 1 is a graphical representation of a nitrogen adsorption-desorption isotherm at -196° C after water extraction for the porous silica samples formed in
  • Example 1 having acid-catalyzed samples
  • Fig. 2 is a graphical representation of a nitrogen adsorption-desorption isotherm at -196° C after water extraction for the porous silica samples formed in
  • Example 2 having base catalyzed samples
  • Fig. 3 is a graphical representation of the relationship between the net surface area and glucose concentration after water-extraction for both acid and base catalyzed samples of Example 1 (O) and Example 2 (D);
  • Fig. 4 is a graphical representation of the relationship between net pore volume and concentration of glucose after water extraction for the acid and base catalyzed samples of Example 1 (O ) and Example 2 (O);
  • Fig. 5 is a graphical representation of BJH pore size distribution derived from nitrogen desorption isotherms which plots differential volume as a function of pore size for the silica matrices formed in Example 1 ;
  • Fig. 6 is a graphical representation of BJH pore size distribution derived from nitrogen desorption isotherms which plots differential volume as a function of pore size for the silica matrices in Example 2;
  • Fig. 7 is a graphical representation of the nitrogen adsorption-desorption isotherms at -196° C for samples after water extraction in Example 3
  • Fig. 8 is a graphical representation of the BJH pore size distributions derived from nitrogen adsorption branches at -196° C for samples after removal of D- glucose with water extraction as in Example 3;
  • Fig. 9 is a graphical representation of the BJH pore size distributions derived from nitrogen desorption branches for samples after removal of D-glucose with water extraction as in Example 3;
  • Fig. 10a is a graphical representation of the relationship between matrix pore diameter and relative enzymatic activity of immobilized ACP as in Example 3;
  • Fig. 10b is a graphical representation of the relationship between matrix pore volume and relative enzymatic activity of immobilized ACP as in Example 3
  • Fig. 1 la is a graphical representation of the relationship between enzymatic activity of free ACP and the concentration of pNPP at pH 6.2 and room temperature as in Example 3;
  • Fig. 1 lb is a graphical representation of the Lineweaver-Burk plot of free ACP at pH 6.2 and room temperature as in Example 3;
  • Fig. 1 lc is a graphical representation of the Eadie-Hofstee plot of free
  • FIG. 1 Id is a graphical representation of the relationship between enzymatic activity of immobilized ACP at various D-glucose concentrations and the concentration of pNPP as in Example 3;
  • Fig. 1 le is a graphical representation of the Eadie-Hofstee plots of immobilized ACP at various D-glucose concentrations assayed at pH 6.2 and room temperature as in Example 3;
  • Fig. 12 is a graphical representation of the nitrogen adsorption- desorption isotherms determined at various relative pressures (P P 0 ) on the mesoporous material samples in Example 5 including an inset which is a graphical representation of the derived BJH desorption pore size distributions;
  • Fig. 13 is a representation of the XRD patterns of the mesoporous materials prepared with 48% of D-maltose in Example 5;
  • Fig. 14 is a representation of TEM micrograph for sample DB50X prepared with 50wt% dibenzoyl-L-tartaric acid in Example 5.
  • an "organometallic compound” means a compound including, at least, a metal element and an organic moiety as described further below.
  • a “sol-gel reaction” includes any reaction for forming a sol-gel matrix, such as processes including hydrolysis of an organometallic precursor in the presence of catalyst and solvent, which may be an aqueous solvent or mixture of organic solvent in aqueous solvent and polycondensation of the inorganic intermediates formed by these reactants to create a porous sol-gel matrix.
  • catalyst and solvent which may be an aqueous solvent or mixture of organic solvent in aqueous solvent and polycondensation of the inorganic intermediates formed by these reactants to create a porous sol-gel matrix.
  • meopore means a pore having a diameter of from about 20 A to about 500 A
  • macropore means a pore having a diameter greater than about 500 A
  • micropore means a pore with a diameter less than 20 A.
  • a “mesoporous material” formed in accordance with the invention may include a material which has substantially all mesopores, a material having a mixture of mesopores and micropores or a material having a mixture of mesopores and macropores.
  • the mesoporous materials in accordance with the invention have substantially all pores as mesopores, however mixtures of mesopores with other pores sizes may be formed in accordance with the invention by controlling the amount of pore forming material described below.
  • the invention includes a versatile, efficient, and preferably biocompatible, non-surfactant method for making mesoporous materials and mesoporous materials having advantageous properties.
  • Non-surfactant, preferably polar, pore forming materials are used in the method such that problems associated with use of surfactants are avoided.
  • pore forming materials are used in the method to provide excellent control over the desired pore diameter and contribute to large surface areas, large pore volumes and narrow pore size distributions in the mesopore range within the mesoporous materials formed by the method.
  • the pore diameter in the mesoporous material can be controlled by varying the concentration of the pore forming material.
  • Such a feature provides flexibility in producing mesoporous materials as well as reproducibility in finished products.
  • the method does not introduce compounds into the mesoporous structure which affect pore size development and/or which could be toxic to biologically active agents which can be immobilized within the pores of the mesoporous materials.
  • the invention further includes a method of using a mesoporous material by immobilizing a biologically active agent within a mesoporous material and introducing it into a biological system.
  • the mesoporous material having a biologically active agent immobilized within the pores allows the biologically active agent to retain from about twice to about 10 times the activity possible using a microporous sol-gel material formed in the absence of a pore forming material for immobilization of a biologically active agent, and in some cases activities of one to three orders of magnitude greater have been observed.
  • the mesoporous material is one in which substantially all of the pores are mesopores.
  • the invention includes a method for making a mesoporous material which includes, as a first step, forming an aqueous solution of an organometallic compound. This step is directed to initiating formation of a sol-gel matrix.
  • the sol-gel matrix is formed by providing an aqueous solution of the organometallic compound and hydrolyzing it in the presence of a catalyst and preferably a solvent. The hydrolysis is preferably acid or base catalyzed and generates intermediates of the organometallic compound in solution which then undergo polycondensation to form a network or sol-gel matrix.
  • the sol-gel matrix is a composite of organic and inorganic material. Such chemistry is known as sol-gel chemistry and is fully described in Y. Wei et al., Phys. Chem.. vol. 101, p. 3318 (1997), which are herein incorporated in full by reference.
  • the aqueous solution including an organometallic compound preferably includes an organometallic compound such as an metal alkoxide compound, preferably an alkoxy silane, or a mixture of such compounds.
  • organometallic compound(s) may be used as the only precursor(s) or used in combination with other sol-gel forming compounds or polymers, including homopolymers and copolymers of styrene, vinyl, acrylic, alkyl acrylate, and the like.
  • the organometallic compound has the following formula:
  • M is preferably a metal capable of forming a sol-gel matrix in aqueous
  • R is a branched, straight chain or cyclic alkyl, alkenyl, alkynyl group of from one to 20 carbon atoms or an aromatic group of from six to 20 carbons.
  • R 1 is R 2 , a halogen, or a polymeric moiety such as homopolymers and copolymers of styrene, acrylics, alkyl acrylates, vinyls, and other similar sol-gel forming polymers known in the art or to be developed.
  • R and R may be unsubstituted or further substituted with functional groups which will not interfere with formation of the sol-gel matrix and, preferably, which would not render the matrix toxic or otherwise form too strong a bond with the selected pore forming material.
  • the metal, M is preferably one of the following metals such as silicon, aluminum, titanium, vanadium, boron, magnesium, iron, niobium and other metals which are useful for forming a sol-gel matrix. More preferably, the metal is silicon, titanium or aluminum, with silicon being most preferred due to the inert nature of these metals with respect to biologically active agents.
  • the valence x + y of the metal M is accordingly preferably 2-6.
  • the organometallic compound may be used to form a mixed sol-gel material, for example, an alkoxysilane and a aluminum alkoxide may be combined to form a mixed matrix of silica and alumina.
  • the organometallic compound is an alkoxysilane, such as a tetraalkylorthosilicate, and is hydrolyzed.
  • a tetraalkylorthosilicate such as tetraethylorthosilicate (TEOS) or tetramethylorthosilicate (TMOS) is used.
  • the hydrolysis reaction is carried out in an aqueous solution using the above-described organometallic compound, a solvent and a catalyst. If available, commercial solutions of these materials may be used, provided they have acceptable weight percentage ratios, as described below, and satisfy the other criteria as describe herein.
  • the catalyst can be any catalyst capable of initiating a sol-gel hydrolysis/polycondensation reaction, preferably an acid or a base catalyst.
  • Preferred acid catalysts include hydrochloric acid, nitric acid, sulfuric acid, methanesulfonic acid, tolylsulfonic acid, quaternary amines, phosphoric acid, polystyrenesulfonic acid, polymethyacrylic acid, and photoacids which are neutral compounds in the dark and become acids upon exposure to light or radiation.
