EP1778192A2 - Systemes de microparticules pour l'administration par voie orale de substances bioactives - Google Patents

Systemes de microparticules pour l'administration par voie orale de substances bioactives

Info

Publication number
EP1778192A2
EP1778192A2 EP05760593A EP05760593A EP1778192A2 EP 1778192 A2 EP1778192 A2 EP 1778192A2 EP 05760593 A EP05760593 A EP 05760593A EP 05760593 A EP05760593 A EP 05760593A EP 1778192 A2 EP1778192 A2 EP 1778192A2
Authority
EP
European Patent Office
Prior art keywords
solution
microparticulate systems
process according
biocompatible
biologically active
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Withdrawn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
EP05760593A
Other languages
German (de)
English (en)
Inventor
Daniele Vigo
Vincenzo Russo
Massimo Faustini
Mario Francesco Pace
Eleonora Munari
Maria Luisa Torre
Ubaldo Conte
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.)
Universita degli Studi di Milano
Original Assignee
Universita degli Studi di Pavia
Universita degli Studi di Milano
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 Universita degli Studi di Pavia, Universita degli Studi di Milano filed Critical Universita degli Studi di Pavia
Publication of EP1778192A2 publication Critical patent/EP1778192A2/fr
Withdrawn legal-status Critical Current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
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A61K9/1605—Excipients; Inactive ingredients
    • A61K9/1629—Organic macromolecular compounds
    • A61K9/1652—Polysaccharides, e.g. alginate, cellulose derivatives; Cyclodextrin
    • 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
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A61K9/1605—Excipients; Inactive ingredients
    • A61K9/1617—Organic compounds, e.g. phospholipids, fats
    • A61K9/1623—Sugars or sugar alcohols, e.g. lactose; Derivatives thereof; Homeopathic globules
    • 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
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A61K9/1605—Excipients; Inactive ingredients
    • A61K9/1629—Organic macromolecular compounds
    • A61K9/1635—Organic macromolecular compounds obtained by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyvinyl pyrrolidone, poly(meth)acrylates
    • 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
    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
    • A61K9/1682—Processes
    • A61K9/1694—Processes resulting in granules or microspheres of the matrix type containing more than 5% of excipient

