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EP0665756A1 - Procede et dispositif de preparation de systemes disperses liquides - Google Patents

Procede et dispositif de preparation de systemes disperses liquides

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Publication number
EP0665756A1
EP0665756A1 EP93922883A EP93922883A EP0665756A1 EP 0665756 A1 EP0665756 A1 EP 0665756A1 EP 93922883 A EP93922883 A EP 93922883A EP 93922883 A EP93922883 A EP 93922883A EP 0665756 A1 EP0665756 A1 EP 0665756A1
Authority
EP
European Patent Office
Prior art keywords
production
liquid
systems according
disperse systems
disperse
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.)
Ceased
Application number
EP93922883A
Other languages
German (de)
English (en)
Inventor
Andreas Sachse
Thomas Schneider
Georg Rössling
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Individual
Original Assignee
Individual
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from DE19924235381 external-priority patent/DE4235381A1/de
Priority claimed from DE19934328331 external-priority patent/DE4328331A1/de
Application filed by Individual filed Critical Individual
Publication of EP0665756A1 publication Critical patent/EP0665756A1/fr
Ceased legal-status Critical Current

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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
    • A61K9/10—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
    • A61K49/00—Preparations for testing in vivo
    • A61K49/04—X-ray contrast preparations
    • A61K49/0433—X-ray contrast preparations containing an organic halogenated X-ray contrast-enhancing agent
    • A61K49/0447—Physical forms of mixtures of two different X-ray contrast-enhancing agents, containing at least one X-ray contrast-enhancing agent which is a halogenated organic compound
    • A61K49/0461—Dispersions, colloids, emulsions or suspensions
    • A61K49/0466—Liposomes, lipoprotein vesicles, e.g. HDL or LDL lipoproteins, phospholipidic or polymeric micelles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K49/00—Preparations for testing in vivo
    • A61K49/06—Nuclear magnetic resonance [NMR] contrast preparations; Magnetic resonance imaging [MRI] contrast preparations
    • A61K49/18—Nuclear magnetic resonance [NMR] contrast preparations; Magnetic resonance imaging [MRI] contrast preparations characterised by a special physical form, e.g. emulsions, microcapsules, liposomes
    • A61K49/1806—Suspensions, emulsions, colloids, dispersions
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K49/00—Preparations for testing in vivo
    • A61K49/06—Nuclear magnetic resonance [NMR] contrast preparations; Magnetic resonance imaging [MRI] contrast preparations
    • A61K49/18—Nuclear magnetic resonance [NMR] contrast preparations; Magnetic resonance imaging [MRI] contrast preparations characterised by a special physical form, e.g. emulsions, microcapsules, liposomes
    • A61K49/1806—Suspensions, emulsions, colloids, dispersions
    • A61K49/1812—Suspensions, emulsions, colloids, dispersions liposomes, polymersomes, e.g. immunoliposomes
    • 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
    • A61K9/107—Emulsions ; Emulsion preconcentrates; Micelles
    • A61K9/1075—Microemulsions or submicron emulsions; Preconcentrates or solids thereof; Micelles, e.g. made of phospholipids or block copolymers
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S977/00—Nanotechnology
    • Y10S977/70—Nanostructure
    • Y10S977/788—Of specified organic or carbon-based composition
    • Y10S977/797—Lipid particle
    • Y10S977/798—Lipid particle having internalized material
    • Y10S977/799—Containing biological material
    • Y10S977/801—Drug
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S977/00—Nanotechnology
    • Y10S977/902—Specified use of nanostructure
    • Y10S977/904—Specified use of nanostructure for medical, immunological, body treatment, or diagnosis
    • Y10S977/906—Drug delivery
    • Y10S977/907—Liposome
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S977/00—Nanotechnology
    • Y10S977/902—Specified use of nanostructure
    • Y10S977/904—Specified use of nanostructure for medical, immunological, body treatment, or diagnosis
    • Y10S977/927—Diagnostic contrast agent
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S977/00—Nanotechnology
    • Y10S977/902—Specified use of nanostructure
    • Y10S977/904—Specified use of nanostructure for medical, immunological, body treatment, or diagnosis
    • Y10S977/927—Diagnostic contrast agent
    • Y10S977/928—X-ray agent
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S977/00—Nanotechnology
    • Y10S977/902—Specified use of nanostructure
    • Y10S977/904—Specified use of nanostructure for medical, immunological, body treatment, or diagnosis
    • Y10S977/927—Diagnostic contrast agent
    • Y10S977/929—Ultrasound contrast agent

Definitions

  • the invention relates to a method and a device for producing liquid, disperse systems.