  • the acid catalyst is hydrochloric acid.
  • Preferred base catalysts include sodium hydroxide, ammonium hydroxide, as well as other Group IA and Group IIA metal hydroxides or alkoxides or salts, for example, carbonate, halide, phosphorate, and acetate salts, triethylamine, and photobases which are neutral in the dark and become basic upon exposure to light or radiation.
  • Suitable solvents are those which are compatible with the sol-gel reactions and which are miscible with or have reasonably good solubility in the aqueous solution.
  • solvents include ethers such as tetrahydrofuran, alcohols such as methanol, ethanol, propanol, and butanol, dimethylformamide, dimethylsulfoxide and the like.
  • ethers such as tetrahydrofuran
  • alcohols such as methanol, ethanol, propanol, and butanol
  • dimethylformamide dimethylsulfoxide and the like.
  • a neutralizing agent be added after the hydrolysis, such as ammonia and sodium hydroxide or hydrochloric acid or a similar buffering or neutralizing agent, to adjust the pH of the aqueous solution to a range of from about 2 to about 9, and preferably from about 5 to about 8 before adding the biologically active agent.
  • pH is adjusted to the value that is appropriate for optimal stability and activity of the biologically active agent chosen to be immobilized in the sol-gel matrix.
  • the rate of polycondensation reaction is known to be high at a neutral pH of about 7. If pH is already acceptable, the neutralization is not necessary.
  • the aqueous solution of the organometallic compound includes the organometallic compound, water, catalyst and solvent.
  • organometallic precursor on the basis of one mole of organometallic precursor used, about 0.1 to more than 10 moles, preferably about 1 to 3 moles of water are used, about 0.00001 to 0.5 moles, preferably from about 0.001 to 0.01 moles of catalyst are used, and from about 0.1 or less to about
  • the solution is preferably initially mixed and undergoes a pre-hydrolysis reaction in which the mixture becomes homogeneous, but is still liquid.
  • the solution is preferably heated to a temperature from room temperature to the reflux temperature of the solvent, preferably from about 25 to about 100°C or higher, and most preferably to the refluxing temperature of the solvent, i.e., the boiling temperture of the solvent, preferably under an inert atmosphere such as nitrogen gas.
  • the hydrolysis temperature may be varied depending on the type of sol-gel to be formed, the catalyst used and the desired pH, among other factors and may be optimized within the ordinary skill in the art to achieve varied results for different applications.
  • the material is heated for a period of time sufficient to achieve a high degree of hydrolysis and partial condensation but without gelation, i.e., the losing of fluidity.
  • the refluxing is to facilitate the hydrolysis reaction. Typically, about 1-2 hours is sufficient for this purpose.
  • the solution is preferably then cooled and combined with a solution of one or more pore forming materials.
  • the solution may be cooled by any conventional means, preferably by sitting.
  • the pH can be adjusted at this point as noted above to a desired level, particularly if a biologically active agent is to be immobilized in the mesoporous material.
  • Other auxiliary agents as well known in the art may also be added.
  • the solution including the pore forming material (also known in the art as a "templating" material) is preferably added after initial pre-hydrolysis as described above.
  • the pore forming material can be a single pore forming material in accordance with the invention or a mixture of such pore forming materials.
  • the pore-forming material which preferably interacts by polar or hydrogen bonding or other weak bonding interactions with intermediates of the organometallic compound in the pre- hydrolyzed solution is inco ⁇ orated in the sol-gel matrix formed when the pre- hydrolyzed solution undergoes polycondensation. Polycondensation may occur under acid or base catalyzed or neutral conditions.
  • the sol-gel matrix formed thereby is then preferably dried by a suitable method to form a solid matrix, which is a solidified matrix formed from the gel. Drying may be accomplished by evaporation, vacuum drying, heating, oven drying, desiccation or other suitable methods, preferably by controlled evaporation over a period of time to allow for a more uniform gelation.
  • the pore forming material-containing solution is sealed in a container with an impermeable cover such as a paraffin film, having small holes to allow the evaporation of solvents and byproducts of the sol-gel reactions or with a semi-permeable membrane allowing solvent evaporation.
  • a monolithic disc typically a transparent, monolithic disc of pore-forming-containing material is obtained.
  • the disc shape is formed by allowing the material to dry in a cylindrical container or mold, however, the shape of the material is not essential to the method as described herein. Evaporation may occur over a period of days, preferably from about 1 to about 10 days, more preferably from about 1 to about 7 days, and most preferably from about 1 to about 3 days.
  • the pore forming material is preferably a biocompatible, non-surfactant material which is also compatible with a biologically active agent if such an agent is to be immobilized in the mesoporous material.
  • the pore forming material is a non-surfactant, polar solid or high boiling liquid. Most preferably, it is capable of forming hydrogen bonding.
  • Such compounds include, for example, monomeric polyols, polyacids, polyamines, carbohydrates, oligopeptides, oligonucleic acids, and mixtures and derivatives thereof.
  • "Monomeric polyols” are aliphatic or aromatic polyols of from about 3 to about 20 carbon atoms such as glycerol, ethylene glycol, diethylene glycol, dihydroxyethyl ether, and bisphenol A.
  • Polyacids may be aromatic or aliphatic and include tartaric acid, EDTA, EGTA, poly- or oligo- styrenesulfonic acid or methacrylic acid, camphorsulfonic acid, oligophosphoric acids and their derivatives.
  • Polyamines may be aromatic or aliphatic and include diaminoethylamine, triaminoethylamine, diaminoaryl ether, and derivatives thereof.
  • Carbohydrates which may be used include sugars such as glucose, maltose, fructose, sucrose, lactose, talose, galactose and the like as well as other di-, tri- and oligo- saccharides and complex carbohydrates such as starches.
  • Oligopeptides which may be used include amino acids, dimers, trimers and higher oligomers of peptides.
  • Oligonucleic acids may include AMP, ADP, ATP and other oligomers of nucleotides and their derivatives.
  • pore forming materials Other organic compounds that include carbonyl groups capable of forming hydrogen bonds such as ketones, aldehydes and esters may also be used as the pore forming materials.
  • Derivatives of the above-mentioned pore forming materials include multifunctional compounds with mixed alcohol, amine, acid and carbonyl groups which may be used as well.
  • the pore forming material may be used alone or in combinations of two or more pore forming compounds.
  • the preferred pore forming materials include D-glucose, D-maltose, D- fructose, sucrose, dibenzoyl-L-tartaric acid, cyclodextrins, and soluble starch.
  • the pore forming material may be added alone or in an aqueous or other solution miscible with the pre-hydrolyzed solution formed as described above.
  • the amount of the pore forming material is preferably from about 20 to about 70% by weight based on the weight of the final, dried pore-forming material-containing solid matrix as determined by theoretical calculation based on the reactants selected for forming the sol-gel matrix in accordance with the ordinary skill in the art.
  • the amount of pore forming material provided can be varied to control the size and number of mesopores within the resulting matrix.
  • the pore forming material may be added at any time prior to the gelling or drying of the sol gel matrix, but it is preferably added after forming the pre-hydrolyzed solution as noted above, followed by neutralizing of the hydrolyzed solution when a biologically active agent is provided in order to minimize unwanted side reactions.
  • An aqueous solution is preferred, because the polarity of the solution solubilizes the pore forming material. While water is preferred, less polar solvents can be used for solubilizing pore forming materials having low polarity provided such solvents are miscible with the pre-hydrolyzed solution.
  • Examples of preferred solutions of pore forming materials include aqueous solutions of D-glucose (0.8 mol/L) and D-maltose monohydrate (1.2 mol/L) and an ethanol solution of dibenzoyl-L-tartaric acid (DBTA) (0.3mol/L) preferably formed while stirring.
  • DBTA dibenzoyl-L-tartaric acid
  • the water used be distilled and/or deionized water to avoid impurities. It is further preferred that the solution be stirred while adding the pore forming material.
  • the step of adding a pore forming material prior to the gelling or drying and formation of a sol gel matrix both enhances the speed of the overall method of making a mesoporous material and provides the very valuable means for controlling the nature of the pores, i.e., the degree of mesoporosity, and the average pore diameter of the resulting material to narrow ranges such as from about 30 A to about 60 A.
  • the concentration of pore forming material added the pore size of the resulting pores can be controlled to achieve pores which are substantially all mesopores, which are a mixture of mesopores and micropores, or which are a mixture of mesopores and macropores for different matrix applications.
  • the method as described above, using the preferred pore forming materials provides a faster route to mesoporous materials than previously obtained in the art by methods which do not use pore forming materials or "templates" and provides a biocompatible, non-toxic mesoporous material which may be used with biologically active agents.
  • the speed of the drying step alone is carried out in as little as from about 1 to about 3 days.