Definitions

  • the present invention relates to the preparation, use and formulation of microparticulate systems for the oral administration of biologically active substances.
  • the proteins, lipids and carbohydrates contained in food must be broken down into their elementary components . in order to be . absorbed by the intestine.
  • non-ruminant polygastric animals such as: the young of ruminant species with nonfunctional pre-stomachs, birds and fish
  • post-omasal digestive tract of ruminants such as: bovids, ovicaprids, camelids and buffalos
  • digestion is principally enzymatic, assisted by mechanical and microbial processes.
  • digestive enzymes are secreted into the lumen of the digestive system ' from the glands associated with it (salivary glands, exocririe pancreas and the liver) or contained in the mucosa.
  • the mechanical processes associated with digestion and, particularly the movements of peristalsis, ,antiperistalsis and segmentation ensure an effective action of the digestive enzymes by increasing the available surface of the foodstuffs, thus promoting improved distribution inside the intestine.
  • Such mechanical processes are also responsible for the expulsion of undigested or unabsorbed materials. Protein digestion begins in the stomach of monogastric and non-ruminant polygastric animals and in the abomasum of polygastric ruminants.
  • the digestive enzymes •are pr ⁇ teolytic enzymes, such as for example pepsin, which are active at acidic pH.
  • All ' protein ' molecules in the gastric lumen endure such action and, hence, the enzymes and structural proteins present in cell walls and/or in plasma membranes also endure this process.
  • the main complex carbohydrates in the feedstuffs for post-weaning subjects are starches.
  • Complex carbohydrates, such as starch are mainly digested in the intestine thanks to the action of special enzymes, such as for example ⁇ -amylase, which are secreted into the intestinal lumen and display maximal activity at pH values about 7.
  • ⁇ -amylase is produced by the exocrine pancreas and secreted in an aqueous solution containing sodium bicarbonate and other electrolytes, having the role of buffering the pH of the intestinal tract in the range of maximal enzyme activity (pH 6.5-7.0) .
  • exocrine pancreas functional development is progressive and induced by growth factors , present in mother's milk: chymotrypsin secretion begins around the first week of life, lipase appears around the second week, and ⁇ -amylase only appears ' from the fourth week onwards .
  • chymotrypsin secretion begins around the first week of life, lipase appears around the second week, and ⁇ -amylase only appears ' from the fourth week onwards .
  • starch digestion is reduced or compromised due to the lack of this enzyme.
  • dietary supplements containing biologically active substances, such as, for example: enzymes, peptides, polypeptides, aminoacids, yeasts, microorganisms, milk enzymes etc., in order to improve digestion and, hence, absorption of nutritional substances; drugs, vaccines, hormones, drug and hormone precursors etc., in order to treat any diseases which might possibly arise.
  • biologically active substances such as, for example: enzymes, peptides, polypeptides, aminoacids, yeasts, microorganisms, milk enzymes etc.
  • drugs, vaccines, hormones, drug and hormone precursors etc. in order to treat any diseases which might possibly arise.
  • the preferred route of administration is orally, in that it is the simplest (said substances are combined with the solid or liquid diet) , the most economical and may even be used for slightly uncollaborative subjects.
  • oral administration has problems due to the fact that the aforementioned biologically active substances may be partially degraded by gastric juices and, hence, either partially or totally, loose their activity.
  • enzymes such as ⁇ -amylase undergo extensive modifications to their primary, secondary and supersecondary structures in acid environment and, by the time they reach the intestine, they are completely inactivated.
  • the technical problem addressed by this invention is that of the successful oral administration of biologically active substances to animals, without incurring in the complete loss of their biological activities .
  • the present invention relates to microparticulate systems consisting of a gastroresistant, biocompatible and biodegradable polymer matrix, comprising a gastroresistant and- enterosoluble polymer, a cryoprotector or lyoprotector, a divalent or trivalent metal salt of a biocompatible and biodegradable polymer having acidic groups and biologically active substances.
  • An additional biocompatible and biodegradable polymer may also optionally be present.
  • biologically active substances animal, plant, bacterial or recombinant derived proteins and peptides and precursors thereof, classified under the following categories: enzymes, coagulation factors, haemopoietic growth factors, interferons, polypeptide and protein ' antibiotics, immunoglobulins, insulin, growth hormones, gonadotropins, protein molecules, antigenic glycoproteins and lipoproteins, prokaryotic cells and viruses, vaccines, drugs, hormones, phyto- and organo-therapeutic ' extracts and mixtures thereof.
  • enzymes enzymes, coagulation factors, haemopoietic growth factors, interferons, polypeptide and protein ' antibiotics, immunoglobulins, insulin, growth hormones, gonadotropins, protein molecules, antigenic glycoproteins and lipoproteins, prokaryotic cells and viruses, vaccines, drugs, hormones, phyto- and organo-therapeutic ' extracts and mixtures thereof.