  • lipid vesicles Due to the hydrophobic interactions, spontaneously closed lipid vesicles, which are called liposomes, are formed after dispersion of phospholipids in water. These are spherical or elliptical hollow bodies with one or more lipid bilayers, which include an aqueous phase. According to their size, a distinction is made between small, unilamellar (small unilamellar vesicles [SUV] with radii from 25 to 50 nm) and large, unilamellar vesicles (large unilamellar vesicles [LUV] with radii greater than 50 nm up to 10 ⁇ m) (Weiner, N ., Martin, F., Riaz, M., Drug Dev. Ind. Pharm. 15_, 1523-1554 (1989)).
  • SUV small unilamellar vesicles
  • LUV large unilamellar vesicles
  • multilamellar liposomes multilamellar vesicles [MLV]
  • MLV multilamellar vesicles
  • MW multivesicular liposomes
  • the liposomes are suitable for the inclusion of both hydrophilic and lipophilic drugs, the extent and location of the inclusion depending on the physicochemical properties of the drug and the lipid composition of the liposomes.
  • lipid film is first formed by rotary evaporation of an organic solution of the lipid or lipid mixture (for example in chloroform, methanol or diethyl ether). To completely remove the residual solvent, a 12-24 hour lyophilization is then often carried out under high vacuum. Subsequent addition of an aqueous phase and simple shaking (the so-called hand-shake method according to Bangham, Bangham, AD, Standish, MM, Watkins, J.C, J. Mol. Biol. 12, 238-252 (1965)) gives an MLV suspension that is extremely heterogeneous in terms of liposome size and lamellarity.
  • the oldest and most widespread method of producing SUVs is the so-called "sonication method" (ultra-sonication method).
  • MLV are crushed by ultrasound (ultrasonic wand or ultrasonic bath).
  • the liposomes thus obtained have an average diameter of 20 to 60 nm and an inclusion capacity of less than 1%.
  • the disadvantages of this method lie above all in the high supply of heat, which can lead to decomposition of the lipid or the pharmaceutical substance, and in the difficulty in reproducibly processing even large amounts of samples.
  • the ultrasound rod there is also the disadvantage of contamination of the sample with titanium chips and the formation of an aerosol (see the already cited publication by R.R.C. New).
  • the main disadvantage is the occurrence of device wear (annular gap etc.) and the difficult control of the product temperature.
  • Microfluidizer TM Mayhew, E., Lazo, R., Vail, WJ, King, J., Green, AM, Biochim. Biophys. Acta 775, 169-174 (1984) ).
  • an MLV dispersion or a coarse, aqueous lipid dispersion is first introduced into a reservoir and pressed by means of a high-pressure pump via a pre-filter (5 ⁇ m) into a so-called interaction chamber, in which the liquid flow in micro-channels is split into two individual flows, which are then split up be reunited at high speed. After exiting the interaction chamber, the dispersion obtained can either be removed or recirculated.
  • the continuous process according to the invention for the production of liquid dispersions does not have the disadvantages of the previously known processes. It is characterized in that a predispersion under high pressure of 6.6 to 250 MPa is sequentially extruded over 1 to 8 filter stages of decreasing pore size between 0.01 and 35 ⁇ m, it being possible to use up to 20 passages per filtration stage.
  • the process according to the invention is preferably carried out at a working pressure of 7 to 80 MPa.
  • the extrusion can take place at each filtration stage using one or a combination of 2-4 filters of the same or different pore size.
  • membrane filters such as polycarbonate membranes (surface filters) from Nucleopore (Tübingen) are used.