  • the method further provides a simple, reliable and reproducible procedure for exerting substantially tight control over pore size diameter and degree of mesoporosity in the resulting mesoporous material, and also provides better control of pore diameter range in pore size distribution.
  • the pore forming material may be removed from the solid matrix by any suitable purification method which would not have a detrimental effect on the matrix, but which will allow for separation of the particular pore forming material used.
  • Preferred purification methods involve procedures known to those skilled in the art or to be developed, and include, but are not limited to calcination and solvent extraction. Most preferably, solvent extraction is used.
  • solvent extraction is used.
  • the matrix first undergoes grinding to form a particulate form of the matrix, most preferably a powder.
  • Preferred extraction can be undertaken with compatible, preferably polar and low boiling solvents including lower molecular weight alkanols, such as methanol, water, saline and buffered solutions .
  • the solvent can be varied to achieve good removal of the pore forming material depending on the type of pore forming material used.
  • the solvents, mixtures of the solvents and extraction conditions are selected so as not to be capable of denaturing biologically active materials, when such materials are to be mobilized within the mesoporous material.
  • Multiple extractions can be carried out by a method such as mixing the ground solid matrix with solvent, separating by centrifugation and decanting to substantially remove the pore forming material. Other extraction or purification techniques may be employed.
  • this step provides a simple and efficient means for effectively removing the pore forming material.
  • the pore forming material is effectively removed to result in a mesoporous material.
  • the degree of removal of pore forming material can readily be monitored by conventional analytical methods such as infrared spectroscopy or thermogravimetric analysis.
  • the invention includes mesoporous materials, which may be either substantially mesoporous, or materials having mixtures of mesopores and micropores, or mesopores and macropores.
  • mesopores can be controlled so as to
  • the mesoporous materials have substantially all mesopores, i.e., they have at least about 50 % mesopores.
  • the materials are formed using sol-gel techniques, such as those described herein, however, the sol-gel technique should not be considered limited to those described herein, provided that the technique used includes the preferred pore forming materials described above to form the mesopores.
  • the invention also includes a method for making a mesoporous material and for using a mesoporous material in which a biologically active agent is immobilized within the mesoporous material.
  • the mesoporous material may be any mesoporous structure formed in accordance with this invention or as described herein provided the mesoporous material is formed using an organometallic compound, such as those described above, and a pore forming material as set forth herein, more preferably, using a monomeric polyol, carbohydrate, oligopeptide, oligonucleic acid, a carbonyl functional compound and mixtures and derivatives thereof.
  • the mesoporous material is preferably formed by the above preferred sol-gel technique, or by any known sol-gel technique or technique to be developed provided the pore forming material is a preferred pore forming material as set forth herein, i.e., a monomeric polyol, a polyacid, a polyamine, a carbohydrate, an oligopeptide, an oligonucleic acid, and mixtures and derivatives of these materials.
  • the biologically active agent to be immobilized within the mesoporous material may be any type of cell, a portion of a cell, a microorganism, a virus, a nucleic acid, an enzyme, a polysaccharide, a polypeptide, a subunit of a polypeptide, a drug, a therapeutic agent, a diagnostic agent and or mixtures or derivatives of these compounds, so long as the agent has biological activity or activity in a biological system.
  • the biologically active agent may be immobilized, i.e., entrapped within the pores, by providing the biologically active agent to the sol-gel matrix while the matrix is being formed.
  • the biologically active agent can be provided to the mesoporous pores after formation of the matrix and removal of the pore forming material. It is preferred, however, that the biologically active agent is added during formation of the pore forming material, and more preferably, after adding the pore forming material. If an acid-catalyzed method is used, it is preferred that a neutralizing step be undertaken as described above, before introducing the biologically active agent.
  • pores will be formed which will entrap the biologically active agent in the mesoporous material, but will allow sufficient porosity for movement within the pore structure to allow material to diffuse inward and outward thereby providing excellent mass transfer properties and good biological activity from the biologically active agent because the mesoporous material is non-toxic.
  • the method allows for adjustment of pore size to control activity or to adjust pore size for different sized biologically active agents. As such, the method is valuable because it results in an immobilized, but still highly active, biological agent entrapped within a mesoporous material which can be used in a wide variety of applications including diagnostic, therapeutic and catalytic applications.
  • the great value of this method is that it results in a material that overcomes the primary problems and limitations of immobilizing biologically active materials.
  • the method and resulting materials minimize restriction of access to substrates or other molecules necessary for biological activity caused by steric hindrance due to pore diameters being too small, because pore size may be adjusted to overcome this difficulty. Further pore size can be adjusted for maximized mass transfer properties.
  • leaching and escape of the immobilized agent due to a too large pore diameter can also be avoided.
  • mesoporous materials and the pore forming materials used in the invention are non- toxic, problems associated with toxicity, denaturation or loss of biological activity due to contact with templating or other pore forming materials which are not biocompatible are further obviated.
  • mesoporous materials with immobilized biologically active agents in accordance with the invention are stable and may not cause denaturation or degradation when exposed to or contacted with physiological elements present in biological systems such as protease, antibodies, and immune system responses or caused by poor thermal stability.
  • the mesoporous materials overcome significant obstacles in the prior art for adopting mesoporous materials for use with biologically active agents.
  • the biologically active agent is an enzyme such as acid phosphatase, alkaline phosphatase or horseradish peroxidase.
  • the biologically active agent is added after adding the solution containing the pore forming material but simultaneous addition or addition prior to the pore forming material is also acceptable, provided the biologically active agent retains its activity.
  • the biologically active agent will usually have to be added after the neutralization step to prevent denaturation and loss of activity.
  • the enzyme can be added as a stock solution and concentrations of agent in solution can be varied using techniques known to those of ordinary skill in the art depending on the biologically active agent chosen, its expense, availability, the targeted application of use for the biologically active agent and the like.
  • This method results in the formation of a mesoporous material with an immobilized biologically active material which substantially or completely retains its natural activity.
  • This immobilized agent in this mesoporous material is now ideally suited for catalytic, diagnostic and/or therapeutic applications in biological systems.
  • This invention thus provides a way to obtain biological activities from about twice to about 10 times, and some cases from 1 to 3 orders of magnitude greater than the activity possible using a microporous sol-gel material formed in the absence of a pore forming material for immobilization of a biologically active agent. Since there are no toxic surfactants or other denaturing chemicals used, the native structure of the biologically active material is maintained, and the materials may be formed easily, reproducibly and controUably to form preferred mesoporous structures for various sized biologically active agents.
  • the preferred pore forming materials result in the property of the pore forming material being readily removable from the matrix.
  • the preferred compounds are readily soluble in aqueous buffers and more importantly, their biocompatibility, in contrast to surfactants which are often denaturants or toxic to biological materials, enables the simultaneous immobilization of biologically active agents in the presence of the pore forming materials.
  • the method further involves contacting the immobilized active agent to a biological system, such as in a human, an animal or an in vitro test specimen.
  • a biological system such as in a human, an animal or an in vitro test specimen.
  • the immobilized biologically active agent in the mesoporous material can be used to function as a biosensor, biocatalyst, therapeutic agent and/or a diagnostic agent as well as in any other application in which improved mass transfer, thermal stability and improved biologically active agent activity would be beneficial.
  • Tetraethylorthosilicate was purchased from Aldrich, Milwaukee, Wisconsin. Ethyl alcohol was supplied by Pharmco Products of Brookfield, Connecticut. D-glucose, sodium hydroxide and hydrochloric acid were obtained from Fisher Scientific, Fair Lawn, New Jersey. All chemicals and reagents were used as received without further purification. The preparation of mesoporous silica-based materials were undertaken as described below.
  • sol-gel reactions were carried out to form mesoporous silica- based materials by using varied D-glucose concentrations.
  • the sol-gel reactants included 0.15 mol of tetraethylorthosilicate, 0.46 mol ethanol, 0.375 mol water and 1.5 mmol (2.0 M) hydrochloric acid as an acid catalyst. These components were mixed in a flask at room temperature under agitation. After about 15 minutes, the mixture became homogeneous accompanied by a temperature increase. The solution was heated to reflux under nitrogen atmosphere for 2 hours and cooled to room temperature to form a prehydrolyzed sol.
  • the prehydrolyzed sol formed a stock solution useful for the subsequent synthesis with varied D-glucose levels.
  • the sol in an amount of 4.0 g, was neutralized with 0.32 ml of 0.25M NaOH (aqueous) to a pH of 6.0. After neutralizing, the sol was combined with a solution of 0.5 g D-glucose in 0.5 g distilled water under stirring. The amount of sol used varied with the desired weight percentage D-glucose.
  • the glucose-containing homogeneous sols formed were sealed in a cylindrical glass container using a paraffin film. From 10-12 small holes were punched with a syringe needle through the film to allow evaporation of volatile molecules of solvents and byproducts of the sol-gel reactions. Upon gelation and drying at room temperature for periods of time of from 3 to 7 days, a transparent and monolithic disc of glucose-containing silica sample was obtained.