  • the enzymes are digestive enzymes selected from: proteases and peptidases, amylases, cellulases, poly- and oligosaccharidases, upases, nucleosidases, phytases.
  • the prokaryotic cells and ' viruses are selected from: yeasts, lactobacilli, probiotic bacteria, antigenic bacteria o viruses, (live, attenuated or killed) .
  • active substances include: enzymes, vaccines, drugs, antibiotics, phyto- and organo- therapeutic extracts and prokaryotic cells and viruse-s or mixtures thereof. Even more preferably, they are enzymes.
  • microparticulate systems are used for the administration, preferably orally, of biologically active substances to animals selected from: porcines, bovines, caprines, ovines, equines, canines, felines, camelids, lagomorphs, rodents, primates and other animal species, such as fish and fowl. Preferred animals are the young of such species.
  • the special composition of such microparticulate systems allows the protection of said biologically active substances from degradation by proteases and gastric acid, allowing their release into the intestine, where they may fulfil their activities.
  • said gastroresistant and enterosoluble polymer is selected from: phthalic acid cellulose esters, (for example: cellulose acetophthalate, hydroxypropyl- methylcellulose phthalate) , trimellitic acid cellulose esters (for example: cellulose trimellitate, hydroxypropylcellulose trimellitate, hydroxypropyl- methylcellulose trimellitate) ; acrylates and polymethacrylates (for example: synthetic anionic and cationic polymers of dimethylaminoethylmethacrylates, of methacrylic acid and ethylmethacrylate) .
  • phthalic acid cellulose esters for example: cellulose acetophthalate, hydroxypropyl- methylcellulose phthalate
  • trimellitic acid cellulose esters for example: cellulose trimellitate, hydroxypropylcellulose trimellitate, hydroxypropyl- methylcellulose trimellitate
  • acrylates and polymethacrylates for example: synthetic anionic and cationic polymers of dimethylaminoethy
  • hydroxypropyl-methylcellulose phthalate synthetic anionic and cationic polymers of methacrylic acid and methylmethacrylate.
  • Said synthetic anionic and cationic polymers of polymethacrylic acid and methylmethacrylate are for example: Eudragit LlOO ® and Eudragit L100-55 ® or Eudragit S100 ® .
  • the cryoprotector or lyoprotector is preferably selected from: glycerol, propanediols, polyethyleneglycols of various molecular weights, sucrose, lactose, mannitol, glycocoll, pentaerythritol, trehalose, sorbitol, xylitol, dimethylsulphoxide, isopropanols, polyvinylpyrrolidones, polyoxyethylenes, hydroxyethylamides, alpha, beta and gamma cyclodextrin and derivatives thereof. More preferably, the cryoprotector or lyoprotector is selected from: sucrose, lactose and mannitol.
  • the divalent or trivalent metal salt of a biocompatible and biodegradable polymer having acidic groups is a calcium, barium, strontium, zinc, aluminium, iron, or chromium salt of alginic acid, hyaluronic acid or xanthan gum.
  • the additional biocompatible and biodegradable polymer is selected from the group consisting of: glucans, scleroglucans, mannans, galactomannans, gellans, carrageenans, pectins, polyanhydrides, polyaminoacids, polyamines, xanthans_,_ tragacanth gum, guar gnm, xanthan gum, celluloses and derivatives thereof, carboxymethylcellulose, ethylcellulose, methylcellulose, hydroxypropylcellulose, hydroxyethylcellulose, hydroxypropylmethylcellulose, polyvinylalcohols, polyoxyethylenes, carboxyvinylpolymers, starches, collagens, chitins and chitosans.
  • the microparticulate systems, described above and obtained by means of the process described below, have dimensions comprised between 30 ⁇ m and 1200 ⁇ m, preferably between 50 ⁇ m and 500 ⁇ m.
  • the present invention relates to a process for the preparation of these gastroresistant microparticulate systems.
  • Said process comprises the following stages: a) combining an aqueous solution of gastroresistant and enterosoluble polymer with an aqueous or physiological solution of cryo- or lyoprotector and the salt of a monovalent metal, prefexably the potassium or sodium salt of a biocompatible and biodegradable polymer, having acidic groups, preferably alginic acid, so as to obtain a colloidal solution or homogeneous suspension; b) solubilising or dispersing the biologically active substance in the solution or suspension from step a); c) nebulising or extruding the solution or suspension from step b) into an aqueous solution of a soluble, inorganic, salt of divalent or trivalent ions.
  • Step a) includes a step al) for the preparation of the monovalent metal salt solution of a biocompatible and biodegradable polymer, having acidic groups and the cryo- or lyoprotector; and a step a2) for the preparation of the gastroresistant and enterosoluble polymer solution.
  • a saline solution (0.5-15% sodium chloride in water, preferably about 0.9% NaCl in water) of the monovalent metal salt of a biocompatible and biodegradable polymer, having acidic groups, in concentration ranging from 0.01% to 10% w/v, preferably from 0.1% to 3% w/v is preferably prepared. This is left stirring using a mechanical s ' tirrer unto the salt is completely dissolved.
  • the monovalent metal salt of a biocompatible ' and biodegradable polymer, having acidic groups is preferably a sodium or potassium salt of alginic acid, hyaluronic acid or xanthan gum, more preferably, it is sodium alginate.
  • an (aqueous or physiological) solution of approx. 2% w/v is prepared, having a viscosity of between 200 cP and 20000 cP at 25°C.