  • filters are metal or polymer membranes or inorganic materials such as glass fiber or Anopore R membranes (Anotec, Banbury Oxon, England).
  • suitable polymer materials are filters made of polytetrafluoroethylene (PTFE), polypropylene (PP), polyvinylidene fluoride or cellulose esters such as cellulose acetate.
  • This device which can be referred to as a continuously operating high-pressure extrusion apparatus (see Fig. 1), is characterized by a storage vessel (1), the drain line of which leads to a high-pressure pump (2) which is designed to build up working pressures of a maximum of 250 MPa. On the output side, this pump is connected to a filter holder (6) intended for receiving the filters used according to the invention, from which the product is connected
  • the high-pressure pump is preferably designed to build up working pressures of up to 80 MPa. Between the high-pressure pump and the filter holder, a pre-filter holder can also be attached, which is designed to hold filters with an average pore size of 2 to 35 ⁇ m. Furthermore, the device according to the invention can also be equipped with ventilation devices and / or temperature and pressure measuring devices.
  • Suitable pumps for the device according to the invention are, for example, pneumatic or hydraulic piston pumps (for example Maximator®, Schmidt, Kranz & Co., Zorge, Germany). Suitable pumps are usually those that have a span factor of around 50 to 750 and can therefore generate working pressures between 5 and 300 MPa from 0.1 to 0.4 MPa inlet pressure (air or nitrogen).
  • the devices according to the invention are expediently provided with a control valve (3), by means of which the pressure to be used in the method according to the invention can be set in a targeted manner.
  • resulting product flows are usually between 0.1 and 10 liters per minute, preferably 0.15 to 3 liters per minute.
  • the high pressures which can be used in the method according to the invention also eliminate the problem of filter clogging which occurs in other methods, as a result of which interruptions in the process for changing the filter are eliminated.
  • the storage vessel used in the device according to the invention can be designed in such a way that it can be temperature-controlled; the lines can be metal tubes or hose lines.
  • the product can also be returned via a two-chamber storage container or a liquid spiral, optionally also with heat exchange.
  • the device is to be used for the production of pharmaceutical preparations, all parts of the product in contact with it must be sterilizable and resistant to the solvents used in it.
  • the device is preferably made of materials which permit heat sterilization.
  • the high-pressure extrusion apparatus enables liquid dispersions to be produced continuously and in large quantities, as a result of which the production outlay for the dispersions is significantly reduced and the economy of the process is thus significantly improved.
  • the apparatus used in the exemplary embodiments has filter holders which allow the use of membrane filters with a diameter of 47 mm.
  • the pressure holding capacity of the filter holder used here is 80 MPa.
  • This device allows the rapid production of dispersions in the range of 100 to 1000 ml (dead volume of the system about 10 ml).
  • flows can be achieved with this device that are well above 150 ml per minute.
  • the dispersion is initially collected in a two-chamber system, in order to then return it to the system by simply opening a corresponding valve.
  • This two-chamber system offers the advantage over direct recirculation, since the total amount of dispersion is always subjected to the shear process and there is no mixing of the undispersed and dispersed phase.
  • the same effect can also take place via the recirculation of the dispersion via a liquid spiral, which comprises the total volume of the batch to be processed.
  • the dispersion can be tempered via the outer walls of the corresponding spiral.
  • the device according to the invention is not only suitable for carrying out the method according to the invention, but can also be useful when using lower pressures in the range from 1 to 6.6 MPa.
  • the method according to the invention allows the production of uni- or multilamellar liposomes over a wide limit range (average diameter usually 25 nm to 5 ⁇ m).
  • the size and homogeneity of the size distribution and the lamellarity of the liposomes obtained here are, among other things, a function of the type of filter used and pore size, the filtration pressure (working pressure), the number of passages through the device, the type and concentration of lipid and the type and amount of used drug.
  • lipid constituents can be used as in the other methods of this type.
  • lipids are generally phospholipids such as, for example, phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylserine, phosphatidic acid, phosphatidylinositol or sphingolipids.