  • the as-synthesized samples (which were from 0.3 to 0.9 g, depending on the glucose content) were ground into a fine powder, and immersed in 15 mL distilled and deionized water under agitation for 15 min. After centrifugation and decantation, the samples were placed in another 15 mL of water for 3 hours under agitation. The sample mixtures were centrifuged and the samples were soaked in 15 mL water overnight. In the following two days, the samples were washed twice a day in the same manner. After such an extraction, with a total of about 100 mL of water in seven portions, the samples were dried in an oven at 115°C overnight. Both infrared and thermogravimetric analysis measurements showed the removal of glucose was complete.
  • the D-glucose concentration in the sol-gel materials were determined from the weight loss at 750°C using thermogravimetric analysis (TGA) on a DuPont 2000 Thermal Analyzer equipped with a TGA 950 Module.
  • TGA thermogravimetric analysis
  • the samples were first crushed with a mortar and pestle into a fine powder of 100-500 microns particle size. Upon drying at 115 °C for 2 hours in an oven prior to TGA.
  • the powdered samples were loaded to the TGA sample container and heated from ambient to 800 °C at a heating rate of 10 °C/min under oxygen atmosphere.
  • the infrared spectra of the samples were measured in the form of KBr powder-pressed pellets on a Perkin Elmer 1600 FT-IR spectrophotometer (Norwalk, Connecticut) under ambient conditions. Both as-synthesized and water-extracted powder samples were used for the spectral measurements.
  • the nitrogen-sorption characterization of the powdered samples before and after removing D-glucose by extraction were conducted on a Micromeritics ASAP 2010 Surface Area Pore-Size Analyzer (available from Micromeritics of Norcross, Georgia) at -196°C (liquid nitrogen). The samples were degassed at 200 °C and 10 torr overnight prior to nitrogen-desorption measurement. The surface and pore parameters were calculated using software from Micromeritics, Inc.
  • BET Brunauer-Emmett-Teller
  • the pore diameter using the Barrett- Joyner-Halenda (BJH) pore size distribution method was determined from the maxima of the BJH desorption pore size distribution curve using the Halsey equation.
  • the micropore area and volume were determined by the t-plot method.
  • compositions of the glucose-containing silica materials prepared by sol-gel reaction and the pore parameters of the porous silicas upon removal of D- glucose using water extraction are summarized below in Table 1 as inventive Examples NEG15 - NEG64.
  • the control sample was formed as set forth above without the pore forming material and data measured without solvent extraction prior to BET measurement (NEG0') and after solvent extraction (NEG0).
  • EXAMPLE 2 The same procedure as set forth above was undertaken, but used sodium hydroxide as a base catalyst for the hydrolysis and/or polycondensation in the formation of the sol-gel matrix.
  • sodium hydroxide was used in place of hydrochloric acid and the same procedure as described in Example 1 was carried out. However, the neutralizing step was not necessary. Varied amounts of 50 weight percent D-glucose solutions were combined with the sol-gel solutions formed with sodium hydroxide. After drying the matrix, transparent, monolithic glucose-containing silica gels were obtained as as- synthesized samples in the same manner as Example 1.
  • the results for the sodium hydroxide samples are also set forth below in Table 1 as samples BEG15 - BEG 64.
  • Example 1 In both Example 1 and Example 2, at high concentrations of D-glucose, the samples sometimes broke into smaller, but still transparent and crack-free pieces during drying. Control samples for Example 2 were also as noted above in Example 1 (samples BEG0' and BEG0). To remove the D-glucose, the as-synthesized samples were ground to fine powders and extracted with a large amount of distilled water at room temperature. The extent of template removal was monitored by infrared spectroscopy and by TGA after extraction.
  • compositions as represented by the glucose concentrations in the as-synthesized materials, calculated from the feed stoichiometry are comparable to those determined from TGA experiments.
  • the small discrepancies could be attributed to incomplete sol-gel reactions or incomplete removal of moisture.
  • the results from nitrogen adsorption-desorption measurements show that addition of D-glucose as the pore forming material in the sol-gel reactions under basic or near neutral conditions modified the microstructure of silica matrices leading to mesoporosity, similar to that achieved using acid catalysis.
  • the nitrogen adsorption-desorption isotherms at -196° C for the porous silica samples after water extraction are shown in Figures 1 and 2.
  • the control samples (NEGO and BEGO) prepared using acid and base catalysts without the pore forming material exhibit reversible Type I isotherms, which are typical of xerogels with microporous structures.
  • the nitrogen sorption isotherms gradually transform from reversible Type I to Type IV isotherms with H2 hysteresis.
  • P/P 0 relative pressures
  • the as-synthesized samples before removing D-glucose exhibit different types of nitrogen isotherms depending on the amount of glucose in the composites. In general, an increase in amount of glucose up to 45 wt% leads to a decrease in surface area and pore volume.
  • the control samples show similar Type I isotherms, surface areas and pore volumes as those after extraction as listed in Table 1. Apparently, the water-extraction procedures may have little effect on the silica structure.
  • the nitrogen absorption isotherms changed from Type I to Type II with H4 hysteresis, which is typical of nonporous solids. The lack of porosity is further evidenced by the very small
  • the pores or channels in the silica materials after removing D-glucose by extraction come from the space previously occupied by the pore forming molecules.
  • analysis on the net pore volume and BET surface which are the differences between the water-extracted and as-synthesized samples, reveals that they are linearly dependent on the D-glucose concentration up to 45 wt%, with good correlation
  • the infrared spectra of both as-synthesized and water-extracted samples show the major absorption bands associated with network Si-O-Si vibrational modes at about 460, 790, 1080, and 1220 cm “1 , along with Si-OH asymmetrical stretching at about 960 cm “ and SiO-H bond stretching at 3400 cm " .
  • intensity of the band at 2940 cm " for C-H stretching of the glucose component in the as- synthesized samples increases with glucose concentration. This band disappeared after water extraction, indicating the removal of the glucose pore forming material.
  • the complete removal of glucose is further supported by the fact that the extracted samples showed little or no weight loss upon heating to 800 °C under air in the TGA measurements.
  • Figures 5 and 6 show the BJH pore size distributions by plotting differential volume as a function of pore size for the silica matrices obtained from the deso ⁇ tion branches of the nitrogen so ⁇ tion isotherms at -196° C according to the BJH method with the Halsey equation for multilayer thickness.
  • the extracted silica matrices possess narrowly distributed mesopores centered at about 3.2 or 3.5 nm, respectively.
  • the BJH pore diameters in Table 1 are around 32 to 35 A at the glucose concentrations of 45 wt% and higher.
  • the BET average pore diameter, derived from V/S- Q ⁇ increases with glucose concentration.
  • Pore size distributions from the adso ⁇ tion branches of isotherms also show a similar pattern as in Figures 5 and 6 but with broader distributions.
  • the pore volumes obtained at P/P 0 of about 1 and the mesopore surface areas calculated from the BJH method are found to increase with the glucose content. Both micropores and mesopores in the materials contribute to the observed pore volumes and areas, which could be differentiated approximately by t- plot analysis. As the glucose content increases, the contribution from the micropores decreases.
  • the pore forming materials of the invention are generally compounds having high affinity for intermediate materials derived from the organosilicon material in the sol-gel matrix, and include the appropriate hydrophilicity and solubility with low volatility. Further, since the pore forming molecules were used in relatively high concentrations and the pore diameters achieved were greater than the size of the individual pore forming molecules, the pore forming molecules are likely present in forms of aggregates whose interactions with the inorganic material in the matrix through hydrogen bonding play an important role in directing mesoporous formation prior to and/or during gelation.
  • the hydrogen bonding between the organic pore forming material and the intermediates of the inorganic species brings the two major components together and forms a homogeneous sol without macroscopic particulation of inorganic species. It may also facilitate the hydrolysis and further condensation of the -OH functional inorganic intermediate species, e.g. silanol groups.
  • the sols in the above Examples underwent gelation faster at higher concentrations of D-glucose which suggests the presence of D-glucose promoting condensation.
  • the affinity of the pore forming material for the intermediate species might keep the inorganic and organic moieties from macroscopic phase separation, while the volatile solvent molecules and reaction by-products, such as alcohol and water, were gradually evaporated from the system.
  • the interactions between the intermediates and the pore forming molecules would stabilize the silicon-based framework and prevent it from fracturing caused by capillary pressure and internal stress build-up during drying.
  • an organic-inorganic composite mesoporous material with bicontinuous networks of silica and the pore forming material were obtained as transparent, monolithic solids, and the removal of the pore forming material provided silica materials with interconnecting mesopores.
