EP1206670B1 - Lyophilization method - Google Patents

Description

The present invention relates to a novel process for freeze-drying (Lyophilization).

Freeze-drying is an important stabilization process hydrolysis-sensitive and thermolabile preparations, as well as materials of biological origin, which are dried under mild conditions should. With the help of freeze-drying materials can be used without major Changes and losses of biological activity are dried. One positive aspect of freeze-drying is that the dried, "lyophilic" Products due to their porous structure and very large specific surface area are very quickly reconstituted and their original properties in solution accept again. Therefore, freeze drying is preferred therapeutic sera, blood products, biologically active substances (hormones, Vitamins, enzymes, medicines), food preparations and flavorings. to Freeze-drying liquid and semi-solid water-containing preparations are suitable, e.g. Solutions, emulsions and suspensions.

• Abkühlung und Kristallisation des Lösungsmittels in der Zubereitung bei Atmosphärendruck.
• Haupttrocknung, d.h. Sublimation des kristallisierten Lösungsmittels.
• Nachtrocknung, d.h. Verdampfung von nicht kristallierten Lösungsmittelanteilen.

Drying from the frozen state combines the advantages of freezing and dehydration at low temperature and is generally carried out as follows:

• Cooling and crystallization of the solvent in the preparation at atmospheric pressure.
• Main drying, ie sublimation of the crystallized solvent.
• Post-drying, ie evaporation of uncrystallized solvent fractions.

The two drying steps differ in principle: In the Primary drying (primary drying) is the sublimation of the frozen Solvent under reduced pressure. At optional drying (Secondary drying), the evaporation of non-frozen solvent at reduced pressure and at elevated temperature.

In the known from the prior art, the method to be dried Preparations in containers, e.g. so-called vials, frozen at atmospheric pressure and The temperature of the product is set to one for the start of the main drying appropriate value set.

The freezing (crystallization) is followed by the main drying, in which, under reduced pressure, the frozen solvent is transferred from the solid to the gaseous state, ie sublimed. The energy that is consumed in the sublimation is replenished, for example, via heatable control panels. During the main drying, the frozen preparation must not heat above its melting point. The main drying can be followed by a final drying, in which non-frozen solvent is removed at elevated temperature and reduced pressure. These are solvents which can be adsorbed on the solid matrix or enclosed in amorphous areas, for example.
Crystallization in the present application is understood as meaning the freezing (solidification) of the solvent fraction in the preparation. In the present application, preparation is understood to mean any type of material which is suitable for freeze-drying.

The temperature profile during the freeze-drying can be controlled by suitable devices. In particular, temperature-controlled control panels are known to those skilled in the art. In this process, the control panels can be brought to the desired glass transition temperature both after loading (cooling variant A) and before loading (cooling variant B). It is also possible to pre-temper the plates and / or the preparation on the plates to a temperature above the actual glass transition temperature to ensure a temperature equalization of the individual vials, or to minimize the cooling time to freezing. This is followed by the actual freezing with further lowering of the plate temperature (cooling variant C).
The variants AC describe the freezing on shelves. Other known methods are freezing methods in cold baths and rotating vessels (shell freezing, spin freezing) or by spraying devices; they differ in principle from the methods described above.
Usually, the preparations to be dried are aqueous systems. In principle, other solvents or mixtures thereof with aqueous systems can be used, such as carboxylic acids (eg glacial acetic acid), dimethyl sulfoxide (DMSO), ethers (eg dioxane), dimethylformamide or alcohols (eg t-butanol).

The various conventional types of freezing and freeze-drying are well described in relevant textbooks, such as lyophilization, vinegar, Oschmann, Scientific Verlagsgesellschaft Stuttgart mbH, 1993; Pages 15-29, freeze-drying, Georg-Wilhelm Oetjen, VCH Verlag, 1997; Page 3-58 and Freeze Drying, Athanasios I. Liapis, in: Handbook of Industrial Drying, ed. By AS Mujumdar, Montreal, pp. 295-326.

In addition, US 3,233,333 describes a process for freeze-drying of meat and other foods that contain cellular structures, but by performing in four steps, the physical quality is not is impaired and taste, smell and texture practically remain unchanged.

All freezing methods have in common that if the preparation is suitable for the Freezing a tempering step (thermal treatment or annealing) can be done. This annealing step serves to crystallize amorphous solidified solids and not to require frozen solvent and thus increased crystallinity and to achieve a reduced residual moisture. To carry out the frozen preparation heated to a temperature above that Glass transition temperature (Tg ') of the amorphous solidified solution and below the Melting point of the solution is. The amorphous phase, which usually contains high levels of contains uncrystallized solvent, goes from the glass state in the rubbery Condition over and the mobility of molecules is increased. Consequence is the Formation of nucleoli that grow into crystals (so-called eruptive recrystallization) and the attachment of solvent molecules to existing ones Solvent crystals.