  • sterols such as choesterol or other components such as fatty acids (e.g. stearic acid, palmitic acid), dicetyl phosphate or cholesterol hemisuccinate can be used as further constituents.
  • amphiphilic substances such as hexadecylpoly (3) glycerol, dialkylpoly (7) glycerol ether and alkyl glucosides, so-called niosomes, ie liposomes from non-ionic vesicle formers, are obtained.
  • Suitable active ingredients are, for example, vitamins, hormones, antimycotics, antiallergics, antphlogistics, antihypertensives, antiarrhythmics, antibiotics, antivirals, anxiolytics, cytostatics, immunomodulators, contraceptives, peptides, proteins and sedatives.
  • hydrophilic drugs these are generally dissolved in the aqueous phase used to prepare the predispersion and, after the predispersion has been prepared, are subjected to the process according to the invention. Surprisingly, liposomes with particularly high inclusions can be obtained here.
  • the method according to the invention thus proves to be particularly suitable, inter alia, for the encapsulation of contrast media for X-rays (or computed tomography) and NMR diagnostics, which have been inadequately encapsulated using the mechanical dispersion methods known to date.
  • contrast media for X-rays or computed tomography
  • NMR diagnostics which have been inadequately encapsulated using the mechanical dispersion methods known to date.
  • RKM iodine-containing X-ray contrast media
  • high inclusion capacities can be achieved even with small liposome diameters and relatively low lipid concentrations.
  • Combining it with one or more freeze-thaw cycles (freezing and thawing) can further increase inclusion.
  • RKM of the triiodobenzoic acid type are lopromide, lohexol, lopamidol, loversol, lopentol, loxaglate, 3-carbamoyl-5- [N- (2-hydroxyethyl) -acetamido] -2,4,6-triiodobenzoic acid - [(1 RS, 2SR) - 2,3-dihydroxy-1-hydroxymethylpropyl] amide and lotrolan.
  • NMR contrast agents which are particularly suitable for encapsulation are Gd-DTPA, Gd-EOB-DTPA, Gd-BOPTA, Gd-DOTA, Gadobutrol and Mn-DPDP (US Pat. Nos. 4,957,939, 5,021,236 and Schuhmann-Giampieri, G., Inv. Radiol. 28, (1993) in press).
  • suitable water-soluble substances can also be encapsulated using so-called active ioading techniques (remote loading).
  • active ioading techniques for example, drug-free liposomes are first produced using the high-pressure extrusion technology, which then, e.g. via a pH gradient, with which the substance to be encapsulated is loaded (Cullis, PR, Mayer, L D., Bally, MB, Madden, TD, Hope, MJ, Adv. Drug Delivery Rev. 3, 267-282 (1989) ).
  • the corresponding active ingredient can be encapsulated in the process according to the invention by dissolving or dispersing it in the lipid predispersion or by subsequent stirring into a finished liposome suspension.
  • Such drugs can also be modified
  • the method according to the invention is distinguished from the previously described production methods by a very good reproducibility of the liposome properties produced.
  • the liposomes produced show only slight fluctuations in their properties (especially inclusion, size and size distribution). This reproducibility of the process is not adversely affected by the enlargement of the manufacturing scale.
  • the method according to the invention is also particularly suitable for being carried out under aseptic conditions. This is particularly important in cases where the desired liposomes cannot be subjected to terminal sterile filtration (0.2 ⁇ m) due to their size.
  • sterile filtration 0.2 ⁇ m
  • the end product can usually be sterile filtered (e.g. 0.2 ⁇ m).
  • the method according to the invention offers the possibility of causing germs to be removed from the outset by extrusion using filters of suitable pore size (less than or equal to 0.6 ⁇ m), as a result of which subsequent sterile filtration could be dispensed with.
  • the inventive method is also particularly suitable for the production of storage-stable liposomes.
  • storage-stable liposomes in which the unencapsulated lopromide portion is not separated off, for example after three months of storage in the refrigerator, no decrease in pH and inclusion and no change in the mean diameters can be determined.
  • the process according to the invention for the first time offers the possibility of continuously producing large quantities of emulsions with reproducible properties.