  • the resulting mesoporous materials formed using the invention include high specific surface areas, pore volumes and narrow pore distributions with BJH pore diameters of 3.2 to 3.5 nm, indicating mesoporosity.
  • the pore parameters generally increase and the nitrogen so ⁇ tion isotherms gradually transform from Type I to Type IV with H2 hysteresis as the glucose concentration increases in synthesis.
  • concentrations of ⁇ 36 wt% glucose both micropores and mesopores contribute to porosity, and at levels of 36 to 64 wt% glucose, mesopores are dominant with narrow pore size distributions.
  • Acid phosphatase is immobilized in a mesoporous material.
  • Acid phosphatase (ACP, EC 3.1.3.2, Type I from wheat germ, 0.4 units/mg, lot 37H7025) and magnesium chloride were purchased from Sigma, St. Louis, Missouri.
  • Tetramethylorthosilicate and p-nitrophenyl phosphate disodium salt hexahydrate (pNPP) were supplied by Aldrich, Milwaukee, Wisconsin.
  • Methyl alcohol, D-glucose, sodium hydroxide and hydrochloric acid were from Fisher Scientific, Fair Lawn, New Jersey.
  • p-nitrophenol was purchased from Acros Organic in New Jersey. All chemicals were used as received.
  • a homogeneous mixture of 5.0 ml tetramethylorthosilicate, 6.0 ml of methanol, 15 ⁇ l of 40 mmol hydrochloric acid and 0.7 ml of water was formed in a beaker. Varied amounts of 50 wt% D-glucose aqueous solution were added under agitation at room temperature, followed by addition of 10 mg of ACP (in a 1.0 ml water solution) upon cooling the mixture to 0°C. The mixture was sealed in the beaker with a piece of plastic film and allowed to reach room temperature under moderate magnetic stirring.
  • the film was pierced with 12-15 holes using a hypodermic syringe needle to allow the evaporation of solvents.
  • the sample-containing beaker was removed from a fume hood and put into a vacuum oven and dried to react constant weight at room temperature in about six days.
  • Transparent or opaque samples of weights ranging from 2.4 to 5.5 g depending on the concentration of D-glucose in the biogels (from 0-60 wt%) were obtained.
  • the samples were crushed with a mortar and pestle into fine powder (10-100 ⁇ m) and kept in sealed vials in a -15°C freezer ready for enzymatic activity assay or other property evaluation.
  • the assay procedure included weighing a certain amount of the immobilized ACP sample powder (30-80 mg) containing nominally 0.06 units of ACP into a test tube.
  • ACP-free silica glass control AMAC Control
  • 50 mg of powder was used.
  • 15 ml of water was transferred into the sample- containing tube and shaken often at room temperature.
  • the sample was centrifuged for 2-3 minutes to ensure the complete settling of the sample powder at the bottom of the tube. Then, the supernatant was carefully decanted by inverting the tube, and absorbing the last droplet of liquid at the trim of the tube using a sheet of tissue paper if necessary. The washing procedures were repeated two more times at one hour intervals to ensure removal of glucose and any free enzyme which was not trapped. Then magnesium chloride and buffer solutions were pipetted along with water, if required for dilution, into the sample tube and were shaken well. After 30 minutes of activation and equilibrium, the substrate pNPP solution was pipetted into the tube and timed. The mixture was shaken often.
  • the determination of the reaction kinetics of the free and immobilized enzyme was carried out in a pH 6.2 citrate buffer.
  • 0.06 units of ACP were mixed with the assay solution containing from 0.02 to 5.0 mmol of the substrate, pNPP.
  • a 50 mmol citrate buffer in the pH range of 4.0-5.9 was used.
  • the immobilized ACP was first washed with water (3 X 10 ml), and then mixed with the assay mixture (pH 6.2) without adding pNPP. Free ACP was dissolved in the same solution.
  • part of the supernatant was separated from the sample powder by centrifugation and decantation when the activity assay was completed.
  • the absorbance change after alkalinizing the supernatant with sodium hydroxide with time was subsequently monitored and compared to that of a blank control to determine any enzymatic activity.
  • the characterization of the microstructure parameters, for example, pore size and distribution, specific surface area and pore volume of the mesoporous matrix was carried out on a Micromeritics 2010 system using procedures as described in Examples 1 and 2.
  • the powder sample (from 0.2 -0.3 g) was extracted with water ( 9 x 10 ml) at room temperature for 3 days before the measurement.
  • the sample was first dried in a 115 °C oven and then degassed at 200 °C and 10 torr overnight prior to nitrogen adso ⁇ tion-deso ⁇ tion measurement at -196°C.
  • the pore parameters were calculated using the Micromeritics software.
  • the weight percentage of D-glucose in dry biogel sample was determined from the weight loss in air at 750° C at which glucose decomposed completely.
  • the sample was first dried in a 110°C oven overnight, and then heated to 750 °C at a heating rate of 10°C/minute in a muffle furnace.
  • the effect of glucose concentration on immobilized ACP activity was determined as follows.
  • the apparent specific enzyme activities, were determined as noted above for all samples and appear in ⁇ mol/min.-mg of ACP for the immobilized and free ACP determined at pH 5.0 and [pNPP] 2.0 mmol at room temperature and are summarized in Table 2.
  • the catalytic activities of the entrapped ACP as well as the free ACP are larger at pH 5.0 than at higher or lower pH. This is in agreement with the optimal pH 5.0 previously reported for ACP obtained from wheat germ.
  • the entrapped ACP retained from 7% to 22 % of the free enzyme activity which was estimated to be 0.596 ⁇ mol/min-mg under the same conditions.
  • the immobilized ACP had similar percentage activity remaining when compared to the free ACP activity, as will be discussed also with respect to the pH profile.
  • the enzymatic activities for the immobilized samples were found to be dependent on the concentration of D-glucose used in the preparation of the biogels. That is, when 36 wt % or less glucose was added, the resultant gels did not exhibit remarkably improved enzymatic activities in comparison with the glucose-free sample.
  • the gels demonstrated higher activity than the glucose-free sample, increasing with the amount of glucose (Table 2).
  • the sample exhibited almost three-fold activity over the glucose-free sample, which is about 22 % of the free ACP activity.
  • the difference in apparent catalytic activity for the entrapped ACP samples was associated with the biogel microstructure.
  • their enzymatic activities were associated with the concentration of D-glucose used in the formulation of the bioactive gels.
  • the matrix pore structure parameters e.g., the specific surface area and pore volume as well as pore size and distribution, were also dependent on the concentration of glucose present in the gels in accord with Examples 1 and 2.
  • the pore parameters of the silica matrix obtained by removing D-glucose from the as-synthesized materials via solvent extraction, revealed further information about the biogel microstructure.
  • the pore structure parameters of the silica matrices, obtained from nitrogen so ⁇ tion isotherms, are also summarized in Table 2.
  • the nitrogen so ⁇ tion isotherms at -196°C (Fig. 7) show that when neither D-glucose nor ACP is added, the resultant matrix exhibits a Type I isotherm and is essentially a microporous silica, similar to the conventional xerogels prepared in the presence of acid catalyst.
  • ACP/gram of Si ⁇ 2) was present in the biogels without adding glucose (the glucose-free sample), micropores and mesopores coexisted in the corresponding product, as evidenced by the t plot.
  • glucose the glucose-free sample
  • micropores and mesopores coexisted in the corresponding product, as evidenced by the t plot.
  • both ACP and glucose were used to formulate the composite biogels, the nitrogen so ⁇ tion isotherms for the matrices shifted from Type I to Type IV with increasing irreversibility as indicated by the growing size of hysteresis loops while the concentration of D-glucose increased.
  • the resultant matrices had smaller pore volumes and pore diameters than the glucose-free sample, but had comparable BET surface areas, similar to the results of Examples 1 and 2.
  • the matrices made with more than 42 wt % of glucose had larger pore volumes and pore diameters than the glucose-free sample and increased with glucose concentration (Table 2). It is noted that at high pore forming material concentrations, the BJH surface areas, reflecting mesopores > 17, also increased with glucose concentration, while the BET surface areas remained about the same.
  • Figures 8 and 9 show, respectively, the BJH adso ⁇ tion and deso ⁇ tion pore size distributions of the biogel matrices.
  • the individual pore size distribution curve was obtained from the adso ⁇ tion or deso ⁇ tion branch of the nitrogen so ⁇ tion isotherms by plotting differential volume as a function of pore size. It is noted that the pore size distribution for a certain sample derived from the nitrogen adso ⁇ tion branch was broader than that from the deso ⁇ tion branch, and the corresponding average pore diameter was also greater when calculated from adso ⁇ tion isotherm than that from the deso ⁇ tion branch.
  • the irreversibility of the nitrogen isotherms may be attributable to the existence of ink-bottle pores and interconnecting pore network structure in the silica matrices prepared using this pore forming material as described above.
