NO171886B - PROCEDURE FOR THE PREPARATION OF A PHARMACEUTICAL PREPARATION IN THE FORM OF AN Aqueous DISPERSION CONTAINING LIPOSOMES CONSISTING OF PHOSPHOLIPIDE COMPONENTS - Google Patents

Abstract

The compositions consist of a) a phospholipid of the formula <IMAGE> in which m is two or three, R1 and R2 are, independently of one another, alkyl, alkenyl or acyl each with 10 to 20 C atoms, and X is a direct bond, C1-C4-alkylene, C2-C4-alkenylene or C1-C4-alkylene or C2-C4-alkenylene substituted by hydroxyl, or a pharmaceutically acceptable salt thereof, b) a phospholipid of the formula <IMAGE> in which R3 and R4 are the acyl group of a saturated or unsaturated carboxylic acid with 10 to 20 C atoms and 1 to 2 double bonds, c) a compound or a mixture of compounds with pharmacological properties and, where appropriate, d) a lipid from the group comprising phosphatidylcholines, phosphatidylserine, phosphatidylinositol, phosphatidylglycerol, cardiolipin and cholesterol and, where appropriate, a pharmaceutically acceptable carrier solution which is buffered to pH 7.0 to 7.8 and, where appropriate, pharmaceutically acceptable auxiliaries. The compositions can be used in the form of liposomes as administration systems, for example for cancer chemotherapy.

Description

The present invention relates to the production of pharmaceutical preparations containing acylated phosphatidylethanolamine derivatives, phosphatidylethanolamine and compounds which have pharmacological properties.

The pharmaceutical preparations produced according to the present invention are administered in the form of liposomes.

Pharmaceutical delivery systems based on liposomes are described in the general review published by G. Gregoria-dis, "Liposome Technology", Volume II, "Incorporation of Drugs, Proteins and Genetic Material"; CRC Press 1984. Such systems have the advantage that biologically active material can be introduced into tissues by phagocytosis, especially into tissues of the reticulo-endothelial system. E.g. is a transport mechanism known by which antibiotics are introduced into infected tissue by phagocytosis so as to cause improved removal or destruction of the infecting microorganism. Endocytosis is also a useful mechanism in the fight against inflammatory centers. Antirheumatic pharmaceutical agents encapsulated in liposomes are preferentially introduced into infected tissue as opposed to "healthy" tissue. Furthermore, cytostatic agents, commonly known as "anti-cancer agents", when encapsulated in liposomes, can be introduced into specific organs of the reticulo-endothelial system (liver, spleen or marrow). Due to filtration in the capillary tubes in the lung and later transport by migrating monocytes, biologically active material can e.g. compounds that have immunomodulatory properties are concentrated in alveolar macrophages. This results in an improved effect on metastatic lung tumors and in a simultaneous reduction of toxicity.

A representative version of previously known technology is constituted by Swedish explanatory document no. 459,228, which describes liposome compositions containing lysophospholipids and other phospholipids. These compounds are known for their carcinostatic properties.

It has now surprisingly been found that the uptake of liposomes and their endocytosis by macrophages, especially alveolar macrophages, is increased when acylated phosphatidylethanolamine derivatives are included in the shell structure of the liposomes.

The present invention relates to a method for producing a pharmaceutical preparation in the form of an aqueous dispersion containing liposomes consisting of

a) a synthetic phospholipid with the formula

where m means two or three, R 1 and R 2 independently of each other

means alkyl, alkenyl or acyl, each having 10 to 20 carbon atoms, X means a direct bond, C₁-C₄alkylene, C₂-C₄alkenylene, or C₁-C₄alkylene or C₂-C₄alkenylene substituted with hydroxy, or a pharmaceutically acceptable salt thereof,

b) a synthetic phospholipid of the formula

where R3 and R4 mean the acyl group of a saturated or an unsaturated carboxylic acid with 10 to 20 carbon atoms and 1-2 double bonds, 1 a ratio phospholipid component (I):phospholipid component (II) from approx. 10:90 up to 50:50, c) a lipophilic compound or a mixture of compounds with pharmacological activity, and, optionally, d) a lipid from natural sources selected from phosphatidylcholine, phosphatidylserine, phosphatidylinositol, phosphatidylglycerol,

cardiolipin and cholesterol, and derivatives thereof, and a pharmaceutically acceptable carrier solution buffered to pH 7.0-8.0 and, optionally, pharmaceutically acceptable additives.

In connection with the description of the present invention, the general terms used above and below preferably have the following meanings: The term "lower" used in definitions of organic residues, e.g. lower alkyl, lower alkylene, lower alkoxy, lower alkanoyl, etc., means that such organic residues, unless otherwise expressly stated, contain up to 7, preferably up to 4, carbon atoms.

The nomenclature for the phospholipids of the formulas I and II is in accordance with the recommendations of the IUPÅC and ITJB commission concerning biochemical nomenclature (CBN) according to Eur. J. of Biochem. 79, 11-21 (1977) "Nomenclature of Lipids"

(sn nomenclature, stereospecific numbering).

Unless otherwise stated, generic names proposed by the World Health Organization (WHO) (Recommended International Non-proprietary Names) have been used to define the active ingredients, these names are taken from the standard textbook "Pharmazeutische Chemie" (E. Schroder, C. Rufer and r. Schmiechen, Thieme Verlag Stuttgart, 1982) and the Merck Index (tenth edition).

In the synthetic phospholipid of formula I (component a), m is preferably two.

Alkyl R 1 and/or R 2 is preferably straight chain with an equal number of from 10 to 20 carbon atoms, e.g. n-decyl, n-dodecyl, n-tetradecyl, n-hexadecyl, n-octadecyl or n-icosyl.

Alkenyl R 1 and/or R 2 is preferably straight chain with an equal number of from 10 to 20 carbon atoms and a double bond, e.g. 9-cis-dodecenyl, 9-cis-tetradecenyl, 9-cis-hexa-decenyl, 6-cis-octa-decenyl, 6-trans-octadecenyl, 9-trans-octadecenyl or 9-cis-icosenyl.

Acyl R 1 and/or R 2 is preferably straight chain with an equal number of from 10 to 20 carbon atoms, e.g. Cio-C20~alltanoyl or Cio-C20~al^en°yl•

Alkanoyl R 1 and/or R 2 are preferably n-decanoyl, n-dodecanoyl, n-tetradecanoyl, n-hexadecanoyl, n-octadecanoyl and n-icosanoyl.

Alkenoyl R 1 and/or R 2 are preferably 9-cis-dodecenoyl, 9-cis-tetradecenoyl, 6-trans-octadecenoyl, 9-cis-octadecenoyl, 9-trans-octadecenoyl, 11-cis-octadecenoyl and 9-cis-icosenoyl .