  • the advantages listed above (liposome production) of the high-pressure extrusion process according to the invention also come into play when producing emulsions.
  • the previously not described use of filter extrusion in this area which was only opened by the high-pressure extrusion process according to the invention, enables the flexible production of emulsions over a wide range of sizes (100 nm-20 ⁇ m average diameter of the dispersed phase) without major expenditure on equipment.
  • the process is characterized by the fact that large amounts of the inner phase can be processed, and mostly direct production (without predispersion) is also possible.
  • this process can be used to produce two or multi-phase emulsions (e.g. W / O, O / W, W / O / W or O / W / O).
  • Vegetable oils such as soybean oil, castor oil, safflower oil or olive oil can be used as the oil phase.
  • Suitable emulsifiers are, for example, egg and soy lecithins or pure phospholipids from such fractions.
  • nonionic surfactants such as e.g. higher fatty alcohols, sorbitan fatty acid esters or polyethylene glycol ethers or esters are used.
  • the water phase can consist of pure water (p.i.
  • emulsions for parenteral nutrition can additionally contain sugars such as glucose and xylitol as well as other salts such as sodium dihydrogen phosphate, magnesium chloride or zinc acetate.
  • fats such as medium chain triglycerides may be present in the emulsion.
  • drugs can be dissolved or suspended in one or both phases before the emulsion is prepared or incorporated after the emulsion has been produced, analogously to the processes described in the literature.
  • the corresponding hydrophilic or lipophilic active ingredients can, for example, belong to the substance classes listed above (liposome production).
  • EPS egg phosphatidylserine, Lipoid EPS, Lipoid KG
  • PCS Pnotonic correlation spectroscopy method for measuring
  • the device is a continuously operating high-pressure extrusion apparatus, shown schematically in Figure 1.
  • a temperature-controlled storage vessel (1) which is connected via a pipe connection to a pneumatic piston air pump (2) which has a transformation factor of approximately 250.
  • the piston air pump is operated with nitrogen, with the inlet pressure being set via an inlet valve (3).
  • a pipe connection leads from the high-pressure pump via a vent valve (4) and a pressure gauge (5) to a high-pressure filter holder (6), which is suitable for holding membrane filter disks with a diameter of 47 mm.
  • the dispersion emerging from the filter holder is discharged via a hose connection (7), which is used either for product removal or for product return to the storage vessel (1).
  • This device shown schematically in Figure 2, is also a continuously operating high-pressure extrusion apparatus.
  • a metal cup pre-filter (6) with a pore diameter of, for example, 35 ⁇ m is installed behind the manometer (5).
  • REPLACEMENT LEAF Two-chamber storage vessel (1) leads.
  • the outlet of the upper of the two vessels (1a) is provided with a three-way valve (1c), which allows the dispersion contained therein to be removed in whole or in part or to be fed through the lower vessel (1b) again to the high-pressure extrusion process.
  • the mean vesicle diameter is determined by means of PCS (Submicron particle sizer autodilute model 370, Nicomp Instr. Corp., Goleta, CA).
  • Example B 1 Preparation of a liposome suspension with 50 mg SPC / ml
  • the predispersion obtained in this way is filtered sequentially with the apparatus according to the invention at a working pressure between 3 and 10 MPa, each 5 times over 2 polycarbonate membranes of decreasing pore size (5.0, 1.0, 0.4, 0.2, 0.1, 0.05 and 0.03 ⁇ m).
  • the liposome suspension obtained is slightly opalescent and the liposomes have an average diameter of 64 nm.
  • Example B 2 Preparation of a liposome suspension with 200 mg SPC / ml
  • the liposome suspension obtained is slightly opalescent and the liposomes have an average diameter of 73 nm.
  • Example B 3 Preparation of a liposome suspension with 400 mg SPC / ml
  • the liposome suspension obtained has a gel-like consistency and the liposomes have an average diameter of 74 nm.
  • Example B 5 Preparation of a liposome suspension with a reduced number of extrusion steps
  • the liposome suspension obtained is highly transparent to slightly opalescent and the liposomes have an average diameter of 68 nm.
