Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K49/00—Preparations for testing in vivo
  • A61K49/001—Preparation for luminescence or biological staining
  • A61K49/0013—Luminescence
  • A61K49/0017—Fluorescence in vivo
  • A61K49/0019—Fluorescence in vivo characterised by the fluorescent group, e.g. oligomeric, polymeric or dendritic molecules
  • A61K49/0021—Fluorescence in vivo characterised by the fluorescent group, e.g. oligomeric, polymeric or dendritic molecules the fluorescent group being a small organic molecule
  • A61K49/0041—Xanthene dyes, used in vivo, e.g. administered to a mice, e.g. rhodamines, rose Bengal
  • A61K49/0043—Fluorescein, used in vivo
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K49/00—Preparations for testing in vivo
  • A61K49/001—Preparation for luminescence or biological staining
  • A61K49/0063—Preparation for luminescence or biological staining characterised by a special physical or galenical form, e.g. emulsions, microspheres
  • A61K49/0069—Preparation for luminescence or biological staining characterised by a special physical or galenical form, e.g. emulsions, microspheres the agent being in a particular physical galenical form
  • A61K49/0076—Preparation for luminescence or biological staining characterised by a special physical or galenical form, e.g. emulsions, microspheres the agent being in a particular physical galenical form dispersion, suspension, e.g. particles in a liquid, colloid, emulsion
  • A61K49/0084—Preparation for luminescence or biological staining characterised by a special physical or galenical form, e.g. emulsions, microspheres the agent being in a particular physical galenical form dispersion, suspension, e.g. particles in a liquid, colloid, emulsion liposome, i.e. bilayered vesicular structure
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K49/00—Preparations for testing in vivo
  • A61K49/22—Echographic preparations; Ultrasound imaging preparations ; Optoacoustic imaging preparations
  • A61K49/222—Echographic preparations; Ultrasound imaging preparations ; Optoacoustic imaging preparations characterised by a special physical form, e.g. emulsions, liposomes
  • A61K49/227—Liposomes, lipoprotein vesicles, e.g. LDL or HDL lipoproteins, micelles, e.g. phospholipidic or polymeric
• • 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
  • A61K9/10—Dispersions; Emulsions
  • A61K9/127—Synthetic bilayered vehicles, e.g. liposomes or liposomes with cholesterol as the only non-phosphatidyl surfactant
  • A61K9/1277—Preparation processes; Proliposomes
• • 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
  • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
  • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
  • A61K9/5005—Wall or coating material
  • A61K9/5063—Compounds of unknown constitution, e.g. material from plants or animals
  • A61K9/5068—Cell membranes or bacterial membranes enclosing drugs