X defined as C 1 -C 4 alkylene is straight chain or branched C 1 -C 4 alkylene, e.g. methylene, 1,1-ethylene, 1,1-, 1,2- or 1,3-propylene, or preferably 1,2-ethylene.

X defined as C2~C4-alkenyl is preferably straight-chain alkenyl, e.g. the vinyl, propenylene, or 1,2- or 2,3-butenylene.

X defined as C1-C4-alkylene or C2-C4-alkenylene substituted with hydroxy is preferably straight-chain C1-C4-alkylene substituted with 1 or, depending on the number of carbon atoms, up to 4 hydroxy groups, e.g. 1-hydroxy-1,2-ethylene, 1,2-dihydroxy-1,2-ethylene, 1- or 2-hydroxy-1,3-propylene or 1,2-dihydroxy-1,3-propylene.

A pharmaceutically acceptable salt of the phospholipid (I) is preferably formed by reaction with one or two equivalents of dilute aqueous alkali metal hydroxide, e.g. sodium or potassium hydroxide and is preferably the mono or disodium salt.

Other pharmaceutically acceptable salts are formed by reaction with amines such as trimethyl, ethyl, diethyl, or triethylamine, piperidine, piperazine, 2-hydroxyethylpiperazine, cyclohexylamine, pyrrolidine, or choline.

In the synthetic phospholipid (I), R 1 and R 2 are preferably straight-chain alkenoyl with an equal number of from 10 to 20 carbon atoms, e.g. 9-cis-dodecenoyl, 9-cis-tetradecenoyl, 9-cis-hexadecenoyl, 6-cis-, 6-trans-, 9-cis-, 9-trans-, or 11-cis-octadecenoyl, or 9-cis- icosenoyl, X is C2~C4 alkylene, e.g. 1,2-ethylene or 1,3-propylene, or C2~C4-alkenylene, e.g. the vinyl.

Most preferred are the sodium or disodium salts of N-[1,2-di-(9-ci s-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxysuccinylamine and N-[1,2-di-(9- cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxyglutarylamine.

In the phospholipid of formula II (component b)), the acyl groups R3 and R4 are preferably straight-chain with an equal number of from 10 to 20 carbon atoms, e.g. Cio-<C>20_alkanoyl or Cio~<c>20~ alkenoyl, especially 9-cis-dodecenoyl, 9-cis-tetradecenoyl, 9-cis-hexadecenoyl, 6-cis-, 6-trans-, 9-cis-, 9-trans- or 11-cis-octadecenoyl.

Most preferred is 1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine.

Compounds or mixtures of compounds that have pharmacological activity (component c)) are preferably drugs selected from the group consisting of antiphlogistic and/or anti-inflammatory agents, antibiotics, fungicides, antifungals, antineoplastic agents, and compounds that have an immunomodulatory effect .

Antiphlogistic and/or anti-inflammatory agents are preferably glucocorticoids, e.g. cortisone, hydrocortisone, prednisone, prednisolone, fluocortolone, triamquinolone, methylprednisolone, prednylidene, paramethasone, dexamethasone, betamethasone, beclomethasone, fluprednylidene, desoximetasone, fluocinolone, flumetasone, diflucortolone, clocortolone, clobetasol or fluorocortin butyl ester, salts of substituted phenylacetic acids or 2-phenylpropionic acids , e.g. alclofenac, ibuprofenac, ibuprofen, clindanac, fenclorac, ketoprofen, fenoprofen, indoprofen, fenclofenac, diclofenac, flurbiprofen, pirprofen, naproxen, benoxaprofen, carprofen or cicloprofen; analgesically active anthranilic acid derivatives, e.g. of the formula:

in which R 1 , R 2 and R 3 , independently of each other, stand for hydrogen, methyl, chlorine or trifluoromethyl, e.g. mefenamic acid, flumenamic acid, tolfenamic acid or meclofenamic acid; analgesically active aniline-substituted nicotinic acid derivatives, e.g. miflumic acid, clonixin, or flunixin; analgesically active heteroarylacetic acids or 2-heteroarylpropionic acids having a 2-indol-3-yl or pyrrol-2-yl residue, e.g.

indomethacin, oxmetacin, intrazol, acemetacin, cinmetacin, zomepirac, tolmetin, colpirac or tlaprofenic acid, analgesically active indenyl acetic acids, e.g. sulindac, analgesically active heteroaryloxyacetic acids, e.g. benzadac.

Antibiotics are preferably tetracycline derivatives of the formula:

in which R 1 stands for hydrogen or pyrrolidin-1-ylmethyl, R 2 stands for hydrogen or hydroxy, R 3 stands for hydrogen, hydroxy or methyl, R 4 stands for hydrogen or methyl, and R 5 stands for hydrogen, chlorine or dimethylamino, e.g. chlortetracycline, oxytetracycline, tetracycline, demethylchlortetracycline, methacycline, doxycycline, minocycline or rolitetracycline, aminoglycosides, e.g. kanamycin, amikacin, gentamicin, cla'c2 or C2D, sisomycin, netilmicin, spectinomycin, streptomycin, tobramycin, neomycin B, dibecacin, or kanendomycin, macrolides, e.g. maridomycin or erythromycin, lincomycins, e.g. clindamycin or lincomycin, penicillanic acid and cephalosporanic acid derivatives having antibiotic activity with 6p<-> or 7p<->acylamino groups, which are present in fermentative, semi-synthetic or synthetically obtainable 6p<->acylaminopenicillanic acid or 7p-acylamino-cephalosporanic acid derivatives or in 7g<->acylaminocephalosporanic acid derivatives modified in the 3-position, e.g. penicillinic acid derivatives have become known as penicillin G or V, pheneticillin, propicillin, nafcillin, oxacillin, cloxacillin, dicloxacillin, flucloxacillin, cyclacillin, epicillin, mecillinam, methicillin, azlocillin, sulbenicillin,

ticarcillin, mezlocillin, piperacillin, carindacillin, azidocillin or clklazillln, or cephalosporin derivatives that have become known under the names cefaclor, cefuroxime, cefazlur, cefacetril, cefazolin, cephalexin, cefadroxil, cephaloglycin, cefoxitin, cephaloridine, cefsulodin, cefotiam, ceftazidin, cefonicide, cefotaxime, cefmenoxime, ceftizoxime, cephalotin, cefradin, cefamandole, cefanone, cefapirin, cefroxadine, cefatrizine, cefazedone, ceftrixone or ceforanid, other β-lactam antibodies of clavam-; the penem or carbapenem type, e.g. moxalactam, clavulanic acid, nocardicin, sulbactam, aztreonam or thienamycin, or other antibiotics of the biomycin, novobiocin, chlor or thiamphenicol, rifampicin, fosfomycin, colicin or vancomycin type.

Anti-soil agents are preferably antimony compounds, e.g. potassium antimonyl tartrate, stibofen, sodium stibocaptate or sodium stibogluconate.