  • Example B 6-B 18 Use of Different Lipids and Lipid Mixtures
  • Placebo liposomes with different lipid compositions are produced as described below:
  • lipid film is carried out by rotary evaporation of an organic lipid solution (ethanol, methanol or chloroform / ethanol - depending on
  • Solubility at elevated temperature (e.g. 50 ° C).
  • the lipid film is dispersed with buffer solution above the phase transition temperature of the lipid mixture used (swelling time at least 15 min, shaking by hand - at least 2 min)
  • Pore size (5.0, 1.0, 0.4, 0.2, 0.1, 0.05 and possibly 0.03 ⁇ m - each 5
  • Example B 19 - B 22 batches with different lipid concentrations
  • Example B 30 - B 33 Influence of the number of passages on the mean vesicle diameter
  • Example B 6-B 18 Four batches are described as described under Example B 6-B 18, but with different numbers of passages (1, 3, 5 and 10) at each extrusion stage.
  • a mixture of SPC, Chol and SPG (6: 3: 1) in Tris buffer serves as lipid. All batches are subjected to a 3-stage extrusion process through membranes with pore sizes of 0.4, 0.1 and 0.03 ⁇ m. The results are shown in Table 4.
  • a 100 ml batch is prepared from SPC: Chol: SPG (6: 3: 1) in Tris buffer with a lipid concentration of 50 mg / ml. Without prior film formation, the lipids are weighed directly into a 100 ml measuring cylinder and mixed with 70 ° C hot Tris buffer. After swelling (30 min), they are dispersed with an Ultraturrax 30 ml at 13500 rpm at the same temperature and then extruded as described in Example B 6-B 18. The end product has an average diameter of 60 nm with a coefficient of variation of 25%.
  • Example B 35 Extrusion using a filter pore size (0.1 ⁇ m)
  • a 100 ml batch is prepared from EPC in Tris buffer with a lipid concentration of 100 mg / ml. Without prior film formation, the lipid is weighed directly into a 100 ml measuring cylinder and Tris buffer is added at room temperature. After swelling (15 min), the mixture is dispersed with an Ultraturrax at 1350 rpm for 10 min at the same temperature and then 10 times each extruded over two superimposed 0.1 ⁇ m polycarbonate filters. The end product has an average diameter of approx. 120 nm with a variation coefficient of 32%.
  • Example B 36 Preparation of a large batch (1 l liposome suspension) with high flow rates
  • a 1000 ml batch is prepared from SPC in Tris buffer with a lipid concentration of 100 mg / ml. Without prior film formation, the lipid is directly in one
  • the flow rate is independent of the pore size
  • Liposomes have an average diameter of approx. 110 nm at one
  • a 100 ml batch is prepared from SPC in Tris buffer with a lipid concentration of 500 mg / ml. Without prior film formation, the lipid is weighed directly into a 100 ml measuring cylinder and Tris buffer is added at room temperature. After swelling (30 min), the mixture is dispersed with an Ultraturrax for 10 min at 13,500 rpm at the same temperature, a gel-like consistency being obtained. This gel is then extruded twice sequentially over two superimposed polycarbonate filters (1, 0 - 0.2 and 0.1 ⁇ m) without the membranes becoming blocked.
  • the liposomes (gel) obtained after 2 passages through the last filter combination (0.1 ⁇ m) have an average diameter of approx. 180 nm with a variation coefficient of 41%.
  • Example B 38 Preparation of a batch using polytetrafluoroethylene (PTFE) filters
  • a 100 ml batch is prepared from SPC in Tris buffer with a lipid concentration of 100 mg / ml. Without prior film formation, the lipid is weighed directly into a 100 ml measuring cylinder and Tris buffer is added at room temperature. After swelling (15 min), the mixture is dispersed with an Ultraturrax for 10 min at 13,500 rpm at the same temperature. This predispersion is then extruded 10 times sequentially over two superimposed PTFE filters (5.0 - 1.2 and 0.2 ⁇ m).
  • the liposomes obtained after 10 passages through the last filter combination (0.2 ⁇ m) have an average diameter of approx. 210 nm with a variation coefficient of approx. 30%.