The annealing process is also known from the literature. Descriptions of the tempering process can be found in The Lyophilization of Pharmaceuticals: A Literature Review, NA Williams and GP Polli, Journal of Parenteral Science and Technology, (1984 Mar-Apr) 38 (2) 48-59, Basic Aspects and Future Trends in the Freeze Drying of Pharmaceuticals , L. Rey, Develop. biol. Standard, Vol. 74, (Karger, Basel, 1991), pp. 3-8 and Fundamental Aspects of Lyophilization, L. Rey, Research and Development in Freeze-Drying, ed. By L. Rey, Paris, 1964, 24-47.

Those prepared by the prior art freeze-drying methods Lyophilisates usually have a high flow resistance, which is the Escape of the gaseous solvent impeded. In addition, that can Crystallize the dissolved ingredients partially to completely omit, so that Products are obtained, which are partly to completely amorphous. The consequences, the This may result in mechanical damage to the product cake by the escaping solvent vapor stream and thereby potential Product loss, as well as collapse and thawing phenomena during drying. In addition, by the end user especially to pharmaceutical and Food preparations also made aesthetic demands, so that strong Damage is not desired.

An object of the present invention was therefore a process for freeze-drying to find, with which lyophilizates can be produced, which are the above do not exhibit said problematic properties and therefore easier are manageable.

Phase 1:

Erniedrigen des Druckes in der Trocknungskammer bis zum Einsetzen einer sichtbaren Kristallisation des Lösungsmittels bei einer Temperatur in der Trocknungskammer, die oberhalb des Festpunktes der Zubereitung liegt.

Phase 2:

Erniedrigen der Temperatur in der Trocknungskammer auf eine Temperatur, die unter dem Festpunkt der Zubereitung liegt oder mit diesem identisch ist, bis zum Abschluß der Kristallisation des Lösungsmittels.

Phase 3:

Sublimation des gefrorenen Lösungsmittels mittels vermindertem Druck.

Surprisingly, it has now been found that mechanically more stable lyophilisates are obtained by carrying out the process for freeze-drying as follows:

Phase 1:

Lowering the pressure in the drying chamber until onset of visible crystallization of the solvent at a temperature in the drying chamber that is above the fixed point of the preparation.

Phase 2:

Lowering the temperature in the drying chamber to a temperature below or equal to the melting point of the formulation until the crystallization of the solvent is completed.

Phase 3:

Sublimation of the frozen solvent by means of reduced pressure.

The fixed point of the preparation in the present application is the one Temperature understood in which the solvent in the preparation in the solid State of aggregation passes.

• durchgezogene Linie, Quadrate = Mannitol
• durchgezogene Linie, Kreise = Saccharose
• unterbrochene Linie, Rauten = Natriumchlorid
• unterbrochene Linie; Kreise = Glycin
• unterbrochene Linie, Dreiecke = Maltose
• Quadrat auf der Ordinate = Lösungsmittel Wasser

According to the invention, the pressure in the drying chamber is initially reduced at a temperature in the drying chamber which is above the fixed point of the preparation to a pressure below the atmospheric pressure (according to FIG. 1). This results in superficial cooling of the preparation by evaporation and partial crystallization of the solvent at the surface (phase 1). In a preferred embodiment, the pressure in aqueous solutions is 0.1 to 6 mbar, in particular 0.2 to 3 mbar. For various preparations, this pressure p is plotted in the drying chamber (measured with a capacitance gauge) as a function of the concentration c (in mol / L) in FIG. The values for various aqueous preparations were shown as follows:

• solid line, squares = mannitol
• solid line, circles = sucrose
• broken line, diamonds = sodium chloride
• broken line; Circles = glycine
• broken line, triangles = maltose
• Square on the ordinate = solvent water

This pressure reduction can e.g. be carried out at room temperature. In In another embodiment, the preparations are before or during the Lowering the pressure to a temperature between room temperature and the Fixed point of the preparation is, pre-tempered. This pre-tempering (e.g. Stellplatten) further ensures that the cooling devices, the z.T. low Have cooling rates, in a short time to the desired crystallization temperature, i.e. can be brought into the range of the fixed point of the preparation. critical is that this Vortemperierung not for the crystallization of the solvent leads.