  • the alginate has a mean molecular weight of between 20,000 and 240,000 Daltons .
  • the cryo- or lyoprotector in such amounts as to obtain a concentration of- from 1 to 40% w/v, preferably from 2% to 20% w/v.
  • Said cryo- or lyoprotector is . as described above.
  • an additional biocompatible and biodegradable polymer in such quantities as to obtain a concentration of from 0.1% to 50% w/v, preferably from 2 to 15% w/v.
  • Said biocompatible and biodegradable polymer is as described above .
  • an aqueous solution of the gastroresistant and enterosoluble polymer is prepared with a concentration ranging from 1% to 50% w/v, preferably from 5% to 15% w/v, with a pH from 6 to 8, even better if from 7 to 7.8.
  • the solution obtained in al) and the solution obtained in a2) are combined in ratios of from 3:1 to; 1:3, preferably in a ratio of' 1:1.
  • step b) the biologically active substance is dispersed in the colloidal solution or homogeneous suspension, thus obtained, at concentrations ranging from 0.01% to 20% w/v, preferably from 0.1% to 5% w/v.
  • step c) nebulisation takes place with the aid of orifices, nozzles, or syringes having sizes ranging ' from 10 ⁇ m to 5000 ⁇ m, preferably from 300 ⁇ m to 2000 ⁇ m..
  • Extrusion takes place with the aid of automatic o ⁇ semiautomatic microencapsulators, peristaltic or piston pumps or alternatives, or by means of a syringe, manually and/or automatically driven at such a speed as to produce from 10 to 250 drops/minute, preferably from 20 to 120 drops/minute .
  • Nebulisation or extrusion determines the formation of very small drops which are collected in an aqueous solution of a soluble divalent or trivalent inorganic salt, kept stirring at a speed of between 10 and 200 rpm, preferably between 20 and 100 rpm.
  • the volumetric ' ratio between the extruded solution and the inorganic salt solution is between 1:1 and 1:6,, preferably, the ratio is 1:4.
  • This divalent or trivalent ion inorganic salt is selected from: calcium, barium, strontium, zinc, aluminium, iron or chromium chloride, preferably calcium chloride, barium chloride or aluminium chloride. Even more preferably, it is calcium chloride.
  • the concentration of said inorganic salt solutions is of between 0.1 M and 2.0 M, preferably between 0.2 M and 0.8 M.
  • the presence of a divalent- or trivalent metal salt leads to the formation of a matrix consisting of insoluble salts of the biodegradable and biocompatible polymer, having acidic groups, with the divalent or trivalent metal used, and thus the attainment of rapidly sedimenting microparticulate systems. These microparticulate systems have a spherical shape and are insoluble.
  • microparticulate systems thus obtained may be subjected to outer surface cross- linking, by means of interfacial polymerisation of the biocompatible . and biodegradable polymer divalent .
  • polyamine-type cross-linking agents such as, for example: protamine sulphate or phosphate, poly-L-lysine hydrobromide (molecular weight range from 1,000 Da to 80,0000 Da), polyvinylamine, chitosans (molecular weight range ' from 15,000 Da to 1,000,000 Da).
  • Said cross-linking agents are preferably used as aqueous solutions at concentrations between 0.01% and 5% w/v.
  • the cross-linking reaction is carried out ' at a temperature between 5°C and 40°C, preferably about 25°C for times between 1 minute and 120 minutes, preferably between 3 and 30 minutes.
  • microparticulate systems may be subsequently subjected to lyophilisation, using techniques known to those skilled in the . art,- or dried by means- of any method known in the art which is not prejudicial to the activity of the encapsulated biologically active substance.
  • Said microparticulate systems may be stored at temperatures between -200°C and 40°C, preferably between
  • the present invention relates to a kit for the preparation of capsules, according to the invention, comprising previously prepared, pre-measured and pre-packaged raw materials, as well as any relevant disposable, sterile, non sterile or sterilisable materials .
  • kit will comprise said gastroresistant and enterosoluble polymer, said monovalent metal salt of a biocompatible and biodegradable polymer having acidic groups, said divalent or trivalent inorganic salt, said cryo- or lyoprotector and said biologically active substance in separate pre-measured packages.
  • the kit may contain said additional biodegradable and biocompatible polymer and said cross-linking agent in separate pre-measured packages.
  • the kit will additionally comprise extrusion devices such as sterile, non-sterile or sterilisable nozzles, needles or syringes.
  • the kit may require storage at temperatures -ranging from -5°C to 40°C, preferably about 0°C, in order to avoid deterioration of the active substance, ' and the immediate use and/or- cryo-desiccation ⁇ of the microparticulate systems immediately upon formation.
  • the microparticulate systems forming the subject of the present invention may be administered- orally, by administration with a liquid diet or as supplements in solid feed.
  • the present invention relates to a pre-packaged animal feed, supplemented with the microparticulate system of the invention.
  • gastroresistant and enterosoluble microparticulate systems may also be used for the administration of biologically active substances to humans .
  • biologically active substances is meant those listed previously.
  • This invention provides microparticulate systems having such dimensions as to allow optimal dispersion in solid and liquid foods without any problems involving the particles aggregating and, hence, ' separating from the solid or precipitating out of the liquid. This allows easy administration to humans and animals.