  • Example B 39 Preparation of a batch using a metal (cup) filter (5 ⁇ m)
  • a 1000 ml batch is prepared from SPC in Tris buffer with a lipid concentration of 100 mg / ml. Without prior film formation, the lipid is weighed directly into a 1000 ml measuring cylinder and Tris buffer is added at room temperature. After swelling (15 min), the mixture is dispersed with an Ultraturrax for 10 min at 13500 rpm at the same temperature and then extruded 10 times through a metal (cup) filter with a nominal pore size of 5 ⁇ m. The liposomes obtained after 10 passages have an average diameter of approximately 1.4 ⁇ m with a coefficient of variation of approximately 80%.
  • a 100 ml batch is prepared with 4 g of VolpoN3 (polyoxyethylene glycol-lauryl alcohol) and 1 g of cholesterol in Tris buffer. Without prior film formation, the lipids are weighed directly into a 100 ml measuring cylinder and Tris buffer is added at RT. After swelling (15 min), they are dispersed with an Ultraturrax for 10 min at 13,500 rpm at the same temperature and then sequentially extruded 5 times each over two superimposed polycarbonate filters of decreasing pore size (5.0 - 0.2 and 0.05 ⁇ m) . The end product has an average diameter of 53 nm with a coefficient of variation of 33%.
  • VolpoN3 polyoxyethylene glycol-lauryl alcohol
  • Example B 41 - B 44 Inclusion of lopromide using various manufacturing methods
  • Example B 45-B 49 lopromide inclusion and vesicle size depending on the pore size of the last extrusion step
  • Example B 43 Batches are prepared and characterized as in Example B 43, but with the change that the pore size of the last extrusion step is varied with each batch.
  • the last pore sizes are 1.0, 0.4, 0.2, 0.1 and 0.05 ⁇ m.
  • the Freeze-Thaw cycles are carried out after extrusion through 5.0 ⁇ m.
  • the lipid concentration is 150 mg / ml.
  • Table 6 The mean values and coefficients of variation of the results from three approaches are summarized in Table 6.
  • Example B 50-B 53 lopromide inclusion and vesicle size as a function of the lipid concentration
  • Example B 43 Batches are prepared and characterized as in Example B 43, but with the change that different lipid concentrations (50, 100, 150 and 160 mg / ml) are used.
  • the mean values and coefficients of variation of the results from three approaches are summarized in Table 7.
  • Example B 54 Three-month stability of lopromide liposomes
  • Table 8 shows the properties of the corresponding liposomes at the respective times.
  • Example B 55- B 57 Inclusion of Gd-DTPA using various manufacturing methods
  • Example B 6-B 18 100 ml of liposome suspensions are prepared as described under Example B 6-B 18, which contain the water-soluble, ionic MRI contrast agent gadopentetic acid dimeglumine salt (hereinafter only called Gd-DTPA).
  • Gd-DTPA water-soluble, ionic MRI contrast agent gadopentetic acid dimeglumine salt
  • the Gd concentration in the end product is 180 ⁇ mol / g
  • SPC and Chol in a molar ratio (7: 3) are used as lipids.
  • the starting solution is with
  • REPLACEMENT LEAF Water 1 1 diluted Magnevist ® used.
  • the pore size of the last extrusion stage is 0.1 ⁇ m.
  • the manufacturing processes differ as follows:
  • Example B 57 Method as described in Example B 6, but after extrusion through a 0.4 ⁇ m pore size, the batch is subjected to 3 freeze-thaw cycles (Freeze-Thaw). Is frozen in glass vials in methanol / dry ice at -70 to -80 ° C, thawed in a water bath at + 70 ° C. Extrusion is carried out at room temperature.
  • Example B 58-B 60 Gd-DTPA inclusion and vesicle size as a function of the pore size of the last extrusion step
  • Example B 57 Batches are prepared and characterized as in Example B 57, but with the change that the pore size of the last extrusion step is varied with each batch.
  • the last pore sizes are 0.2, 0.1 and 0.05 ⁇ m.
