DE4023241A1 - STABLE ACTIVE SUBSTANCE FORMULATION - Google Patents

Abstract

Described is a method of preparing stable formulations of active substance. In this method, a positively or negatively charged active substance is dissolved in molar amounts in an organic solvent with an oppositely charged lipid, optionally adding to the active-substance/lipid mixture thus obtained a lipid forming a double-layer membrane, also optionally adding other membrane components which increase the mechanical and/or chemical stability of colloidal particles when the particles are in contact with biological fluids, and finally removing the solvent.

Description

The present invention relates to new stable active substance formulations and their manufacture.

Liposomes consist of membrane bilayers that are in the form of tiny cavities spheres an aqueous interior from the surrounding water phase separate. Liposomes form spontaneously when dispersing suitable amphi phile lipids in aqueous systems. Depending on the preparation Conditions can be liposomes with diameters from about 20 nm to about 10 microns are formed. Basic work on this was carried out by i.a. Bangham (J. Mol. Biol. 13 (1965), pp. 238-252) and Papahadjopoulos (Biochem. Biophys. Acta 135 (1967), 624-638).

The spherical closed liposomes enable encapsulation water-soluble compounds, e.g. B. Active ingredients in the aqueous interior. In a number of patents, such liposomal active ingredient encapsulations described lungs (including: US 39 93 754, US 41 45 410, US 42 35 871). There the active ingredient is in an encapsulated form, but it often succeeds existing local toxicity (e.g. vein irritation after IV injection) favorable to influence. However, such liposomal encapsulations prepare problems both in manufacturing and in storage. So can with the usual methods rarely more than 30% of those used Amount of active ingredient are encapsulated. The non-encapsulated active ingredient must always in a further process step by gel filtration or Ultrafiltration can be removed. Finally, the storage stability of such preparations limited by the premature active ingredient reflux.

The invention relates to a method for producing stable active Substance formulations, characterized in that an active ingredient which can form positively or negatively charged salts, in molar amounts with an oppositely charged lipid in an organic solvent dissolves, optionally the active ingredient-lipid mixture thus obtained double-layer membrane-forming lipid and optionally another membrane components that affect the mechanical and / or chemical stability of the increase colloidal particles on contact with biological fluids, added and the solvent removed. The subject of the invention are further the active ingredient formulations thus obtained.

The process according to the invention can be carried out using non-neutral active ingredients perform, i.e. Active substances that can form cations or anions. Such are, for example, basic active ingredients such as amonafide, mitonafide,  Emopamil, N-acetylamonafide, anipamil and acidic ingredients such as acetyl salicylic acid, ibuprofen, diclofenac, penicillins, prostaglandin E₁.

The lipid used for the preparation of the formulations must be a Active substance have opposite charge, so that salt formation with the Active ingredient is possible. All are suitable for binding basic active ingredients acidic lipids that are capable of salt formation through proton release. Examples include saturated and unsaturated, preferably natural occurring fatty acids with more than 11 carbon atoms such as B. oleic acid, Double-layer membrane-forming amphiphiles containing carboxyl groups (cf. EP 3 31 092) such as N- (4-oxobutanoic acid) -L-aspartic acid dioleylester (= lipid No. 1) and phosphatidic acids such as. B. 1,2-Dihexadecyl-glycerol-3-phosphate (= Lipid No. 3). Basic agents are suitable for binding acidic active ingredients Lipids. Examples include: stearylamine and phosphate diylethanolamine.

Aprotic solutions are suitable as solvents for salt formation agents in which the lipids are readily soluble, such as dichloromethane, ethanol, Tetrahydrofuran, isopropanol.

Active ingredient and lipid are generally in a molar ratio of about 1: 1 implemented with each other, regardless of how many charges the active ingredient ion occupied.

It is generally necessary to extend the lipid-active ingredient salts widely Res lipid add a homogeneous distribution of the solid lipid effect mixture when dispersed in aqueous systems. Suitable Lipids for this are double-layer membrane-forming lipids like the natural one occurring phospholipids or synthetic double layer membrane forming lipids such as N- (4-oxobutanoic acid) -L-aspartic acid-dioleylester- Potassium salt (= Lipid No. 2) or cholesterol hemisuccinate salts. This sub are punched, provided that the lipid-active salt is not already in aqueous systems can be distributed homogeneously, added in amounts that about 0.5 to 2 times the amount of lipid used for salt formation be.

As components of the cell membrane, which the mechanical stability of Par Increase particles, especially cholesterol and its esters should be mentioned, which the stability of the drug carrier especially when in contact with biological liquids, e.g. B. plasma, increase.