Have crystals, e.g. in the form of a water / ice mixture or on formed the surface floating ice layer, the pressure in the Drying chamber raised to ambient pressure and the temperature in the drying chamber for crystallization at or below the fixed point of Preparation (phase 2). It is also possible the pressure during to keep the crystallization reduced; this has no relevant effects the crystallization of the solvent. In principle, every crystallization is Temperature suitable, which is below the benchmark of the preparation or with him is identical. In a preferred embodiment, the temperature is Crystallization in aqueous solutions between -60 ° C and 0 ° C.

After crystallization, the preparation is optionally to the final Temperature brought to the beginning of drying. This temperature is from the present product and the vapor pressure curve of the solvent from Pressure to be used in the main drying depends. In a preferred embodiment, this temperature is in aqueous solutions -60 ° C to 0 ° C.

Then the main drying follows. This runs in principle as with the Method according to the prior art. In a further embodiment has the process additionally a post-drying phase (phase 4) following the Main drying on. However, this is in the presence of an annealing phase (phase 2a) in not necessary in some cases.

According to another embodiment, an annealing process such as connected above. This annealing process is referred to as Phase 2a referred. Annealing provides products with higher crystallinity and lower residual moisture after the main drying and shortens the post-drying or makes them redundant.

The inventive method will be explained in more detail by the figures 2 to 7 Here are the temperature (T) and the pressure (p) in millibars (mbar) in the Drying chamber applied against the time t, with the temperature as solid, the pressure is shown as a broken line. For better Clarification of the method, the figures always show embodiments Main and post-drying.

Fig. 2 shows a conventional manufacturing method of the prior art.

FIG. 3 shows a conventional manufacturing process with annealing step after State of the art.

4 shows the method according to the invention with pressure reduction (phase 1), Crystallization (phase 2), annealing step (phase 2a) and subsequent main and Post-drying (phases 3 and 4).

5 shows the method according to the invention with pre-tempering and pressure reduction (Phase 1), crystallization (phase 2) and subsequent main and Post-drying (phases 3 and 4).

Fig. 6 shows the inventive method with pre-tempering and Pressure reduction (phase 1), crystallization (phase 2), annealing step (phase 2a) and subsequent main and post-drying (phases 3 and 4).

7 shows the method according to the invention with pressure reduction (phase 1), Crystallization (phase 2), followed by main and post-drying (phases 3 and 4).

The lyophilizates which can be prepared by the process according to the invention have a improved structure cohesion and are escaping from the Steam flow, even at increased sublimation rates, less mechanically strong damaged as lyophilisates by methods of the prior art getting produced. They show less pronounced collapse phenomena.

The residual moisture contents which can be achieved by the process according to the invention are in principle comparable to those obtained by freeze-drying to the state technology can be achieved (see Tab. 3)

Preparations with or without so-called scaffold formers are suitable for use in the process according to the invention. With such scaffold formers, a porous scaffold or a matrix can be produced during freeze-drying. Freeze-drying products which are prepared using scaffold formers or other substances which are suitable as scaffolding agents on account of their physicochemical properties are preferred. Particularly preferred are freeze-drying products which are prepared using skeleton formers selected from the classes of compounds amino acids, carbohydrates (mono-, disaccharides, sugar alcohols, oligosaccharides, polysaccharides), peptides, polymeric compounds and salts. Most preferred are those prepared using frameworks selected from the group consisting of mannitol, sucrose, maltose, glycine and sodium chloride. List of preferred scaffold builders amino acids glycine alanine aspartic acid peptides gelatin collagen albumin monosaccharides glucose lactose disaccharides maltose sucrose trehalose oligosaccharides cyclodextrins maltodextrins polysaccharides Starch and starch derivatives Cellulose and cellulose derivatives polymers polyvinylpyrrolidones polyethylene glycols salts sodium chloride calcium carbonate sugar alcohols mannitol sorbitol xylitol

For the process according to the invention, a multiplicity of solvents are used Question. For the sake of better understanding, the description will be overwhelming treated aqueous systems. However, the invention also relates expressly nonaqueous systems. Preference is given to using aqueous solutions.

embodiments Durchführungsschemta of freeze-drying with the invention "Vacuum-induced" freezing and state-of-the-art processes technology Starting materials, materials and equipment

• There was prepared a 5% aqueous solution of mannitol and a 0.2μm membrane filter sterile filtered.
• The solution was filled to 3 ml in 10R tube glass vials and lyophilization stoppers placed.
• For the freeze-drying, the filled vials were placed in a freeze drier of the company Kniese (footprint 0.6 m ² ).