  • the BJH adso ⁇ tion pore size distributions may reflect the cavity size distributions, while the deso ⁇ tion pore size distributions may correspond to the throat size distributions. If this is the case, then Figures 8 and 9 demonstrate that both cavity and throat sizes increase with D-glucose concentration when glucose is 20 wt % or higher in the biogels. When D-glucose accounts for 42-60 wt % of the biogels, the extracted silica matrices possess narrowly distributed pore openings centered at 32.5 to 34.5 A.
  • [pNPP] 0.5-5.0 mmol/ L.
  • the BET surface area in the range of 650-820 m /g for the ACP- containing samples, does not seem to be a significant factor affecting the rate in the immobilized enzyme-catalyzed reaction as in the conventional chemical reactions using heterogeneous catalysts. This may be attributable to the difference in the active sites of these two kinds of catalysts. It is generally accepted that active sites on the surface are important for framework-confined catalysts which require large areas exposed to and attainable by reactants for an effective reaction. But for the enzyme-catalyzed reactions, it seems that the transfer of mass from the bulk solution to the active site of the immobilized enzyme is critical. Therefore, the pore diameter and pore volume of the carrier, instead of its BET surface area, are the more important factors affecting the overall catalytic activity of a highly active enzyme.
  • This Example demonstrates the direct immobilization of the enzyme molecules in mesoporous silica glasses through use of the neutral organic pore forming materials in accordance with the invention.
  • the biologically active composites comprising enzyme macromolecules, low-molecule-weight organic compound and highly crosslinked silica networks
  • the aggregation or assembly of the aggregates of the pore forming molecules and the hydrogen bonding between the intermediate silicate species and the pore forming molecules are believed to direct the formation of the nanophases in the hybrid materials as discussed above.
  • the enzyme macromolecules only comprising 0.5 wt % of silica though, may play a similar role.
  • the degree of freedom for the migration of chemicals inside the glucose-formed matrix would be greater than within the glucose-free sample.
  • the channels or pore sizes of the silica matrix shall be adjusted so as to facilitate the penetration of small molecules such as substrate but not to allow the leakage of the entrapped enzyme macromolecules to the bulk solution.
  • the effect of the substrate pNPP concentration on the reaction kinetics of free and immobilized ACP was investigated to further discern if the immobilized enzyme-catalyzed reaction was pore-diffusion controlled, kinetically controlled, or a combination of both.
  • the assay mixture consisted of pNPP in the range of 0.02 to 5.0 mmol/L in a pH 6.2 citrate buffer. The plot of specific enzyme activity vs.
  • pNPP for free and several immobilized ACP samples synthesized in the presence of high concentrations of glucose (50, 55, and 60 wt% respectively), are demonstrated in Figs. 11a and l id.
  • the various derived forms of the Michealis-Menten equation such as Lineweaver-Burk, Eadie-Hofstee or Hanes-Woolf plot, may be used to construct the enzyme activity versus the substrate concentration graphs and to estimate the maximum velocity (V max ) and Michealis constant (K m ) under the assay condition used.
  • V ma ⁇ and K m values were 0.017 ⁇ mol/min-mg and 0.75 mmol, 0.023 ⁇ mol/min-mg and 0.85 mmol, and 0.030 ⁇ mol/min-mg and 0.91 mmol, respectively, for the immobilized ACP samples made with 50, 55, and 60 wt % of glucose.
  • V ma ⁇ and K m would be larger if they were derived from the Eadie-Hofstee as shown in Fig. 1 le or the Hanes-Woolf plots.
  • V max and K m would be associated with the matrix pore size in the same manner regardless of which of the three plots were used. That is, both V ma ⁇ and K m increased with pore diameter of the matrices. But, V ma ⁇ for the immobilized ACP was smaller than that of free ACP while K m was larger than that of the free enzyme. This is in agreement with the expected effect of immobilization on the apparent V max and K m values.
  • Table 3 shows that both free and immobilized ACP have similar pH profiles, with optimal pH 5.0. No shifting in optimal pH for the immobilized ACP was observed. This result is in agreement with the reported value of optimal pH 5.0 for wheat germ ACP indicating neither significant electrostatic interactions between the matrix and the enzyme nor the occurrence of significant pH difference inside and outside the matrix. Table 3 also clearly demonstrates that the apparent activities for all samples in the pH range 4.0-5.9 are related to the matrix pore diameters which are associated with concentration of D-glucose used in the preparations. The immobilized ACP activities under different pH values display similar dependence on glucose concentration as previously discussed. The relative activity among the immobilized ACP samples was basically independent of the pH of assay mixture, indicating the apparent activity is limited by the mass transport barrier of the matrix.
  • thermal stability of enzymes due to immobilization is presumed to be caused by restricting the segmental motion of protein chains, driven by the gain of entropy by freeing of bound water, so as to reduce the possibility of an irreversible structure change, or by reducing denaturing segmental collision with the surface of the host through attachment to the surface.
  • Evaluation of the stability of ACP from wheat germ showed loss of activity was the result of surface inactivation together with a sensitivity to dilution.
  • Table 4 shows that thermal stability of ACP remarkably improved upon immobilization.
  • the glucose-free sample displayed the highest percentage retention of activity (87 %) after treatment at 50 °C in citrate buffer for one hour.
  • the thermal stability of immobilized ACP prepared in the presence of glucose did not su ⁇ ass that of the glucose-free sample.
  • the samples made with either low (30 % or below) or high concentration (55-60 wt %) of glucose showed over 50 % of their initial activities after treatment at 50° C for one hour.
  • the large free space may increase the chance of denaturing segmental collision of ACP with the surface of the host during heat treatment and consequently result in lower thermal stability and enzymatic activity.
  • the large free space in the near vicinity of the active site of ACP may improve catalytic activity through enhanced rate of mass transfer within the matrix, as previously discussed. Therefore, it is favorable to have relatively small free space surrounding the protein molecules in order to reduce the possibility of denaturing segmental collisions during thermal treatment but to have spacious channels to help ease of internal mass transport during assay. Compromise of these two factors leads to the trend in the thermal stability of immobilized ACP shown in Table 4.
  • the percentage retention of activity in the third cycle was 80% for the glucose-free sample, and 80, 90 and 98 % for the samples made, respectively, with 50, 55 and 60 wt % of glucose.
  • the loss in activity for the other samples was larger after each cycle of assay.
  • ACP was directly immobilized in microporous and mesoporous silica glasses without leakage via sol-gel processing.
  • the immobilized ACP retained 7-22 % of its native activity at its optimal pH 5.0, depending on the glucose concentration used in the preparation of the biogels.
  • D-glucose served as an appropriate pore-forming material for the synthesis of silica gel-immobilized biologically active agents.
  • the addition of glucose as the pore forming material not only modified the pore structure parameters but also enhanced the apparent activity of the immobilized enzyme.
  • the pore size and pore volume of the silica matrix, as well as the catalytic activity of the immobilized ACP, increased with the concentration of glucose in the preparations.
  • D-glucose is an economical and environmentally-friendly pore forming material with good biocompatibility with enzymes, it may be widely used in the preparation of immobilized biologically active agents via the sol-gel reactions described herein in accordance with the invention.
  • mesoporous biogels with increased pore size, pore volume and mesopore surface area formed in accordance with the invention exhibit improved catalytic activity owing to greater mass transfer of chemicals within the pores.
  • this is of great importance in applications requiring monolithic or bulky bioglasses.
  • the increased pore size in the mesoporous matrix makes penetration of relatively large substrates within the gel possible due to reduced steric hindrance.
  • the mesoporous materials provide an effective shell which may prevent the migration of protein macromolecules into or out of the open pores.
  • the mesoporous matrix with can achieve a specified pore diameter and relatively narrow pore distribution, which likely causes the preferable abso ⁇ tion and selective reaction of molecules with critical sizes.
  • This Example also demonstrates that it is very difficult, if not impossible, to determine the degree of denaturation or activity remaining of an enzyme after immobilization in a highly crosslinked organic-inorganic matrix since the activity assay only gives the overall apparent activity, controlled by internal diffusion-limited kinetics, that is often used to estimate the activity remaining.
  • the true extent of deactivation or activity remaining of an enzyme upon immobilization in a porous carrier might be ascertained if one can find a proper homogenizing method. It shall be pointed out that the immobilization of ACP in this Example is only an illustration of principles of this novel improved synthesis technique and the preparation conditions may be further optimized to achieve higher retention of ACP activity or to immobilize other biologically active materials.
  • Horseradish peroxidase was directly immobilized in mesoporous silicon-based matrices using the procedure for sol-gel reactions as described in Example 3 in the presence of various pore forming materials according to the invention, including D-fructose (samples FH16-FH60), D-glucose (Samples GH33 and GH42), sucrose (Samples SH33 and SH42), and glycerol (Samples YH33 and YH42).