Antimyotics are e.g. thiocarbonic acid derivatives, e.g. dibenzthion, tolnaftate, or tolcidate, imidazole derivatives, e.g. clotrimazole, miconazole, econazole, iconazole, or ketoconazole, or polyene derivatives such as nystatin, natamycin, or amphotericin B.

Antineoplastic agents are preferably alkylating agents containing the bis-(2-chloroethyl)-amine group such as chlormethine, chlorambucil, melphalan, uramustine, mannomustine, extramustine phosphate, mechlorethamine oxide, cyclophosphamide, ifosfamide, or triphosphamide, alkylating agents containing a substituted aziridine group, e.g. tretamine, thiotepa, triazicon or mitomycin, alkylating agents of the methanesulfone ester type such as busulfan, alkylating N-alkyl-N-nitrosourea derivatives, e.g. carmustine, lomustine, semustine, or streptozotocin, alkylating agents of the mitobronitol, dacarbazine, or procarbazine type, complexing agents such as cis-platin, antimetabolites of the folinic acid type, e.g. methotrexate, purine derivatives such as mercaptopurine, thioguanine, azathioprine, thiamiprine, vidarabine, or puromycin, pyrimidine derivatives, e.g. fluorouracil, floxuridine, tegafur, cytarabine, idoxuridine, flucytosine, antibiotics such as dactinomycin, daunorubicin, doxorubicin, mithramycin, bleomycin Å2 or B2 or etoposide, or vinca alkaloids, possibly together with chlormethamine, prednisolone, prednisone or procarbazine.

Compounds or mixtures of compounds which have an immunomodulating effect are e.g. muramyl peptides of the formula:

in which H1 stands for L-Ala-D-isoGln-L-Ala-2-(1,2-dipalmitoyl-sn-glycero-3-hydroxyphosphoryloxy)-ethylamide, L-Ala-D-Glu-(Cg flTp-ma- L-Ala-2 - (1 ,2-di palm i toyl-sn-glycero-3-hydroxyph osphoryl-oxy )-ethylamide, L-Ala-D-isoGln-OH, L-Ala-D-Gln-NB^ -oc-n-butyl ester, L-Ala-D-isoGln-L-(stearoyl)-Lys, L-Val-D-Gln-NB^-oc-n-methyl ester, L-Ala-D-isoGln-L- Ala-1,2-dipalmitoyl-sn-glycerin ester or L-Ala-D-isoGln-L-Ala-cholesterol ester group, R2 stands for hydrogen, methyl or n-propyl, R3 stands for hydrogen, n-stearoyl, 10- (2,3-dimethoxy-1,4-dioxo-5-methyl)-2,5-cyclohexadienoyl, 2-behenoyloxy-2-methyl-propanoyl or n-octanoyl, and R4 represents hydrogen or n-octanoyl, as well as its 2-palmitoylthio derivative, lipo-peptides such as n-lauroyl-L-Ala-D-isoGln-(m-DAP-Gly)-NH2, n-lauroyl-L-Ala-D-i soGln-(L-DAP-Gly )-NH2, n-lauroyl-L-Ala-D-isoGln-(L-Lys-D-Ala)-NH2, n-octanoyl-L-Ala-D-isoGln-(L-Lys-D-Ala)- NH2 or palmitoyl-Cys-((2R)-2,3-dilauroyloxypropyl)-Ala-D-Glu-(Gly-taurine-Na)-NH2, or are lymphokines as

produced by lymphocytes, monocytes or macrophages after stimulation by antigens or mitogens.

Lymphokines are especially gamma interferon, especially natural or recombinant human gamma interferon, especially human gamma interferon obtainable according to European Patent Publication Nos. 63,482, 77,670, 83,777, 88,540, 89,676, 95,350, 99,084, 110,044 and 112,967 and the international (PCT) the public zones (W0) 83/04.053 or W0 84/02.129.

Preferred is human gamma interferon of the following amino acid sequences:

according to European patent publication 121,157 and

according to GB patent 2,107,718, human interleukin 2 which can be obtained e.g. from culture filtrates of cell cultures of leukemia or lymphoma cells after activation or stimulation with human T-cell mitogens and by purification by reversed phase EPLC, culture filtrates containing mixtures of compounds known as migration inhibitory factor (MIF), leukocyte migration inhibitory factor, macrophage activating factor (MÅF), colony stimulating factor, so as well as interleukin 1 and 2 and gamma-interferon which can be obtained from cultures of human T-lymphocytes from spleens or from peripheral blood after stimulation with antigens or mitogens, or e.g. human T-cell leukemia-lymphoma virus (HTLV I or II), phytohemagglutinin, or concanavalin, especially those culture filtrates or isolates containing a high percentage of macrophage-activating factor (MAF).

Preferred are N-acetylmuramyl-L-alanyl-D-isoglutaminyl-L-alanine-2-(1,2-dipalmitoyl-sn-glycero-3-hydroxyphosphoryloxy)-ethylamide, sodium-N-acetyl-D-muramyl-L- alanyl-D-isoglutamine or sodium-N-acetyldesmethylmuramyl-L-alanyl-D-isoglutamine of formula V, optionally in combination with purified, natural or recombinant human gamma interferon.

Lipids (component d)) selected from the group consisting of phosphatidylcholine, phosphatidylserine, phosphatidylinositol, phosphatidylglycerol and cardiolipin, are synthetic phospholipids or are mixtures of phospholipids which have different acyl groups of different molecular weight and structure, e.g. soybean or chicken egg phosphatidylcholine or phosphatidylquinoline from bovine brain, bovine liver or pig liver, phosphatilyserine from bovine brain, phosphatidylinositol from soybeans or yeast, phosphatidylglycerol from egg yolk, or cardiolipin from bovine heart.

Derivatives of cholesterol are, for example, cholestane, coprostane, ergosterol or stigmasterol.