  • the gastroresistant microparticulate systems of the invention may be administered orally and afford, - in gastric acid environments, effective protection of the biologically active substances vehicularised, and the rapid release of the aforesaid substances, with high biological activity, in the enteric environment (small or ' large intestine) .
  • Said gastroresistant microparticulate systems have significant application potential in the sector of veterinary gastroenterology and nutrition, especially in monogastric animal species, but also in polygastric non- ruminants and in those ruminants with still non-functional pre-stomachs . •
  • Such preparations may be classified among the zootechnical feed additives (as described in the 1 st enclosure to Reg. EC N° 1831/2003) .
  • the invention resolves the essential problem of the administration of digestive enzymes (e.g. ⁇ -amylase) in young, where enzyme expression is still not entirely efficient, and in adults with reduced digestive capacity.
  • digestive enzymes e.g. ⁇ -amylase
  • it is ' possible to- supplement the maternal milk diet of the young, with feeds promoting faster growth, thus obtaining improved quality of life and health for the— animals and significant economic advantages for the grower.
  • GASTRORESISTANCE AND BIOLOGICAL ACTIVITY TESTING In order • to assess whether said microparticulate systems are effectively gastroresistant and whether the enzyme or biologically active substance maintains its activity, a sample of the microparticulate systems has been subjected to the assay provided in the Pharmacopoeia (FUI XI) for gastroresistant pharmaceutical, forms.
  • the sample has firstly been incubated at 37 °C in hydrochloric acid at pH 1 for two hours and subsequently transferred to phosphate buffer at pH .6.8 for one hour. Following incubation in hydrochloric acid, the microparticulate systems showed no morphological changes but, after a few minutes of incubation at pH 6.8, they begun to dissolve rapidly, completely solubilising and hence liberating the- vehicularised active substance. The quantity and activity of the remaining active substance has been determined on such solution.
  • Solution A To isotonic saline (physiological solution) (NaCl 0.9% w/v) is added low viscosity sodium alginate (250 cps, 2% solution, 25°C) (alginic acid, sodium • salt, low viscosity, SIGMA Chemical CO, St.- Louis, Missouri,- USA) so- as to obtain a concentration of 5% w/v and it is left stirring with the aid of a magnetic stirrer at -100 rpm at room temperature until the polymer is completely, dissolved. To the resulting solution is then added hydroxypropylmethylcellulose (Methocel E3- Dow Chemical) so as to obtain a concentration of 1% w/v.
  • hydroxypropylmethylcellulose Mehocel E3- Dow Chemical
  • lactose (Carlo Erba, Milan, Italy) is then added to the above solution in such quantities as to give a concentration of 4.5% w/v.
  • Solution B A 10% w/v solution of polymethacrylate (Eudragit S100®, Rohm Pharma, GmbH, Darmstadt, D) in phosphate buffer at pH 7.5 is prepared by stirring at room temperature.
  • Solution B is then added to solution A, whilst kept stirring with the aid of a magnetic stirrer, in a volumetric ratio of 1:1, to give a suspension containing 2.5% sodium alginate, 0.5% hydroxypropylmethylcellulose, 2.25% lactose and 5.0 % polymethacrylate ' .
  • ⁇ -amylase from Hog Pancreas; Sigma, Milan, Italy
  • the resulting suspension is nebulised, by means of a suitable spray system fitted with a 0.5 mm diameter nozzle, into a 0.3 M solution of calcium chloride in physiological solution at 0.9% w/v NaCl, whilst kept stirring at 100 rpm with the aid of a magnetic stirrer.
  • the nebulisation nozzle is positioned approx. 10 cm from the surface of the solution.
  • the volumetric ratio between the nebulised solution and the calcium chloride solution is 1:4; the pressure applied is 4 atm.
  • Microparticulate systems, which sediment rapidly, are obtained: they are separated by removal of the supernatant by means of aspiration, washed twice with physiological solution, filtered and placed in suitable stainless steel containers in order to be subsequently subjected to lyophilisation.
  • the lyophilisation process is carried out in accordance with techniques known to those skilled in the art .
  • the lyophilised product appears as a fine powder, with good properties of flow and flowability.
  • the composition of the lyophilisate calculated from the composition of the nebulised solution, is as follows: ⁇ -amylase: 2.8% Calcium alginate: 23.7% Lactose: 21.3% Hydroxypropylmethylcellulose : 4.7% Polymethacrylate: 47.5%
  • the lyophilisate consists of microparticulate systems, insoluble in water, with normal granulometric distribution and mean diameter of approx. 150 ⁇ m, (Coulter LS230 laser scattering granulometer, Beckman-Coulter Inc., Fullerton, CA, USA) .
  • the powder has good free-flow and wetting properties, and is hence particularly suitable to be added to solid and liquid feeds, in order to obtain homogeneous mixtures or suspensions.
  • the ⁇ -amylase maintains practically unchanged levels of enzymatic activity, being equal to 90%, with respect to the activity of the enzyme prior to encapsulation.
  • the 10% loss of enzymatic activity is due to the process to which the enzyme has been subjected in order to incorporate into the microparticulate systems.
  • the microparticulate systems have subsequently been subjected to the above mentioned assay envisaged in the