  • each pore forming material is expressed below in Table 5 based on the amount of silicon dioxide and organic compound, along with the mean V max based on the average V max from the Eadie-Hofstee plot and the Hanes-Woolf plot, as described in other Examples herein.
  • Table 5 also provides the standard deviation for the V ma ⁇ data.
  • Table 5 also sets forth the mean K m value, as described in other
  • the apparent catalytic activities of the entrapped enzyme showed up to more than 20% of the free HRP when assayed with a calorimetric method using 4-aminoantipyrine (4-AAP)-phenol (PHOH) - hydrogen peroxide (H2O2) reagents at a pH of 6.5 and room temperature.
  • the activity was also measured as activity remaining after the sample underwent thermal treatment at 60 °C for 30 minutes before assaying at room temeprature, in comparison with the corresponding sample without thermal treatment as set forth as "Activity Remaining" in Table 5.
  • Free HRP in solution retained 50%) of its activity after such thermal treatment.
  • Table 5 also includes the dry mass (g) of the total as-synthesized dry samples obtained after drying in vacuum at room temperature.
  • the experimental data demonstrated that immobilized HRP exhibited maximum activities at a higher concentration of hydrogen peroxide than free enzyme, due to the substrate, hydrogen peroxide, inhibition and diffusion limitation within the matrix pores.
  • the matrices obtained from removal of the pore forming materials set forth above from the sol-gel matrices formed and dried, using water extraction at room temperature, had interconnecting mesopore structures, as evidenced by nitrogen so ⁇ tion measurements conducted using nitrogen adso ⁇ tion-deso ⁇ tion isotherms at -196°C on an ASAP 2010 as described above. The measurement data are provided in Table 6.
  • the type of isotherms generated in accordance with such procedures as described in the other Examples herein are also listed in Table 6.
  • the mesoporous structures and pores prevented leaching of HRP from the mesoporous materials and provided easy access to low molecular weight reactants.
  • the catalytic activities of immobilized HRP were found to be closely related to the pore sizes and volumes of the mesoporous matrix materials which, in turn, were associated with the concentration of the pore forming compounds used in the synthesis of the composite biogels in accord with the findings of Example 3. Thermal stability of HRP upon immobilization was greatly improved.
  • Example and Example 3 demonstrate that the synthesis of mesoporous materials having immobilized biologically active agents using the pore forming materials of the invention provide versatility in terms of the biologically active agents and pore forming compounds, and indicate usefulness of these immobilized biologically active agents as various biocatalysts and biosensors.
  • EXAMPLE 5 In this Example, various preferred pore forming materials were used to make mesoporous materials, specifically D-glucose, dibenzoyl-L-tartaric acid, and D- maltose. Upon removing the pore forming materials, it was found that mesoporous materials were achieved. The samples were analyzed using XRD diffraction patterns which suggested a disordered channel assembly. The pore diameters achieved varied from 2 ⁇ A to about 6 ⁇ A depending upon pore forming material concentration.
  • the organometallic compound used to form the mesoporous material in this Example was tetraethylorthosilicate.
  • the tetraethylorthosilicate was dissolved in an ethanol solution and combined with distilled water and a hydrochloric acid as the acid catalyst in molar ratios of tetraethylorthosilicate:water:catalyst:ethanol of 1 :4:0.01 :3.
  • the reactants were prehydrolyzed at 60°C for about 1-2 hours, during which the initial phase-separation disappeared and the mixture became homogeneous. This was followed by reflux for 2 hours.
  • the solution was combined with an aqueous solution of D-glucose (0.8 mol/1), an aqueous solution of D-maltose (1.2 mol/1) or an ethanol solution of dibenzoyl-L-tartaric acid (0.3 mol/1) while stirring.
  • the homogeneous solutions obtained were cast into cylindrical polystyrene molds followed by sealing of the molds with a cover having 2-3 pinholes for allowing evaporation of volatile solvents and byproducts.
  • the reaction was allowed to stand at room temperature for gelation and slow drying for 15-20 days to provide a colorless, transparent (>80% transmission in visible light range), monolithic disk of the pore forming compound-containing silicon- based matrix.
  • the mesoporous materials were formed by removing the pore forming materials.
  • the materials were removed by first grinding the disks to fine powder, followed by Soxhlet extraction with methanol or water for 2-3 days followed by drymg at 100° C overnight.
  • the extent of pore forming material removed was monitored using TGA as described above for weight loss at 750 °C at which the pore forming materials decomposed completely.
  • the BET surface areas and pore volumes were determined before and after extraction by nitrogen adso ⁇ tion-deso ⁇ tion isotherm measurements.
  • the unextracted samples had relatively small BET surface areas and pore volumes.
  • the surface areas and pore volumes increased drastically.
  • the surface areas were found to correlate linearly with the extent of pore forming material removed at various extraction times. For example, the surface are of Sample DB40 in Table 7
  • the nitrogen adso ⁇ tion-deso ⁇ tion isotherms were determined at various relative pressures (P/P 0 ) on the mesoporous material samples.
  • P/P 0 relative pressures
  • the samples DB40X, DB50X, and DB60X prepared with high dibenzoyl-L-tartaric acid concentrations of ⁇ 40 wt% exhibited Type IV isotherms with Type H2 hysteresis loops
  • sample DB20X prepared with low pore forming concentration of dibenzoyl-L-tartaric acid of 16 wt% shows a completely reversible Type I isotherm, typical of microporous structures.
  • the position of the well-defined step in the nitrogen deso ⁇ tion isoterms appears to shift to higher relative pressures as the pore forming material concentration is increased. These characteristics are similar to those observed for prior art surfactant pore formers and can be attributed to capillary condensation within narrow tubular mesopores of 30-60 A in effective diameter such that the pores may be tubular. Furthermore, the ratio of pore volume to surface area in this Example was found to be close to one half the average radius as indicated by (P7S B E ⁇ )/(r av /2) values approximating unity which is in agreement with tubular channels of average radius. The pore size and its distribution were also found to be influenced by concentration of pore forming materials as shown in Fig. 12.
  • the dominant pore diameter increased from about 34 to 60 A.
  • the fidelity of pore distribution ranges from 3 to 8 A, which is comparable to values achieved using surfactant pore formers.
  • the type and size of pore forming materials seemed to have relatively less effect on the pore size distribution, for example, at approximately the same pore forming concentration (50 wt%), the D-glucose and D- maltose formed mesoporous structures (DB50X and MT50X, respectively) have almost identical BJH pore size distributions although their molecule sizes are very different.
  • the variation in the pore diameter is only 4A between DB50X and MT50X as shown in Table 7. These observations differ from similar observations based on surfactant pore forming materials.
  • the XRD pattern of the mesoporous materials prepared with 48% of D- maltose is shown in Fig. 13.
  • the pattern appears to have a peak (100 reflection) with a ⁇ ?-spacing of 62 A and a broad, low intensity peak (d of about 35-31 A) which could possibly be caused from overlapped 110 and 200 reflections.
  • Such a pattern is consistent with mesoporous structures.
  • Consideration of the thickness of the pore walls would indicate that the -spacing value for the 100 reflection is comparable to the BJH pore diameter (32A).

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Abstract

La présente invention concerne des matières mésoporeuses et un procédé de fabrication de ce type de matières selon lequel elles sont fabriquées par la formation d'une solution aqueuse contenant un composé organométallique. On y ajoute une solution comprenant une matière porogène sélectionnée dans le groupe constitué par des polyols monomères, des polyacides, des polyamines, des hydrates de carbone, des oligopeptides, des acides oligonucléiques, des composés organiques à fonction carbonyle, et des mélanges et dérivés de ces matières, de manière à former une matrice sol-gel par polycondensation. On fait sécher ensuite la matrice sol-gel et on retire de cette matrice ainsi séchée la matière porogène, de manière à former une matière mésoporeuse. Les matières mésoporeuses présentent des diamètres de pores compris entre environ 20 Å et env iron 100 Å et peuvent être utilisées avec un agent biologiquement actif immobilisé dans les pores de la matière mésoporeuse et introduit dans un système biologique.