The pharmaceutical preparations according to the present invention, when used in the form of liposomes, are characterized by their excellent phagocytosis. For example, phagocytosis of multi-lamellar liposomes consisting of a 3:7 molar mixture of sodium-N-[1,2-di-(-9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N -hydroxysuccinylamine (I) and dioleoylphosphatidylethanolamine (II) or sodium-N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxyglutarylamine (I) and dioleoylphosphatidylethanolamine (II) by mouse peritoneal macrophages higher than phagocytosis for multilamellar liposomes consisting exclusively of phosphatidylethanolamine or of a 3:7 molar mixture of phosphatidylserine and phosphatidylcholine. This can be demonstrated in vitro by incubating mouse peritoneal macrophages with multilamellar liposomes containing trace amounts of <125>j as an Uposome marker. At regular intervals the cultures are washed and the amount of cell-associated radiation determined. Furthermore, liposomes consisting of phospholipids of formulas I and II in a molar ratio of 3:7 containing immunomodulators such as MDP and gamma-interferon show higher activation of macrophages to tumoricidal activity at lower doses than liposomes consisting of phosphatidylcholine and phosphatidylserine containing the same amount of MDP and gamma interferon. This can also be shown in vitro by coating mouse peritoneal macrophages in culture wells and activating the macrophages with multilamellar liposomes consisting of sodium-N-[l,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol] -N-hydroxysuccinylamine (I) and dioleoylphosphatidylethanolamine (II) in a molar ratio of 3:7 containing MDP and gamma interferon and with multilamellar liposomes consisting of phosphatidylcholine and phosphatidylserine in the same molar ratio containing MDP and gamma interferon. The liposome preparations are used at a concentration of 100 nmol total phospholipid per well and contains 6 units of recombinant immune gamma interferon and 0.2 g of MDP. After washing the wells, IO<4> [<125>I]iododeoxyuridine - labeled BL6 melanoma cells are added. The cytotoxicity is determined after 72 hours of cultivation by measuring the radioactivity associated with the adherent viable target cells after washing the cultures three times with Hank<*>'s balanced salt solution. Percent cytotoxicity can be calculated with regard to the number of counts per minute in control cultures containing unactivated macrophages and target cells. ;The pharmaceutical preparations produced according to the present invention are, when used in the form of liposomes, further characterized by excellent release properties at low pH values. The release properties of liposomes composed of disodium N-[1,2-di-(9-c i s-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxysuccinylamine and 1,2-di-(9-cis-octadecenoyl) )-sn-glycero-3-phosphoethanolamine in a molar ratio of 3:7 is analyzed by the fluorescence method to measure the leakage from liposomes at low pH values as described by Ellens et al., Biochemistry 1984, 23, 1532-1538. To investigate the pH dependence of the leakage, liposomes are injected into buffer solutions varying from pH 4.0 to 7.4 and the percentage of captured 8-aminonaphthalene-1,3,6-trisulfonic acid (ANTS) and p-xylylene-bis- pyridinium dibromide (DPX) that replaces the encapsulated pharmaceutical compounds in this assay is determined. Complete encapsulation of the water-soluble fluorophore ANTS complexed with the "quencher" DPX removes most of the ANTS fluorescence. Leakage of ANTS from the liposomes can be followed by the increase in fluorescence caused by the release of DPX "quenching". It has now been found that essentially no leakage occurs above pE 6.0. However, when the pH is reduced, a simultaneous increase in the amount of ANTS/DPX released from the liposomes takes place. The half maximum value takes place at approx. pH 4.5 and complete release at approx. 4.0. ;It is known that pathological tissue has an ambient pH that is significantly lower than that of normal tissue. For example, primary tumors, metastases, inflammation and infections have reduced local pE environments. The liposomes pentastable according to the present invention will therefore deliver their contents, e.g. anti-inflammatory drugs or immunomodulators specifically at the site of inflammation, the primary tumor or metastasis and release these drugs into the acidic environment of this pathological tissue. E.g. release liposomes consisting of a 3:7 molar mixture of disodium-N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxysuccinylamine (I) and dioleoylphosphatidylethanolamine more of its content, e.g. an effective dose of recombinant human gamma interferon, at low pH values, e.g. pH 4, than at higher or neutral pH values. This can be demonstrated in vitro in buffer solutions of pH 7.4 and pH 4 by determining at regular intervals the amount of labeled ^<25>I-interferon released from the lipids. At a pH value of approx. 4, released e.g. more than 25$ of enclosed interferon gamma from liposomes after 180 minutes. The pharmaceutical preparations produced according to the present invention are therefore, when used in the form of liposomes, excellent administration systems for drug delivery to the local areas with reduced pE of pathological tissue. Therefore, they are particularly useful in cancer chemotherapy to overcome metastatic tumor cells. Aqueous liposome dispersions in which the phospholipids of formulas I and II are the encapsulating material and compounds or a mixture of compounds that have pharmacological activity are encapsulated, optionally after concentration or isolation of the liposomes, e.g. in the (ultra)centrifuge, are suitable for therapeutic purposes for parenteral (buccal, lingual, sublingual, i.v., i.c., topical, s.c., i.m. or nasal) final administration. For parenteral (topical) administration, the liposome-containing aqueous dispersion may be mixed with common thickeners, e.g. hydroxypropylmethylcellulose, suitable preservatives, antioxidants and perfumes, and can be used in the form of a lotion or a gel for application to the skin or mucous membranes. For parenteral administration, the aqueous dispersion of the enriched liposomes can be suspended in a suitable carrier liquid, e.g. sterile, calcium-free, isotonic sodium chloride or glucose solution, optionally buffered to pH 7.2-7.4. ;The dose of the active ingredient to be administered is generally the highest and lowest dosage amount as prescribed e.g. in the Deutsches Arzneimittelbuch (DAB) [German Pharmacopoieia] for the specific active ingredient for the particular form of administration, the patient's age and state of health. However, aqueous liposome dispersions also have the advantage that the active ingredient administered in smaller doses can nevertheless be transferred to the receptors and provide a therapeutic effect, or on the other hand, that the administration of high doses is possible without unwanted side effects. ;The preferred dosage amount for the liposome-encapsulated immunomodulators of the muramyl peptide or lipopeptide type is approx. 0.001 up to 10 mg/kg body weight per dose. For human gamma interferon or mixtures containing MAF, the preferred dosage amount is approx. 0.01 ml of liposome dispersion per kilogram of body weight, containing 100-1000 units of gamma interferon or MAF. If muramyl peptides are administered together with gamma interferon, it is estimated that the highest dose that can be administered to a human being of approx. 70 kg is approx. 