  • Solution A To isotonic saline (physiological solution) (NaCl 0.9% w/v) is added low viscosity sodium alginate (250 cps, 2% solution, 25°C) (alginic acid, sodium salt, low viscosity, SIGMA Chemical CO, St. Louis, Missouri, USA) so as to obtain a concentration of 5% w/v and it is left stirring with the aid of a magnetic stirrer at 100 rpm at room temperature until the polymer is completely dissolved. As a cryoprotector, lactose (Carlo Erba, Milan, Italy) is then added to the above solution in such quantities as to give a concentration of 4.5% w/v.
  • Solution B A 10% w/v solution of polymethacrylate (Eudragit
  • resulting suspension is then extruded, in the form of drops, through needles (25Gx5/8") with the aid of a peristaltic pump, into a 0.3 M solution of calcium chloride containing 0.9% w/v sodium chloride, kept stirring with the aid of a magnetic stirrer at 100 rpm.
  • the extrusion, needle is positioned approx. 10 cm from the surface of the solution.
  • the volumetric ratio between the extruded solution and the calcium chloride solution is 1:4; the speed of. the peristaltic pump is such so as to produce 60 drops of extrudate/min.
  • Microparticulate systems, which sediment rapidly, are thus obtained: they are separated by removal of the supernatant by means of aspiration, .
  • the lyophilisation process is carried out in accordance with techniques known to those skilled in the art.
  • the lyophilised product appears as an easy flowing, fine powder.
  • the composition of the lyophilisate calculated from the composition of the extruded solution, is as follows: ⁇ -amylase: 3.0% Calcium alginate: 24.8% Lactose: 22.3% Eudragit S100 ® : 49.9%
  • the lyophilisate is constituted by microparticulate systems with diameters of approx. 500 ⁇ m (Coulter LS230 laser scattering granulometer, Beckman-Coulter Inc.,
  • the ⁇ -amylase maintains practically unchanged levels of enzymatic activity, being equal to 90%, with respect to the activity of the enzyme prior to encapsulation.
  • the 10% loss of enzymatic activity is due to the process to which the enzyme has been subjected in order to incorporate into the microparticulate systems.
  • the microparticulate systems have subsequently been subjected to the assay provided in the Pharmacopoeia (FUI XI) for the above mentioned gastroresistant pharmaceutical forms, so as to assess the enzyme's in vitro stability.
  • Mean ⁇ -amylase enzyme activity has been evaluated as being equal to 29.5%.
  • EXAMPLE 3 PREPARATION OF LACTATE DEHYDROGENASE (LDH) CONTAINING, MICROPARTICULATE SYSTEMS BY NEBULISATION.
  • LDH LACTATE DEHYDROGENASE
  • MICROPARTICULATE SYSTEMS BY NEBULISATION PREPARATION OF LACTATE DEHYDROGENASE (LDH) CONTAINING, MICROPARTICULATE SYSTEMS BY NEBULISATION.
  • Solution A To isotonic saline (physiological solution) (NaCl 0.9% w/v) is added low viscosity sodium alginate (250 cps, 2% solution, 25°C) (alginic acid, sodium salt, low viscosity, SIGMA Chemical CO, St.
  • Solution B is added to solution A, kept stirring with the aid of a magnetic stirrer, in a volumetric ratio of 1:1, to • give a suspension containing.2.5% sodium alginate, 2.25% lactose and 5.0% Eudragit S100®.
  • LDH in the form of a fine, easily soluble powder, ' is added to this , suspension ' in such quantities as to give a concentration of 0.3% w/v.
  • the resulting suspension is nebulised by means of a spray system fitted with a 0.5 mm diameter needle into a 0.3 M solution of calcium chloride in 0.9% w/v sodium chloride, kept stirring with the aid of a magnetic stirrer at 100 rpm.
  • the nebulisation nozzle is positioned approx. 10 cm from ' the surface of the solution.
  • the volumetric ratio between the nebulised solution and the calcium chloride solution is 1:4; the pressure applied is 4 atm.
  • Microparticulate systems, which sediment rapidly, are obtained: they are separated by removal of the supernatant by means of aspiration, washed twice with physiological solution, filtered and placed in suitable stainless steel containers in order to be subsequently subjected to lyophilisation.
  • the lyophilisation process is carried out according to techniques known to those skilled in the art.
  • the lyophilised product has good free-flowing and wettability properties.
  • the composition of the lyophilisate calculated from the composition of the nebulised solution, is as follows: Lactate dehydrogenase: 3.0% Calcium alginate: 24.8% Lactose: 22.3% Eudragit. S100 ® : 49.9%
  • the lyophilisate consists of microparticulate systems with normal granulometric distribution and mean diameter of 130 ⁇ m, (Coulter LS230 laser scattering granulometer,
  • the powder has good . free-flow and wettability properties, and is hence particularly suited to being mixed with other excipients necessary for the formulation of solid pharmaceutical forms, as well as to being used as the dispersed phase in the formulation of suspensions or to being added to solid or liquid feeds.
  • the LDH maintains practically unchanged levels of enzymatic activity, being equal to 90%, with respect to the activity of the enzyme prior to encapsulation-.
  • the 10% loss of enzymatic activity is due to the process to .which the enzyme has been subjected in order to incorporate into the microparticulate systems.
  • the microparticulate systems have subsequently been subjected to the above mentioned assay envisaged in the Pharmacopoeia (FUI- XI) for the ' gastroresistant pharmaceutical forms, so as to assess the enzyme's in vitro stability.
  • Mean LDH enzyme activity has been evaluated as being equal to 75%.