Are the substances used to make the new molds easily oxidizable, so is the addition of radical scavengers, such as tocopherols or carotene, appropriate.  

The new active ingredient formulations are generally successful at room temperature.

The active ingredient formulations obtained according to the invention are suitable for Manufacture of drugs.

The new process has the following advantages:

• 1. Poorly soluble active ingredients can be dissolved in significantly higher concentrations in aqueous systems by ionic binding to colloidal carriers (e.g. liposomes or emulsions) than by simple dissolution. So z. B. liposome solutions or emulsions with an ionic bond with the active ingredient Mitonafide in concentrations of 6 mg / ml (based on the active ingredient base) while z. B. even the mitonafide hydrochloride can only be dissolved in concentrations of up to 0.4 mg / ml in the same buffer system.
  So z. B. also produce liposome solutions or emulsions with ammonia fide in concentrations of 6 mg / ml at physiological pH values from 6 to 7, in which the active ingredient largely flocculates in the form of the base without using the method according to the invention. Anipamil base (solubility of the HCl salt in water at room temperature <5 ppm) can also be dissolved at physiological pH values in a concentration of 8 mg / ml by using the method according to the invention.
• 2. The manufacture of relatively concentrated drug solutions is therefore also possible at physiological pH values, which are normally used for Precipitation of many active ingredients. This makes it difficult to dissolve Active ingredients such. B. by using micelle-forming detergents in solubilize aqueous systems. The common ones Solubilizers (e.g. Tween 80®, Cremophor EL®) have to do this but used in concentrations that are undesirable Side effects such as sensitization, histamine release, hemolysis cause. Double-layer membrane-forming lipids, however, such as. B. the Naturally occurring phospholipids show these side effects not because they are an integral part of every cell membrane.
• 3. The active ingredient solutions according to the invention show significantly improved Compatibility with the purely aqueous active ingredient solutions same molar dose of active ingredient. This is particularly important for them intravenous administration.  
• 4. The production methods of the liposome solutions according to the invention are significantly simplified compared to the known methods, since a Encapsulation or the like is not necessary and therefore complex Process steps (gel filtration, ultrafiltration, etc.) are omitted. By dispensing with the encapsulation, high efficacy can also be achieved achieve substance contents. Salt formation of the active ingredient also takes place with the lipid in the formulation. A previous preparation or Isolation of these salts is not necessary.
• 5. Stability problems during storage are minimized because the Ver encapsulation known and disruptive stability-limiting active ingredient Efflux from the liposomes does not occur. Sensitive drug solution gene can with the known methods after the addition of antifreeze agents (sucrose, trehalose, glucose) etc. can be lyophilized.

The following examples illustrate the invention:

Example 1

120 mg amonafide base (0.424 mmol), 310.6 mg lipid No. 1 (0.424 mmol), 346.8 mg of Lipid No. 2 (0.444 mmol) and 24.0 mg of Tocopherol (0.056 mmol) were clearly dissolved in a little dichloromethane. The solvent was then eats (last in vacuum) removed. The residue was treated with 20 ml of buffer solution (NaCl 8.30 g / l + EDTA-2Na 0.50 g / l, adjusted to pH 7.5) ver sets and sonicated at 45 ° C for 20 min. The still warm solution filtered then through a 0.2 micron syringe filter and then stored in the cool closet. The pH of the solution was 7.1.

Example 2

The preparation was carried out as in Example 1, but he was sonicated followed in 9.25% sucrose solution. After sterile filtration the samples in 1 ml protions in 2 ml vials lyophilized. The particles sizes were 104 nm before lyophilization with mean, after Lyophi lization in the redisperse after adding 1 ml of water by shaking samples averaged 119 nm (measured with photon correlation spectro scopie).

Example 3 (comparative example)

30.0 mg amonafide base (0.111 mmol) and 162.7 mg (0.222 mmol) Lipid No. 1 were dissolved in a little dichloromethane. After completely removing the loosening by means of ultrasonication at 40 ° C in 10 ml of buffer solution (0.9% NaCl + 10 mmol phosphate, pH 7.2). After 30 min sonication was obtained  a cloudy solution, from which when cooling to room temperature inside of 20 min the substances flocculated.

Example 4

The preparation was carried out as in Example 3, but the mixture of substances ent also held 223.1 mg (0.290 mmol) of Lipid # 2 and 49.9 mg (0.128 mmol) cholesterol. After 20 min of ultrasound, a clear Obtain opalescent solution, even after cooling after 3 months No flocculation occurred (refrigerator storage).