10 mg liposomes per kg body weight, containing 3 micrograms of the muramyl peptide and 1,500 units of gamma interferon. The highest and the lowest dose of the encapsulated material, the concentration of the phospholipids in the aqueous phase as well as the concentration of the encapsulated compounds may vary according to results that can be determined experimentally in clinical trials. The present invention preferably relates to a method for the production of pharmaceutical preparations in the form of an aqueous dispersion containing liposomes consisting of: a) a phospholipid of formula I, in which m stands for two, R^ and R£ are as defined above, X stands for C^-C^alkylene, C2-C4-alkenylene or C1-C4-alkylene substituted by hydroxy or a pharmaceutically acceptable salt thereof, b) a phospholipid of formula II, in which R3 and R4 independently of each other stand for the straight chain <C>10 ~c20~alkan°yl or C10<-C>20<->;alkenoyl, ;c) a compound or a combination of compounds that have pharmacological activity and optionally a pharmaceutically acceptable carrier solution buffered to pH 7.2-7.4 . More preferably, the present invention relates to a method for the production of pharmaceutical preparations in the form of an aqueous dispersion containing liposomes consisting of a) a phospholipid of formula I, in which m stands for two, Rj and R2 are defined above, X stands for C2~C4 -alkylene or C2-C4-alkenylene, or a pharmaceutically acceptable salt thereof, b) a phospholipid of formula II, in which R3 and R4 independently of each other stand for the straight chain <C>io-C20~alkan°y1 or C10<-C> 20<->;alkenoyl, ;c) a compound or a combination of compounds selected from the group consisting of antiphlogistic and/or anti-inflammatory agents, antibiotics, antimalarial agents, antineoplastic agents and immunomodulators and a pharmaceutically acceptable carrier solution buffered to pH 7.2-7 ,4. The present invention specifically relates to a method for the production of pharmaceutical preparations in the form of an aqueous dispersion containing liposomes consisting of: a) a phospholipid of formula I, in which m stands for two, R^ and R2 stand independently of each other for straight-chain alkanoyl or ; alkenoyl with an equal number of from 10 to 20 carbon atoms, X stands for C2-C4~alkylene, e.g. 1,2-ethylene or 1,3-propylene, or C2-C4~alkenylene, e.g. the vinyl, or a pharmaceutically acceptable salt thereof, b) a phospholipid of formula II, in which R3 and R4 stand for the straight chain CiQ-C2o~alken°y1 with an equal number of from 10 to 20 carbon atoms, c) a compound or a combination of compounds selected from the group consisting of antiphlogistic and/or anti-inflammatory agents, antibiotics, neoplastic agents and immunomodulators and a pharmaceutically acceptable carrier solution buffered to pH 7.2-7.4. The invention relates more specifically to a method for the production of pharmaceutical preparations in the form of an aqueous dispersion containing liposomes consisting of a) a phospholipid of formula I, in which m stands for two, R^ and R2 stand independently of each other for 9-cis-dodecenoyl, 9-cis-tetradecenoyl, 9-cis-hexadecenoyl, 6-cis-, 6-trans-, 9-cis-, 9-trans-, or 11-cis-octadecenoyl, or 9-cis-icosenoyl, X stands for C2 -C4-alkylene, e.g. 1,2-ethylene or 1,3-propylene, or C2~C4-alkenylene, e.g. vinylene, or a pharmaceutically acceptable salt thereof, b) a phospholipid of formula II, in which E3 and R4 independently stand for 9-cis-dodecenoyl, 9-cis-tetradecenoyl, 9-cis-hexadecenoyl, 6-cis-, 6 -trans-, 9-cis-, 9-trans-, or 11-cis-octadecenoyl, or 9-cis-icosenoyl, c) a compound or a combination of compounds selected from the group consisting of antiphlogistic and/or anti-inflammatory agents, antibiotics , antineoplastic agents and immunomodulators and a pharmaceutically acceptable carrier solution buffered to pH 7.2-7.4. In particular, the present invention relates to a method for the preparation of pharmaceutical preparations in the form of an aqueous dispersion containing liposomes consisting of a) sodium or disodium-N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3 -phosphoethanol]-N-hydroxysuccinylamine or sodium or disodium-N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxyglutarylamine, b) 1,2-di -(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine, c) a compound or a combination of compounds selected from the group consisting of antiphlogistic and/or anti-inflammatory agents, antibiotics, antineoplastic agents, and immunomodulators and a pharmaceutically acceptable carrier -solution buffered to pH 7.2-7.4. Of high preference is a method for the production of pharmaceutical preparations in the form of an aqueous dispersion containing liposomes consisting of a) sodium or disodium-N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3 -phosphoethanol]-N-hydroxysuccinylamine or sodium or disodium-N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxyglutarylamine, b) 1,2- di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine, c) a compound or a combination of compounds selected from the group consisting of diclofenac, pirprofen, mitomycin, cytarabine, dactinomycin, daunorubicin, doxorubicin, etoposide, N- acetyl-D-muramyl-L-alanyl-D-isoglutaminyl-L-alanine-2-(1,2-dipalmitoyl-sn-glycero-3-hydroxyphosphoryloxy)-ethylamide, N-acetyl-D-muramyl-L-alanyl- D-glutamic acid-CC^-L-alanine-2-(1,2-dipalmitoyl-sn-glycero-3-hydroxy-phospho oryl-oxy)-ethylamide disodium salt, N-acetyl-D-muramyl-L-alanyl- D-isoglutamine sodium salt, N-acetyldesmethyl-muramyl-L-alanyl-D-isoglutamine sodium salt, N-acetyl-D-mura myl-L-alanyl-D-glutamine-a-n-butyl ester, Na<->(N-acetyl-D-muramyl-L-alanyl-D-isoglutaminyl)-N^-stearoyl-L-lysine, 6-O-stearoyl -N-acetylD-muramyl-L-alanine-D-isoglutamine and lymphokines and a pharmaceutically acceptable carrier solution buffered to pH 7.2-7.4. Particularly preferred is a method for the production of pharmaceutical preparations in the form of an aqueous dispersion containing liposomes consisting of a) sodium or disodium-N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3- phosphoethanol]-N-hydroxysuccinylamine or sodium or disodium-N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxyglutarylamine, b) 1,2-di- (9-cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine, c) a compound or a combination of compounds from the group consisting of diclofenac, pirprofen, mitomycin, cytarabine, dactinomycin, daunorubicin, doxorubicin, etoposide, N-acetyl -D-muramyl-L-alanyl-D-isoglutaminyl-L-alanine-2-(1,2-dipalmitoyl-sn-glycero-3-hydroxyphosphoryloxy)-ethylamide, sodium N-acetyl-D-muramyl-L-alanyl -D-isoglutamine, sodium-N-acetyldesmethyl-muramyl-L-alanyl-D-isoglutamine, purified, natural or recombinant human gamma interferon, interleukin 2, and compounds obtained from cultures of human ;T-lymphocytes from the spleen or from per transfused blood after stimulation with antigens or mitogens and which is characterized by a high percentage of macrophage activating factor (MAF) and a pharmaceutically acceptable carrier solution buffered to pH 7.2-7.4. Most preferred is a method for the production of pharmaceutical preparations in the form of an aqueous dispersion containing liposomes consisting of: a) sodium or disodium-N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero- 3-phospho-ethanol]-N-hydroxysuccinylamine or sodium- or disodium-N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxyglutarylamine, b) 1 ,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine, c) a compound or a combination of compounds; consisting of N-acetyl-D-muramyl-L-alanyl-D-isoglutaminyl-L -alanine-2-(1,2-dipalmitoyl-sn-hydroxyphosphoryloxy)-ethylamide, sodium N-acetyl-D-muramyl-L-alanyl-D-isoglutamine, sodium