Landscapes

  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Epidemiology (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Medicinal Chemistry (AREA)
  • Chemical & Material Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Biophysics (AREA)
  • Molecular Biology (AREA)
  • Medicinal Preparation (AREA)

Abstract

La présente invention concerne des systèmes de microparticules constitués d'une matrice polymère gastrorésistante, biocompatible et biodégradable contenant un polymère gastrorésistant et entérosoluble, un cryoprotecteur ou un lyoprotecteur, un sel de métal divalent ou trivalent d'un polymère biocompatible et biodégradable présentant des groupes acides et des substances bioactives. Lesdits systèmes de microparticules gastrorésistants sont utilisés pour l'administration, de préférence par voie orale, de substances bioactives à des espèces animales. La composition spéciale de ces systèmes à microparticules permet la protection desdites substances bioactives contre la dégradation par les protéases et l'acide gastrique, permettant leur libération dans l'intestin où leur action peut avoir lieu.
EP05760593A 2004-06-22 2005-06-20 Systemes de microparticules pour l'administration par voie orale de substances bioactives Withdrawn EP1778192A2 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
ITMI20041255 ITMI20041255A1 (it) 2004-06-22 2004-06-22 Sistemi microparticellari per somministrazione orale di sostanze biologicamente attive
PCT/IT2005/000353 WO2005123034A2 (fr) 2004-06-22 2005-06-20 Systemes de microparticules pour l'administration par voie orale de substances bioactives

Publications (1)

Publication Number Publication Date
EP1778192A2 true EP1778192A2 (fr) 2007-05-02

Family

ID=35510243

Family Applications (1)

Application Number Title Priority Date Filing Date
EP05760593A Withdrawn EP1778192A2 (fr) 2004-06-22 2005-06-20 Systemes de microparticules pour l'administration par voie orale de substances bioactives

Country Status (3)

Country Link
EP (1) EP1778192A2 (fr)
IT (1) ITMI20041255A1 (fr)
WO (1) WO2005123034A2 (fr)

Families Citing this family (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN105147626A (zh) * 2007-12-21 2015-12-16 Cnj控股有限公司 含海藻糖的稳定化因子ix制剂
JP5724108B2 (ja) 2009-04-22 2015-05-27 メドスキン ソリューションズ ドクター ズベラック アーゲーＭｅｄＳｋｉｎ Ｓｏｌｕｔｉｏｎｓ Ｄｒ．Ｓｕｗｅｌａｃｋ ＡＧ 凍結乾燥組成物
PL2675287T3 (pl) 2011-02-18 2017-01-31 Dupont Nutrition Biosciences Aps Kompozycja dodatku paszowego
GB201102857D0 (en) 2011-02-18 2011-04-06 Danisco Feed additive composition
GB201102865D0 (en) 2011-02-18 2011-04-06 Danisco Feed additive composition
GB201213801D0 (en) 2012-08-03 2012-09-12 Dupont Nutrition Biosci Aps Feed additive composition
CN104735999A (zh) 2012-08-03 2015-06-24 杜邦营养生物科学有限公司 饲料添加剂组合物
US20160243154A1 (en) * 2013-10-23 2016-08-25 Donald W. Jessup Hyaluronic acid formulation
US20200281225A1 (en) 2015-11-09 2020-09-10 Dupont Nutrition Biosciences Aps Feed additive composition
CN108472351A (zh) 2015-12-29 2018-08-31 英特维特国际股份有限公司 球虫病疫苗
CN113115952A (zh) * 2021-05-27 2021-07-16 江苏恒康生物科技有限公司 一种含微晶球的耐胃酸爆珠及其制备方法
US11833224B1 (en) 2023-02-08 2023-12-05 Leuvian Llc Lyoprotectant compositions and uses thereof