  • the mean values and variation coefficients of the results from three approaches are summarized in Table 10.
  • Example B 61 - B 63 Gd-DTPA inclusion and vesicle size depending on the lipid concentration
  • Example B 57 Batches are prepared and characterized as in Example B 57, but with the change that different lipid concentrations (100, 150 and 200 mg / ml) are used.
  • the mean values and variation coefficients of the results from three approaches are summarized in Table 11.
  • Example B 64 Preparation of liposomes with a lipophilic drug (methyiprednisolone aconate - MPA)
  • the liposomes thus obtained have an average diameter of 189 nm with a coefficient of variation of 30%.
  • the MPA is completely encapsulated in the liposomes.
  • Lipoid E 80 1.5 g of Lipoid E 80 are suspended in about 50 ml of double-distilled water and then 5 ml of filtered soybean oil (both Lipoid KG, Ludwigshafen) are added. After adding double-distilled water to 100 ml, the mixture is predispersed for 10 minutes with an Ultra-Turrax (13500 rpm). This predispersion is then extruded 10 times over two superimposed polycarbonate filters (0.1 ⁇ m). The droplet size of the homogeneous, yellowish-cloudy O / W emulsion thus obtained is approximately 430 nm.
  • Example B 66 Preparation of a 5% O / W emulsion using a PTFE filter
  • Lipoid E 80 1.5 g of Lipoid E 80 are suspended in about 50 ml of bidistilled water and then 5 ml of filtered soybean oil (both Lipoid KG, Ludwigshafen) are added. After adding double-distilled water to 100 ml, the mixture is predispersed for 10 min with an Ultra-Turrax (13500 rpm). This predispersion is then extruded 10 times each over two superimposed PTFE filters (0.2 ⁇ m). The droplet size of the homogeneous, yellowish-cloudy O / W emulsion thus obtained is approximately 230 nm.
  • Example B 67 Preparation of a 20% O / W emulsion
  • Cremophor S9 1 g of Cremophor S9 (BASF) is suspended in about 50 ml of double-distilled water and then 20 ml of filtered soybean oil (Lipoid KG, Ludwigshafen) are added. After adding distilled water to 100 ml, the mixture is predispersed for 1 min with an Ultra-Turrax (8000 rpm). This predispersion is then extruded 10 times over two superimposed polycarbonate filters (0.1 ⁇ m). The droplet size of the homogeneous, milky, cloudy O / W emulsion thus obtained is approximately 880 nm.

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Abstract

L'invention concerne un procédé continu de préparation de systèmes dispersés liquides, qui se caractérise en ce que l'on procède à l'extrusion d'une prédispersion, à une pression élevée comprise entre 6,6 et 250 MPa, de manière séquentielle en faisant passer la préparation par 1 à 8 étages de filtration de grosseur de pore décroissante comprise entre 0,01 et 35 νm. Il peut y avoir jusqu'à 20 passages par étage de filtration. L'invention concerne en outre un dispositif pouvant servir à mettre ledit procédé en ÷uvre.
EP93922883A 1992-10-16 1993-10-13 Procede et dispositif de preparation de systemes disperses liquides Ceased EP0665756A1 (fr)

Applications Claiming Priority (5)

Application Number Priority Date Filing Date Title
DE4235381 1992-10-16
DE19924235381 DE4235381A1 (de) 1992-10-16 1992-10-16 Apparatur zur Herstellung flüssiger, disperser Systeme
DE4328331 1993-08-18
DE19934328331 DE4328331A1 (de) 1993-08-18 1993-08-18 Kontinuierliches Hochdruckextrusionsverfahren zur Herstellung von Liposomen und Emulsionen
PCT/DE1993/000997 WO1994008626A1 (fr) 1992-10-16 1993-10-13 Procede et dispositif de preparation de systemes disperses liquides

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EP0665756A1 true EP0665756A1 (fr) 1995-08-09

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AU (1) AU5174193A (fr)
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WO (1) WO1994008626A1 (fr)

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CA2146963A1 (fr) 1994-04-28
US6241967B1 (en) 2001-06-05

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