Definitions

• the invention relates to the subject-matter mentioned in the claims, i.e. a process for the production of liposomes and / or biological cells which absorb substances added externally during production. In both cases, substance absorption takes place under gentle conditions.
• the loaded liposomes and or cells are suitable as targeted Phaima carriers.
• Liposomes are made up of lipid molecules, preferably phospholipids. These spontaneously form lipid bilayers in the aqueous medium, which are closed and lamellar. One or more lamellae separate the inner compartment from the outer solution.
• the polar hydrophilic head groups of the lipids e.g. ethanolamine for cephalin, choline for lecithin
• the bilayers are two molecular layers or 4 to 5 ⁇ m thick.
• SUV small unilamellar vesicles
• LUV large unilamellar vesicles
• MLV multilamellar liposomes
• phospholipids accumulate without assistance to form multilamellar liposomes, in which onion-like several lipid bilayers are separated by water layers.
• the formation of lamellar lipid bilayers is a result of the balance of physical forces and steric effects.
• the lipid bilayer also forms the formation block of the plasma membranes, which envelop biological cells and in which proteins are embedded in the lipid matrix as a second building block.
• liposomes The physical and physiological properties of liposomes are determined by their lipid composition, size, temperature, pH, type and strength of the buffer, production conditions, history and age. The use of liposomes that have been loaded with pharmaceuticals has found great interest in diagnostics and therapy. As targeted carriers of pharmaceuticals, liposomes open up new diagnostic and therapeutic forms (see eg R_ Lawaczeck “Liposomes as targeted pharmaceutical carriers", Deutsche maschiner Science 127, 1771-1773 (1987), MJ. Ostro “Liposomes", Sei. Am 256, 90-99 (1987)). The permeability of the lipid membrane and the stability of the liposomes are of interest for pharmaceutical purposes; physiologically, the fusogenicity and the kinetics of cell uptake are also important.
• the gases used must be chemically inert with regard to the lipids and the water phase and not interact with water like carbon dioxide or ammonia.
• the use of easily manageable, technically available gases which leave no toxic residue in a pressure range up to 1000 bar, preferably up to 200 bar, are suitable. It may be advantageous to exceed the boiling curve in the p, T diagram in the liquid state.
• Suitable gases are preferably nitrous oxide (N2O), halothane, argon, xenon, sulfur hexafluoride (SFß), alkanes such as methane, ethane, propane, butane, hexane, heptane, alkenes such as ethene, propene, butene, hexene, heptene.
• N2O nitrous oxide
• halothane halothane
• argon argon
• xenon sulfur hexafluoride
• SFß sulfur hexafluoride
• alkanes such as methane, ethane, propane, butane, hexane, heptane
• alkenes such as ethene, propene, butene, hexene, heptene.
• a pressure of up to 55 bar is selected as it is available in technical pressure bottles.
• the gas molecules mentioned exercise a "detergent-like" one "Effect and lead to an opening and / or solubilization of the lipid bilayer and / or membrane.
• new liposomes spontaneously form or close the membranes again, so that the added substances are taken up. This process can be repeated cyclically.
• liposomes it is possible to narrow the liposome size by depressurizing to atmospheric pressure using size-selective separation filters.
• the lipid composition of the liposomes can be optimized depending on the therapeutic and / or diagnostic use and on the destination.
• the biological cells in addition to the cells of the bloodstream such as erythrocytes, leukocytes, lymphocytes and thrombocytes, cells which are kept in culture are preferred.
• the process is technically easy to implement and inexpensive.
• the method is suitable for liposomal and cellular addition, incorporation and / or substitution of hydro- and lipophilic substances.
• the loaded liposomes can be surface-modified and used as target or organ-specific pharmaceutical carriers or pharmaceutical stores. Examples
• Example 1 20 mg of dimyristoyl lecithin are taken up in 1 ml of chloroform. The solution is placed in the pressure vessel. The chloroform is filled with purified nitrogen and subsequently evaporated under vacuum. 2 ml of water containing 1 mM carboxyfluorescein (purified and recrystallized from ethanol), 20 mM Tris HC1 are adjusted to pH 8. CO2 and O2 are expelled by flushing with laughing gas. The pressure vessel is closed and a nitrous oxide pressure of 50 bar is applied at room temperature. The solution is shaken for 0.5 hours, then is slowly and isothermally depressurized to atmospheric pressure. The process is repeated 5 times. The solution is removed from the pressure vessel and passed through an anion exchanger to remove the externally present caiboxyfluorescein. The Iiposome fraction contains the included dye and can be used for fluorescence studies.
• Example 2 as example 1, 200 mg of a 4: 1 mixture of egg yolk lecithin and cholesterol are used instead of dimyristoyl lecithin. Carboxyfluorescein is replaced by 20 mM gadolinium DTPA as an MR contrast medium. The liposomes obtained can be used in MR diagnostics.
• Example 3 Preparation of erythrocytes or erythrocyte ghosts loaded with carboxyfluorescein under blood isotonic conditions.
• Fresh bovine blood is washed 3 times in PBS buffer pH 7.4 and freed of other components except for the erythrocytes.
• the dark red, cloudy erythrocyte suspension is diluted with PBS buffer containing carboxyfluorescein so that the concentration of carboxyfluorescein is 10-5 M.
• the solution is placed in a pressure measuring cell. After rinsing with N2O, 50 bar of N2O are stirred at 37 ° C. and the optical density at 675 nm is followed. Lysis of the cells occurs between 400 and 600 min, which is visible through the typically sigmoidal drop in optical density.
• Example 4 As example 3, but sheep erythrocytes are used instead of the bovine erythrocytes in 2 mM KH2PO4, 9 mM Na2HP04, 3 mM KCl, 137 mM NaCl pH 7.5. Lysis occurs between 55 to 70 min. The results are comparable to those from example 3.

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• Health & Medical Sciences (AREA)
• Life Sciences & Earth Sciences (AREA)
• Public Health (AREA)
• Epidemiology (AREA)
• Animal Behavior & Ethology (AREA)
• General Health & Medical Sciences (AREA)
• Veterinary Medicine (AREA)
• Engineering & Computer Science (AREA)
• Chemical & Material Sciences (AREA)
• Biomedical Technology (AREA)
• Dispersion Chemistry (AREA)
• Medicinal Chemistry (AREA)
• Bioinformatics & Cheminformatics (AREA)
• Pharmacology & Pharmacy (AREA)
• Acoustics & Sound (AREA)
• Radiology & Medical Imaging (AREA)
• Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
• Physics & Mathematics (AREA)
• Biophysics (AREA)
• Cell Biology (AREA)
• Molecular Biology (AREA)
• Virology (AREA)
• Botany (AREA)
• Zoology (AREA)
• Medicinal Preparation (AREA)
• Manufacturing Of Micro-Capsules (AREA)
• Micro-Organisms Or Cultivation Processes Thereof (AREA)

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Cited By (14)

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Citations (5)

• 1988
  • 1988-04-16 DE DE3812816A patent/DE3812816A1/de not_active Withdrawn
• 1989
  • 1989-04-13 WO PCT/DE1989/000230 patent/WO1989009593A1/de not_active Ceased
  • 1989-04-13 JP JP1504065A patent/JPH03505201A/ja active Pending
  • 1989-04-13 DE DE8989730103T patent/DE58903315D1/de not_active Expired - Fee Related
  • 1989-04-13 AT AT89730103T patent/ATE84709T1/de not_active IP Right Cessation
  • 1989-04-13 AU AU34168/89A patent/AU634470B2/en not_active Ceased
  • 1989-04-13 ES ES198989730103T patent/ES2039929T3/es not_active Expired - Lifetime
  • 1989-04-13 EP EP89730103A patent/EP0338971B1/de not_active Expired - Lifetime
• 1990
  • 1990-10-01 DK DK236790A patent/DK236790D0/da not_active Application Discontinuation
  • 1990-10-15 NO NO904456A patent/NO178653C/no unknown
• 1993
  • 1993-02-23 GR GR930400364T patent/GR3007128T3/el unknown

Patent Citations (5)

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