Execution:

• The solution was pre-tempered on the control plates at + 10 ° C.
• Thereafter, the chamber pressure was lowered to 0.65 mbar (for parameter selection see Fig. 1).
• After reaching the pressure value of 0.65 mbar and inserting the partial Freezing at the product surface was vented to ambient pressure and at the same time bringing the control panels to a temperature e.g. for mannitol May be -7.5 ° C.
• The products were each held at the respective temperature for 1 hour and then cooled to -40 ° C (freezing variant III).
• The freezing was followed by a main drying for 8 hours at + 40 ° C and 1.6 mbar without drying. The observed abnormalities were in Tab. 2.
• As a reference, appropriate solutions were used which did not correspond to the subjected to vacuum-induced freezing (freezing variant III) according to the invention but with 2K / min to -40 ° C (freezing variant I) and occurring Frozen undercooling or fast in the cold bath to -60 ° C (freezing variant II) and then lyophilized under the same conditions were.

Undesirable changes and damage of the product cake during different freezing processes and main drying with plate temperature (+ 40 ° C). A weighting of severity and frequency of damage occurred was indicated by (-), i. none, (+) weak, (++) strong and (+++) very strong. freezing 5% mannitol solution Variant I Surface torn open on many samples, lyophilisates on the vial bottom collapsed (++) Variant II Many lyophilisates on the vial bottom collapse; Product cake torn apart (+++) Variant III (-) Implementation Schemes of freeze-drying with the invention "Vacuum-induced" freezing (variant III), as well as with the invention "Vacuum-induced" freezing with subsequent annealing process (Variant IV) Starting materials, materials and equipment

• It was prepared an aqueous, 2% solution of mannitol and a 0.2μm membrane filter sterile filtered.
• The solution was filled to 3 ml in 10R tube glass vials and lyophilization stoppers placed.
• For freeze-drying, the vials were placed in a freeze drier from the company Kniese (footprint 0.6 m ² ).

Execution:

• The solutions were pre-tempered on the control plates at + 10 ° C.
• Thereafter, the chamber pressure was lowered to 0.65 mbar (for parameter selection see Fig. 1).
• After reaching the pressure value of 0.65 mbar and inserting the partial Freezing at the product surface was vented to ambient pressure and at the same time bringing the control panels to a temperature e.g. for mannitol May be -7.5 ° C.
• The products were each held at the respective temperature for 1 hour and then cooled to -40 ° C (variant III).
• For variant IV, samples were taken following the procedure of Variant III heated to -3 ° C, annealed at this temperature for 4 hours and then cooled to -40 ° C.
• As a reference, samples were frozen to -40 at a cooling rate of 2 K / min. The samples undercooled. (Variant I).
• On freezing to variant I, III or IV, followed by a main drying over 20 hours at -10 ° C and 0.2 mbar and a final drying at + 40 ° C and 0.2 mbar for 2 hours.

After freeze-drying, the residual moisture of the lyophilisates was additionally determined by a Karl Fischer titration: Dependence on residual moisture and drying time of the freezing process. Mannitol 2% Residual moisture [%] Sublimation duration [min] vacuum-ind. Freezing (variant III) 1.41 810 vacuum-ind. Freezing and tempering process (variant IV) 0.18 864 Reference (variant I) 0.86 1066

Claims (9)

1. Method for freeze-drying preparations in a drying chamber comprising the steps cooling the preparation and crystallizing the solvent and sublimation of the frozen solvent by means of reduced pressure, in which the method is carried out as follows:

   phase 1:

   reducing the pressure in the drying chamber until the onset of a visible crystallization of the solvent at a temperature in the drying chamber which is above the solidification point of the preparation,

   phase 2:

   reduction of the temperature in the drying chamber to a temperature which is below the solidification point of the preparation or is identical to this, until completion of crystallization of the solvent,

   phase 3:

   sublimation of the frozen solvent by means of reduced pressure.

2. Method according to Claim 1, characterized in that an aqueous solution is present.
3. Method according to Claim 1, characterized in that the pressure in phase 1 is reduced to 0.1 to 6 mbar in the case of aqueous preparations.
4. Method according to Claim 3, characterized in that the pressure in phase 1 is reduced to 0.2 to 3 mbar.
5. Method according to Claim 1, characterized in that the temperature in phase 2 is set to -60 to 0°C in the case of aqueous preparations.
6. Method according to Claim 1, characterized in that a tempering step (phase 2a) is carried out, in which a temperature is set which is above the glass transition temperature (Tg') of the amorphously solidified solution and below the solidification point of the preparation.
7. Method according to Claim 1, characterized in that the temperature at the start of the primary drying is set to -60 to 0°C in the case of aqueous preparations.
8. Method according to Claim 1, characterized in that a cake-forming agent is used.
9. Method according to Claim 1, characterized in that the cake-forming agent is mannitol, sucrose, maltose, glycine or sodium chloride.

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