Example 5

The preparation was carried out as in Example 4, but only with 81.35 mg (0.111 mmol) Lipid No. 2. The storage stability was that of Example 4 identical.

Example 6

The preparation was carried out as in Example 4, but with 30.0 mg (0.096 mmol) Mitonafide base and 70.2 mg (0.096 mmol) of lipid No. 1. The storage stability was identical to that of Example 4.

Example 7

100.0 mg (0.200 mmol) S-Emopamil and 219.2 mg (0.299 mmol) Lipid No. 1 were with 397.8 mg (0.509 mmol) of lipid No. 2 in a little dichloromethane solved. After the solvent had been completely removed, the residue became 20 ml buffer solution (0.9% NaCl + 10 mM phosphate, pH 7.2) at 45 ° C for 15 min ultrasound. The still warm solution was passed through a sterile 0.45 µm Syringe filters were filtered into sterile polypropylene tubes and stored then without precipitation / flocculation for 6 months at room temperature.

Example 8

60.0 mg (0.223 mmol) of Amonafide base and 163.5 mg (0.223 mmol) of Lipid No. 1 were treated with 3000 mg of soybean oil (Sigma), 173.4 mg (0.222 mmol) of lipid No. 2 and 12.0 mg (0.028 mmol) of tocopherol mixed and aqueous in 20 ml Ultrasonically exposed to glycerol solution (2.6%) at 60 ° C for 15 min. One was get homogeneous emulsion, the particle sizes even after 3 months Storage time (refrigerator) was still 88% less than 1 µm. When waiving lipid # 1 used for salt formation, however, did not become stable Receive emulsion.  

Example 9

185.0 mg (0.237 mmol) of Lipid No. 2 were mixed with 51.0 mg (0.070 mmol) of Anipa mil base dissolved in a little dichloromethane. After removing the solvent in the The residue was vacuumed with 10 ml of buffer solution (0.9% NaCl + 10 mM Phosphate, pH 7.2) was added and sonicated for 20 min at 45 ° C. The solution was then filtered warm through a 0.45 µm syringe filter and then stored in the refrigerator for 1 year without precipitation.

Example 10

120.0 mg (0.424 mmol) amonafide base, 275.1 mg 1,2-dihexadecyl-glycerol 3-phosphoric acid (Lipid No. 3) (0.444 mmol) and 346.8 mg (0.444 mmol) lipid No. 2 were dissolved in a little dichloromethane. After removing the loosening the residue was removed in vacuo with 20 ml of phosphate buffer (0.142 mM; pH 7) sonicated for 20 min at 45 ° C. After filtration through a 0.45 µm Syringe filters stored the solution in the refrigerator for 3 months without precipitation closet.

Example 11

173.4 mg (0.222 mmol) of lipid No. 2 was combined with 163.0 mg (0.222 mmol) of lipid No. 1 and 72.2 mg (0.222 mmol) N-acetylamonafide base in a little dichlor dissolved methane. After removal of the solvent in vacuo, after Zu administration of 10 ml of buffer solution (phosphate 0.142 mM; pH 7.0) for 20 min at 45 ° C. The solution was then warmed by a The 0.45 µm syringe filter was filtered and then stored for 3 months without precipitation in the fridge.

In the absence of the lipid No. 1 used for salt formation and below simultaneous increase in the amount of lipid No. 2 to 346.8 mg (0.444 mmol) no stable solutions could be obtained.

Example 12

120.0 mg amonafide base (0.424 mmol), 119.8 mg oleic acid (0.424 mmol), 401.0 mg of Lipid No. 1 (0.514 mmol), 6.0 g of soybean oil and 24.0 mg of tocopherol (0.056 mmol) was added with 40 ml glycerol solution (2.6% in water) and sonicated for 20 min at 60 ° C. This resulted in a homogeneous emulsion preserved, which remained stable even after 1 month of refrigerator storage.

The omission of the oleic acid used for salt formation was opposed no stable emulsion obtained.

Claims (3)

1. A process for the preparation of stable active ingredient formulations, characterized in that an active ingredient which can form positively or negatively charged salts is dissolved in molar amounts with an oppositely charged lipid in an organic solvent, if appropriate the active ingredient / lipid mixture thus obtained a double-layer membrane-forming lipid and optionally other membrane components that increase the mechanical and / or chemical stability of the colloidal particles upon contact with biological fluids, and remove the solvent. 2. Active ingredient formulations according to claim 1. 3. Use of the active ingredient formulations according to claim 1 for the manufacture provision of medicines.