N-acetyldesmethylmuramyl-L-alanyl-D-glutamine , and purified, natural or recombinant human gamma interferon and a pharmaceutically acceptable carrier solution buffered to pH 7.2-7.4. The method for producing the pharmaceutical preparations mentioned above is characterized by the fact that: a) a homogeneous mixture consisting of phospholipids of the formulas I and II, a lipophilic compound or a mixture of compounds that have pharmacological activity and, optionally, a lipid mentioned above from natural sources and the homogeneous mixture which is thereby obtained by dispersion in an aqueous phase, or ;<p>) a homogeneous mixture consisting of phospholipids of the formulas I and II and, optionally, a lipid mentioned above from natural sources is prepared and the homogeneous mixture which thereby obtained is optionally dispersed in an aqueous phase containing a hydrophilic compound or mixture of compounds having pharmacological activity and, if necessary after carrying out any of the process steps a) or <p>), the aqueous dispersion obtained is buffered to pH 7.0-7.8 and, if desired, separating non-encapsulated lipids and/or compounds having pharmacological activity tet from the aqueous phase and/or the liposomes thereby obtained are concentrated or separated from the aqueous phase. The homogeneous mixture is produced by forming a film or a lyophilisate. ;The film is produced by method a) by dissolving the phospholipids (I) and (II) and the lipophilic compound or mixture of compounds and, optionally, a lipid mentioned above from natural sources or according to method <p>) by dissolving the phospholipids (I ) and (II) and, optionally, a lipid mentioned above from natural sources in an organic solvent and cleaving the solvent. Suitable solvents are e.g. unsubstituted or substituted, e.g. halogenated, aliphatic or cycloaliphatic hydrocarbons, e.g. n-hexane, cyclohexane, methylene chloride or chloroform, alcohols, e.g. methanol or ethanol, lower alkane carboxylic acid esters or amides, e.g. acetic acid ethyl ester or dimethylformamide, or ethers, e.g. diethyl ether, tetrahydrofuran or dioxane, or mixtures of these solvents. The organic solvent is then split off by applying a vacuum, preferably a high vacuum, or by blowing off with an inert gas, e.g. nitrogen. The lyophilisate is prepared according to method a) by dissolving the phospholipids (I) and (II) and the lipophilic compounds or the mixture of compounds or according to method <p>) by dissolving the phospholipids (I) and (II) in an organic solvent according to the method described in TJS patent no. 4,311,712. Suitable solvents are in solid form together with the phospholipids (I) and (II) at the temperature of the lyophilization process and have a melting point of more than 0°C, e.g. glacial acetic acid, benzene or dioxane, especially tert-butanol. A homogeneous mixture can also be prepared by spray drying a solution of the phospholipids (I) and (II) and of the encapsulated material in an organic solvent having a low boiling point, such as chloroform. A powder is obtained by this process. ;The ratio between the phospholipid component (I) and the phospholipid component (II) in the homogeneous mixture is approx. 10 to 90 up to 50 to 50 mol-#. Preferably the ratio is 30 to 70 mol-#. The approximate ratio between the molar amounts of the encapsulated material (gamma-interferon) divided by the total amount of the phospholipids (I) and (II) is approx. 0.0001 to 0.1 to 1.0, preferably 0.005 to 0.01:0.1. ;The dispersion is carried out by mechanical shaking (stirring, shaking, "Vortex" mixer) of the aqueous phase to which, according to method a) the homogeneous mixture of the phospholipids (I) and (II) and the lipophilic compounds or the lipophilic mixture of compounds having pharmacological activity is added. According to method <p>), the aqueous phase containing the hydrophilic compounds or the mixture of compounds having pharmacological properties is added to the homogeneous mixture of the phospholipids (I) and (II). ;A mixture of small, large, unilamellar or multilamellar liposomes is formed spontaneously at high speed without the input of external energy. 0.1 to 40% by weight, preferably 2 to 20% by weight, of the homogeneous mixture relative to the total weight of the aqueous dispersion may be dispersed in the aqueous phase. Such dispersions can be further diluted to approx. 1 micromole of lipid per ml. Such liposome dispersions contain approx. 2.5 microliters of the aqueous phase per micromoles of the lipid. Acidic or basic aqueous dispersions are buffered to pH 7.0-7.8, preferably 7.2-7.4. Preferably, the dispersion is carried out in an aqueous phase having a pH of from 7.2 to 7.4. Method a) is preferred in the case that lipophilic, water-insoluble compounds are encapsulated in liposomes, e.g. lipophilic muramyl tripeptides. ;Procedure <p>) is preferred in the case that hydrophilic water-insoluble compounds are encapsulated in liposomes, e.g. cytarabine or cytostatic compounds such as triphosphamide. The production of the pharmaceutical preparations according to the present invention in the form of liposomes can also be carried out by other methods which are known in the art for the production of liposomes, e.g. by ultrasound treatment with ultrasound waves, with infusion procedures or reversed phase evaporation. ;The dispersion step is carried out at temperatures below 60°C, preferably at room temperature. In case of a potential thermal sensitivity of the encapsulated material, the dispersion is carried out under cooling and, optionally, under an atmosphere of an inert gas, e.g. nitrogen or argon atmosphere. The obtained liposomes can be made storage stable in the aqueous phase for up to several weeks or months after the addition of stabilizers, e.g. mannitol or lactose. The size of the liposomes that are formed depends, among other things, on the structure of the active ingredient and the lipid component, among other things, the ratio between the components and the concentration of these components in the aqueous dispersion. By e.g. to increase or decrease the concentration of the lipid components, aqueous phases are formed which have a high content of small or large liposomes. The separation of small liposomes from large liposomes is effected by means of conventional separation methods, e.g. sedimentation of the large liposomes in an ultracentrifuge, by gel filtration or extrusion through straight-pore filters. When centrifuging, e.g. from 5 to 60 minutes in a rotational field that gives rise to an inertial force equivalent to a gravitational field of 5000-40,000 x g, large liposomes are deposited at the bottom of the container, while small liposomes remain dispersed and can be decanted. After repeated centrifugation, complete separation of large liposomes from small liposomes is achieved. Liposomes are preferably separated from the aqueous phase in the event that the aqueous phase according to method p) contains non-encapsulated water-insoluble compounds or pharmaceutical agents. In particular, water-soluble antineoplastic agents, e.g. alkylating agents such as cyclophosphamide are separated by filtration, ultrafiltration, dialysis or by centrifugation to prevent any side effects caused by non-encapsulated compounds. The liposome fraction can be mixed with a carrier solution buffered to pH 7.2-7.4, e.g. isotonic, sterile sodium chloride solution buffered to pH 7.2-7.4.; Liposomes in the aqueous phase having a diameter greater than 6.0 x 10-* m, e.g. large mul ti lami Army liposomes, can be separated by gel filtration, e.g. with "Sepharose" or "Sephacryl" as carriers.