Family Cites Families (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR960011236B1 (ko) * 1987-05-08 1996-08-21 스미스 클라인 앤드 프렌취 라보라토리스 리미티드 제약학적 조성물 및 고체 제형
US5230901A (en) * 1988-03-23 1993-07-27 Knoll Ag Sustained release tablet of a mixture of alginates and polyacrylates
US5876754A (en) * 1992-01-17 1999-03-02 Alfatec-Pharma Gmbh Solid bodies containing active substances and a structure consisting of hydrophilic macromolecules, plus a method of producing such bodies
US5695781A (en) * 1995-03-01 1997-12-09 Hallmark Pharmaceuticals, Inc. Sustained release formulation containing three different types of polymers
US6251430B1 (en) * 1998-02-04 2001-06-26 Guohua Zhang Water insoluble polymer based sustained release formulation

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO2005123034A3 *

Also Published As

Publication number Publication date
ITMI20041255A1 (it) 2004-09-22
WO2005123034A8 (fr) 2006-03-30
WO2005123034A3 (fr) 2006-09-14
WO2005123034A2 (fr) 2005-12-29

Similar Documents

Publication Publication Date Title
WO2005123034A2 (fr) Systemes de microparticules pour l'administration par voie orale de substances bioactives
JP4463109B2 (ja) 栄養補助食品および水生動物の給餌方法
US20190328670A1 (en) Microparticles for Oral Delivery
US20090053317A1 (en) Microparticulate systems for the oral administration of biologically active substances
DK2665377T3 (en) MICROCRAINED PROBIOTIC SUBSTANCE AND PROCEDURE FOR PREPARATION
JP6081374B2 (ja) 酸性劣化からの微生物細胞の保護
Masoomi Dezfooli et al. Encapsulation for delivering bioactives in aquaculture
US5597564A (en) Method of administering a microgranular preparation to the intestinal region of animals
Ray Potential aspects of chitosan as pharmaceutical excipient
US20120301546A1 (en) Acid-resistant soft gel compositions
AU609419B2 (en) Microgranular preparation useful in the delivery of biologically active materials to the intestinal regions of animals
WO2002000035A1 (fr) Complexe alimentaire bioactif, procede servant a preparer un complexe alimentaire bioactif et procede servant a lutter contre les maladies
TWI584824B (zh) A capsule for delivery to a large intestine and a method of manufacturing the same
US20080138419A1 (en) Method for preparing an orally administrable formulation for controlled release
JPWO2006016595A1 (ja) ラクトフェリン複合体及びその製造方法
WO2002085415A1 (fr) Produits enrobes de sucres non digestibles et procede associe
RU2688136C1 (ru) Композиция для пероральной доставки биоактивных агентов
CN109452652A (zh) 耐胃微胶囊及其在刺激哺乳动物内回肠glp-1释放的用途
JP5779762B2 (ja) 経口送達のための生細胞の配合物
WO2005025609A1 (fr) Composition d'un produit a la lactoferrine
WO1995018602A1 (fr) FORME PHARMACEUTIQUE DE DESMOPRECYNE (dDAVP) POUR ADMINISTRATION PAR VOIE ORALE
US20050084519A1 (en) Feed for fish and shellfish, additive therefor, and method for producing additive
Chomová et al. Development and evaluation of a fish feed mixture containing the probiotic Lactiplantibacillus plantarum prepared using an innovative pellet coating method
WO1997040702A1 (fr) Composition pour administration orale presentant une haute efficacite d'absorption et procede de preparation de ladite composition
JPH119196A (ja) キトサンによる家畜の損耗防止の方法

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: 20070119

AK Designated contracting states

Kind code of ref document: A2

Designated state(s): AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HU IE IS IT LI LT LU MC NL PL PT RO SE SI SK TR

RAP1 Party data changed (applicant data changed or rights of an application transferred)

Owner name: UNIVERSITA'DEGLI STUDI DI MILANO

DAX Request for extension of the european patent (deleted)
17Q First examination report despatched

Effective date: 20090702

GRAP Despatch of communication of intention to grant a patent

Free format text: ORIGINAL CODE: EPIDOSNIGR1

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: 20120619