PCT/US1999/001116 1998-01-20 1999-01-20 Matieres mesoporeuses et leurs procedes de fabrication WO1999036357A1 (fr)

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WO2000051724A1 (fr) * 1999-02-27 2000-09-08 The Procter & Gamble Company Procede de protection et/ou de liberation controlee d'ingredients actifs
WO2002010218A1 (fr) * 2000-07-31 2002-02-07 Drexel University Encapsulation directe de biomacromolecules dans des materiaux mesoporeux et nanoporeux a matrice tensioactive
EP1358873A1 (fr) * 2002-04-30 2003-11-05 National Institute of Advanced Industrial Science and Technology Silice mésoporeuse ayant une fonction de controle permettant la mise en ou hors-service de la libération controllée de composés, méthode pour sa préparation et son utilisation
US6989254B2 (en) 2000-07-31 2006-01-24 Drexel University Direct encapsulation of biomacromolecules in surfactant templated mesoporous and nanoporous materials
US7163611B2 (en) 2003-12-03 2007-01-16 Palo Alto Research Center Incorporated Concentration and focusing of bio-agents and micron-sized particles using traveling wave grids
US7309410B2 (en) 2003-12-03 2007-12-18 Palo Alto Research Center Incorporated Traveling wave grids and algorithms for biomolecule separation, transport and focusing
EP1789366A4 (fr) * 2004-07-06 2008-11-26 Agency Science Tech & Res Nanoparticules mesoporeuses
FR2942622A1 (fr) * 2009-02-27 2010-09-03 Commissariat Energie Atomique Procede de preparation de particules de silice poreuses, lesdites particules et leurs utilisations
US20110086099A9 (en) * 2003-06-27 2011-04-14 K.U. Leuven Research & Development Crystalline mesoporous oxide based materials useful for the fixation and controlled release of drugs
US7981441B2 (en) 2004-02-18 2011-07-19 The Board Of Trustees Of The Leland Stanford Junior University Drug delivery systems using mesoporous oxide films
US8097269B2 (en) 2004-02-18 2012-01-17 Celonova Biosciences, Inc. Bioactive material delivery systems comprising sol-gel compositions
US8258137B2 (en) 2008-01-29 2012-09-04 Katholieke Universiteit Leuven Process for release of biologically active species from mesoporous oxide systems
CN101653975B (zh) * 2003-06-24 2013-02-13 斯攀气凝胶公司 凝胶片的制造方法
CN104523643A (zh) * 2014-12-23 2015-04-22 南京先宇科技有限公司 一种稳定的羟苯磺酸钙制备方法
US9114125B2 (en) 2008-04-11 2015-08-25 Celonova Biosciences, Inc. Drug eluting expandable devices
CN105025928A (zh) * 2013-03-15 2015-11-04 普西维达公司 用于递送治疗剂的可生物蚀解的硅基组合物
US9808421B2 (en) 2010-11-01 2017-11-07 Psivida Us, Inc. Bioerodible silicon-based devices for delivery of therapeutic agents
US9962396B2 (en) 2009-05-04 2018-05-08 Psivida Us, Inc. Porous silicon drug-eluting particles
CN110813286A (zh) * 2018-08-14 2020-02-21 中国石油化工股份有限公司 载体为含有面包圈介孔材料和硅胶的复合材料的异丁烷脱氢催化剂及其制法和应用
CN110813285A (zh) * 2018-08-14 2020-02-21 中国石油化工股份有限公司 载体为球形面包圈介孔材料硅胶复合材料的异丁烷脱氢催化剂及其制法和应用
CN114497585A (zh) * 2022-01-27 2022-05-13 中国科学院青岛生物能源与过程研究所 一种具有结构耦合效应的铂基协同催化剂的制备方法
WO2023068928A1 (fr) 2021-10-19 2023-04-27 Tijani Holding B.V. Polymère ou particule biosoluble pour l'administration d'un agent actif et son procédé de production

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WO2000051724A1 (fr) * 1999-02-27 2000-09-08 The Procter & Gamble Company Procede de protection et/ou de liberation controlee d'ingredients actifs
US6989254B2 (en) 2000-07-31 2006-01-24 Drexel University Direct encapsulation of biomacromolecules in surfactant templated mesoporous and nanoporous materials
WO2002010218A1 (fr) * 2000-07-31 2002-02-07 Drexel University Encapsulation directe de biomacromolecules dans des materiaux mesoporeux et nanoporeux a matrice tensioactive
US6902806B2 (en) 2002-04-30 2005-06-07 National Institute Of Advanced Science And Technology Mesoporous silica having controlled-release on-off control function, production method thereof and method using same
EP1358873A1 (fr) * 2002-04-30 2003-11-05 National Institute of Advanced Industrial Science and Technology Silice mésoporeuse ayant une fonction de controle permettant la mise en ou hors-service de la libération controllée de composés, méthode pour sa préparation et son utilisation
CN101653975B (zh) * 2003-06-24 2013-02-13 斯攀气凝胶公司 凝胶片的制造方法
US8273371B2 (en) * 2003-06-27 2012-09-25 Johan Adriaan Martens Crystalline mesoporous oxide based materials useful for the fixation and controlled release of drugs
US20110086099A9 (en) * 2003-06-27 2011-04-14 K.U. Leuven Research & Development Crystalline mesoporous oxide based materials useful for the fixation and controlled release of drugs
US7163611B2 (en) 2003-12-03 2007-01-16 Palo Alto Research Center Incorporated Concentration and focusing of bio-agents and micron-sized particles using traveling wave grids
US7309410B2 (en) 2003-12-03 2007-12-18 Palo Alto Research Center Incorporated Traveling wave grids and algorithms for biomolecule separation, transport and focusing
US7981441B2 (en) 2004-02-18 2011-07-19 The Board Of Trustees Of The Leland Stanford Junior University Drug delivery systems using mesoporous oxide films
US8097269B2 (en) 2004-02-18 2012-01-17 Celonova Biosciences, Inc. Bioactive material delivery systems comprising sol-gel compositions
EP1789366A4 (fr) * 2004-07-06 2008-11-26 Agency Science Tech & Res Nanoparticules mesoporeuses
US8258137B2 (en) 2008-01-29 2012-09-04 Katholieke Universiteit Leuven Process for release of biologically active species from mesoporous oxide systems
US10285968B2 (en) 2008-04-11 2019-05-14 Celonova Biosciences, Inc. Drug eluting expandable devices
US9949944B2 (en) 2008-04-11 2018-04-24 Celonova Biosciences, Inc. Drug eluting expandable devices
US9114125B2 (en) 2008-04-11 2015-08-25 Celonova Biosciences, Inc. Drug eluting expandable devices
FR2942622A1 (fr) * 2009-02-27 2010-09-03 Commissariat Energie Atomique Procede de preparation de particules de silice poreuses, lesdites particules et leurs utilisations
JP2010202505A (ja) * 2009-02-27 2010-09-16 Commissariat A L'energie Atomique & Aux Energies Alternatives 多孔質シリカ粒子を調製するプロセス、その粒子およびその使用
EP2228127A1 (fr) * 2009-02-27 2010-09-15 Commissariat à l'Énergie Atomique et aux Énergies Alternatives Procédé de préparation de particules de silice poreuses, lesdites particules et leurs utilisations
US9962396B2 (en) 2009-05-04 2018-05-08 Psivida Us, Inc. Porous silicon drug-eluting particles
US9808421B2 (en) 2010-11-01 2017-11-07 Psivida Us, Inc. Bioerodible silicon-based devices for delivery of therapeutic agents
US11026885B2 (en) 2010-11-01 2021-06-08 Eyepoint Pharmaceuticas, Inc. Bioerodible silicon-based devices for delivery of therapeutic agents
EP2968571A4 (fr) * 2013-03-15 2016-09-07 Psivida Inc Compositions à base de silicium bioérodables pour l'administration d'agents thérapeutiques
JP2016512839A (ja) * 2013-03-15 2016-05-09 シヴィダ・ユーエス・インコーポレイテッドＰｓｉｖｉｄａ Ｕｓ， Ｉｎｃ． 治療物質の送達のための生体内分解性ケイ素系組成物
US9980911B2 (en) 2013-03-15 2018-05-29 Psivida Us, Inc. Bioerodible silicon-based compositions for delivery of therapeutic agents
AU2014235051B2 (en) * 2013-03-15 2019-01-17 Eyepoint Pharmaceuticals Us, Inc. Bioerodible silicon-based compositions for delivery of therapeutic agents
CN105025928A (zh) * 2013-03-15 2015-11-04 普西维达公司 用于递送治疗剂的可生物蚀解的硅基组合物
CN104523643A (zh) * 2014-12-23 2015-04-22 南京先宇科技有限公司 一种稳定的羟苯磺酸钙制备方法
CN110813286A (zh) * 2018-08-14 2020-02-21 中国石油化工股份有限公司 载体为含有面包圈介孔材料和硅胶的复合材料的异丁烷脱氢催化剂及其制法和应用
CN110813285A (zh) * 2018-08-14 2020-02-21 中国石油化工股份有限公司 载体为球形面包圈介孔材料硅胶复合材料的异丁烷脱氢催化剂及其制法和应用
WO2023068928A1 (fr) 2021-10-19 2023-04-27 Tijani Holding B.V. Polymère ou particule biosoluble pour l'administration d'un agent actif et son procédé de production
CN114497585A (zh) * 2022-01-27 2022-05-13 中国科学院青岛生物能源与过程研究所 一种具有结构耦合效应的铂基协同催化剂的制备方法

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