When extruding through straight-pore filters, e.g. membrane filters of the "Åcrodisc", "Nucleopore" or polycarbonate type which have a pore diameter of approx. 1.0 x 10"^ - 1.0 x 10"^ m at a pressure of approx. 0.1 to 1.5 bar and a filtration speed of approx. 20 ml/hour, a particularly uniform size distribution of the liposomes is achieved.

The formation of liposomes and their content in the aqueous phase can be detected in a manner known per se by using various physical analytical methods, e.g. by microscopy of freeze-fracture samples and thin sections in an electron microscope, by X-ray refraction, by dynamic light scattering, by mass determination of the filtrate in an analytical ultracentrifuge and, especially, by spectroscope!, e.g. in the nuclear magnetic resonance spectrum (^H, ^C Qg 31p).

Synthetic phospholipids of formula I are known. Their preparation as intermediates is described in European Patent Publication No. 56992.

The phospholipids of formula II are all known. Some of them are commercially available (Avanti, Fluka, Serva, Sigma).

The pharmaceutical preparations mentioned above, especially the antiphlogistic, antirheumatic, anti-marking agents, antifungals, antibiotics or antineoplastic agents are all known, see e.g. "MERCK Index", 10th edition.

The preparation of muramyl peptides of formula V is described in British Patent Specification No. 1,570,625, and in European Patent Publications 25,495 and 21,367. Immunomodulators of the lipopeptide type are also known, see European Patent Publication No. 114,787 and European Patent No. 330.

The production of purified, natural or recombinant, gamma interferon is described in European Patent Publications 63,482, 77,670, 83,540, 89,676, 95,350, 99,084, 110,044, 112,976 or 121,157, 1 British Patent No. 2,107,718 as well as in the International ( PCT) publications (WO) 83/04053 or WO 84/02129.

The production of purified interleukin 2 is described in European Patent Publication 106,179 and in US Patent No. 4,448,879.

The buffer solutions of pH 7.0 to 7.8 are preferably sterile phosphate buffer solutions based on the dihydrogen phosphate/hydrogen phosphate equilibrium (KH 2 PO 4 /Na 2 HPO 4 ). The preparation of these buffer solutions is described in standard reference books, e.g. "Hager's Handbuch der Pharmazeutischen Praxis", Springer Verlag, volume 1, pages 357-359. In particular, sterile, isotonic calcium-free buffer solution of pH 7.2 (Dulbecco) or Hank's balanced salt solution (M.A. Bioproducts, Walkersville MD, USA) is used.

The following examples illustrate the invention.

Example 1: a) Dissolve 84.70 mg (0.098 mmol) disodium N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxysuccinylamine and 168.10 mg (0.226 mmol) of 1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine in a sufficient amount of tert-butanol until both lipids are dissolved. The solution is filtered under sterile conditions over "Acrodisc" filter (2.0 x 10"^ m) and poured into a sterile bottle. This bottle is frozen at -45°C. A vacuum is applied to the frozen bottle and the solvent is removed until room temperature is reached The bottle is sealed under an inert gas atmosphere, eg argon atmosphere.

To this bottle containing a lyophilisate of the lipid components mentioned above, add 2.5 ml of a sterile, phosphate-buffered (pH 7.2-7.4), calcium-free sodium chloride solution (Dulbecco) containing doxorubicin at a concentration of 4 g/l with a sterile syringe. The bottle is then shaken for 10 minutes on a standardized laboratory shaker ("Vortex", speed 6) and placed in a centrifuge. After centrifugation in a gravity field of approx. 40,000 x g for approx. 60 minutes, the supernatant is decanted. The liposome dispersion is resuspended in 2.5 ml of 0.8556 sterile, phosphate-buffered (pH 7.2-7.4) sodium chloride solution (Dulbecco). The centrifugation and resuspension are repeated until the supernatant is free of doxorubicin. The liposome dispersion obtained is suitable for parenteral administration.

b) Production of sodium-N- fl. 2- di-(9- cis- octadecenoyl)- sn- glvcero- 3- phosphoethanol1- N- hydroxysuccinylamine

200.6 mg (270 micromoles) of 1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine are dried under nitrogen and placed under high vacuum for two hours. The dry lipid is dissolved in 10 ml of freshly distilled pyridine containing 54 mg (540 micromoles) of succinic anhydride. The reaction mixture is stirred for 2 hours under a nitrogen atmosphere at 55° C. The excess of pyridine is removed by evaporation under reduced pressure and the excess of succinic anhydride is hydrolyzed by suspending the residue in a mixture of chloroform-methanol-0.5856 aqueous sodium chloride solution (1:2: 0.8 volume/volume). The product is extracted by partitioning after adding one part by volume of chloroform and one part by volume of 0.5856 aqueous sodium chloride solution. The lower phase is washed three times with a mixture of chloroform-methanol-0.5856 aqueous sodium chloride solution (3:48:47 v/v). The solvent is removed by rotary evaporation and the residue is resuspended in chloroform. Rf (Merck silica gel 60 plates) ,375 (chloroform, methanol, water 65/25/4; DV: 254.6 nm (broad); mp 154°C.

Example 2: In a method similar to example 1, aqueous liposome dispersions are prepared containing 84.70 mg (0.098 mmol) disodium N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N -hydroxysuccinylamine and 168.10 mg (0.226 mmol) 1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phospho-ethanolamine and 0.1 mg up to 10 mg N-acetyl-L-muramyl- L-alanyl-D-isoglutamine sodium salt or 0.1 mg up to 10 mg N-acetyldesmethylmuramyl-L-alanyl-D-isoglutamine sodium salt or 1000 to 100,000 units of recombinant human immune gamma interferon obtained according to EP-A- 121,157 (Kyowa Hakko Kogyo Co.) or a combination of 1000 to 1,000,000 units of this recombinant human immune gamma interferon with 50-200 micrograms sodium-N-acetyl-D-muramyl-L-alanyl-D-isoglutamine or sodium -N-acetyldesmethylmuramyl-L-alanyl-D-isoglutamine.

Example 3: a) Dissolve 84.70 mg (0.098 mmol) of disodium N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxysuccinylamine and 168.10 mg (0.226 mmol) of 1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine in a sufficient amount of tert-butanol until both lipids are dissolved. The solution is filtered under sterile conditions over an "Acrodisc" filter (2.0 x 10~<7 >m) and filled into a sterile bottle. The bottle is rotated at 1750 rpm (revolutions per minute) and the solvent is removed in a steam cleaned, filtered (at a pressure of 1 bar) dry nitrogen. The bottle is evacuated in a high vacuum of 6.0 x 10-<2> mbar and is stable under an inert gas atmosphere of argon.

To this bottle containing a thin film of the lipid components discussed above is added 2.5 ml of a sterile, phosphate-buffered p(pH 7.2-7.4), calcium-free sodium chloride solution (Dulbecco) containing diclofenac at a concentration of 2 g/ l with a sterile syringe. The bottle is then shaken for 10 minutes on a standardized laboratory shaker ("Vortex", speed 6) and placed in a centrifuge. After centrifugation in a gravity field of approx. 40,000 x g for approx. 60 minutes, the supernatant is decanted. The liposome dispersion is resuspended in 2.5 ml of 0.85% sterile, phosphate-buffered (pH 7.2-7.4) sodium chloride solution (Dulbecco) and is suitable for parenteral administration.

Example 4; In a round-bottomed bottle, dissolve 0.1 mg of N-acetylmuramyl-L-alanyl-D-isoglutaminyl-L-alanine-2-(1,2-dlpalmitoy 1-sn-glycero-3-hydroxyphosphoryloxy)-ethylamide (preparation according to European Patent No. 25,495), 84.70 mg (0.098 mmol) disodium N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxysuccinylamine and 168.10 mg (0.226 mmol) of 1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine in a sufficient amount of sterile tert-butanol until all components are dissolved. The solution is filtered under sterile conditions over "Acrodisc" filter (2.0 x

10~<7> m) and filled into a sterile bottle. The bottle is rotated at 1750 rpm and the solvent is blown off in a stream of purified, filtered (at a pressure of 1 bar) dry nitrogen. The bottle is evacuated in a high vacuum of 6.0 x 10~<2> mbar and sealed under an inert argon gas atmosphere.

To this bottle containing a thin film of the components discussed above, 10 ml of a sterile, phosphate-buffered (pH 7.2-7.4), calcium-free, sodium chloride solution (Dulbecco) is added with a sterile needle. The bottle is then shaken for 10 minutes on a standardized laboratory shaker ("Vortex", speed 6). The resulting liposome dispersion is storage stable at 4°C and is suitable for parenteral administration.

Example 5: 0.1 mg of N-acetylmuramy 1-L-alanyl-D-isoglutaminyl-L-alanine-2-(1,2-dipalmitoyl-sn-glycero-3-hydroxyphosphoryloxy)ethylamide (prepared according to European Patent No. 25,495), 84.70 mg (0.098 mmol) disodium N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxysuccinylamine and 168.10 (0.226 mmol) of 1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine in a sufficient amount of sterile tert-butanol until all components are dissolved. The solution is filtered under sterile conditions over an "Acrodisc" filter (2.0 x 10"<7 >m) and filled into a sterile bottle. This bottle is frozen at

-45"C. A vacuum is applied to the frozen bottle and dissolve-

the igniting agent is removed until room temperature is reached. The bottle is sealed under an inert argon gas atmosphere.

To this bottle containing a lyophilisate of the components discussed above, 10 ml of a sterile, phosphate-buffered (pH 7.2-7.4), calcium-free sodium chloride solution (Dulbecco) is added with a sterile syringe. The bottle is then shaken for 10 minutes on a standardized laboratory shaker ("Vortex", speed 6). The resulting liposome dispersion is storage stable at 4°C and is suitable for parenteral administration.

Example 6: In a method analogous to examples 4 or 5, liposome dispersions are prepared containing 0.1 mg to 10 mg of N-acetylmuramyl-L-alanyl-D-isoglutaminyl-L-alanine-2-(1,2-dipalmitoyl-sn-glycero- 3-hydroxyphosphoryioxy)-ethylamide, 84.70 mg (0.098 mmol) disodium N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxysuccinylamine and 168.10 mg (0.226 mmol) of 1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine.

Example 7: By a method analogous to example 1, liposome dispersions containing 86.3 mg (0.098 mmol) of disodium N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N- hydroxysuccinylamine and 168.1 mg (0.026 mmol) 1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine and 0.1 mg up to 10 mg N-acetyl-L-muramyl-L-alanyl -D-isoglutamine sodium salt or 0.1 mg up to 10 mg N-acetyl-desmethyl-muramyl-L-alanyl-D-isoglutamine sodium salt.

Claims (2)

1. Process for the production of a pharmaceutical preparation in the form of an aqueous dispersion containing liposomes consisting of phospholipid components, characterized in that a) a homogeneous mixture consisting of a) a synthetic phospholipid with the formula where m means two or three, and R 2 independently means alkyl, alkenyl or acyl, each having 10 to 20 carbon atoms, X means a direct bond, C 1 -C 4 alkylene, C 2 -C 4 alkenylene, or C 1 -C 4 alkylene or The C2~C4 alkenyl substituted with hydroxy, or a pharmaceutically acceptable salt thereof, b) a synthetic phospholipid of the formula where R3 and R4 mean the acyl group of a saturated or an unsaturated carboxylic acid with 10 to 20 carbon atoms and 1-2 double bonds, in a ratio of phospholipid component (I) to phospholipid component (II) from approx. 10:90 up to 50:50, c) a lipophilic compound or a mixture of compounds with pharmacological activity, and, optionally, d) a lipid from natural sources selected from phosphatidylcholine, phosphatidylserine, phosphatidylinositol, phosphatidylglycerol, cardiolipin and cholesterol, and derivatives thereof, and a pharmaceutically acceptable carrier solution buffered to pH 7.0-7.8 and, where appropriate, pharmaceutically acceptable additives, are prepared and that the homogeneous mixture thus obtained is dispersed in an aqueous phase, or p) a homogeneous mixture consisting of synthetic phospholipids with formula I and II in the ratio indicated above and, optionally, a lipid mentioned above from natural sources is prepared and that the homogeneous mixture thus obtained is dispersed in an aqueous phase containing a hydrophilic compound or mixture of compounds with pharmacological activity, and if necessary, after carrying out either process step a) or p), the dispersion thus obtained is buffered to pH 7.0-7.8 and, if desired, non-encapsulated lipids and/or compounds with pharmacological activity are separated from the aqueous phase and/or the liposomes thus obtained are concentrated or separated from the aqueous phase. 2. Process according to claim 1, characterized in that a) a homogeneous mixture consisting of a) sodium or disodium-N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N- hydroxysuccinylamine or sodium or disodium N-[1,2-di-(9-cis-octadecenoyl)-sn-glycero-3-phosphoethanol]-N-hydroxyglutarylamine (I), b) 1,2-di-(9 -cis-octadecenoyl)-sn-glycero-3-phosphoethanolamine (II) and c) a lipophilic compound or a combination of compounds with pharmacological activity is prepared, or<p>) a homogeneous mixture consisting of synthetic phospholipids of formula (I) and (II) is prepared and the homogeneous mixture thus obtained is dispersed in an aqueous phase containing a hydrophilic compound or a mixture of compounds with pharmacological activity, and if desired, after carrying out one of the process steps a) or <p>), the thus obtained aqueous dispersion is buffered to pH 7.2-2.4.