CN106880592B - Trabectedin nanoemulsion and preparation method thereof - Google Patents

Trabectedin nanoemulsion and preparation method thereof Download PDF

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CN106880592B
CN106880592B CN201611144699.4A CN201611144699A CN106880592B CN 106880592 B CN106880592 B CN 106880592B CN 201611144699 A CN201611144699 A CN 201611144699A CN 106880592 B CN106880592 B CN 106880592B
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nanoemulsion
oil
composition
solubilizer
group
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CN106880592A (en
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孙琼
邵长丽
简彤
刘凯
史海贤
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Jiangsu Hengrui Medicine Co Ltd
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/4995—Pyrazines or piperazines forming part of bridged ring systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/08—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing oxygen, e.g. ethers, acetals, ketones, quinones, aldehydes, peroxides
    • A61K47/10—Alcohols; Phenols; Salts thereof, e.g. glycerol; Polyethylene glycols [PEG]; Poloxamers; PEG/POE alkyl ethers
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/08—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing oxygen, e.g. ethers, acetals, ketones, quinones, aldehydes, peroxides
    • A61K47/14—Esters of carboxylic acids, e.g. fatty acid monoglycerides, medium-chain triglycerides, parabens or PEG fatty acid esters
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/24—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite containing atoms other than carbon, hydrogen, oxygen, halogen, nitrogen or sulfur, e.g. cyclomethicone or phospholipids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
    • A61K47/26—Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/44—Oils, fats or waxes according to two or more groups of A61K47/02-A61K47/42; Natural or modified natural oils, fats or waxes, e.g. castor oil, polyethoxylated castor oil, montan wax, lignite, shellac, rosin, beeswax or lanolin
    • 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
    • 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
    • A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
    • A61K9/19—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles lyophilised, i.e. freeze-dried, solutions or dispersions

Abstract

The invention relates to a trabectedin nanoemulsion and a preparation method thereof. In particular, the nanoemulsion in the trabectedin nanoemulsion comprises a solubilizer and an emulsifier, and improves the stability of the trabectedin in the preparation and storage processes of a pharmaceutical preparation.

Description

Trabectedin nanoemulsion and preparation method thereof
Technical Field
The invention relates to a Trapertidine nanoemulsion and a preparation method thereof.
Background
Tribetidine (ecteinascidin 743, abbreviated as ET-743 or ET743) is a tetrahydroisoquinoline alkaloid isolated from marine tunicates and has the following specific structure. The antitumor activity of the compound is mainly realized by alkylating and modifying DNA through self-forming imine positive ions or similar intermediates (chem.Rev.2002,102,1669), and ET-743 has inhibitory activity on breast cancer, non-small cell lung cancer, colon cancer, kidney cancer, ovarian cancer, endometrial cancer, melanoma, osteosarcoma, prostate cancer and the like, and is a novel anticancer drug capable of being combined with DNA. Ecteinascidin-743 was developed by the company Spanish FammaMa (PharmMar) and approved for marketing in the European Union in 2007 as a rare drug for the treatment of soft tissue sarcoma in patients with advanced soft tissue sarcoma who failed traditional chemotherapy.
Figure BDA0001178611340000011
ET-743 is a complex compound with limited water solubility that would increase the solubility of ET-743 in water by adjusting the pH of the solution to 4. However, the stability of ET-743 in biocompatible forms and formulations is difficult to predict and achieve. Therefore, it is a challenge for those skilled in the art to prepare a stable and effective ET-743 formulation. The main impurities in Trabectet are ET-743, ET-745, ET-759B and ET-789A (Synthesis of Natural Ecteinascidins (ET-729, ET-745, ET-759B, ET-736, ET-637, ET-594) from Cyanossafracin B, Journal of Organic Chemistry 200368 (23) page 8859-8866), wherein the impurity ET-701 is the main impurity in the freeze-drying process and storage of the preparation, and is a hydrolysate; the impurity ET-745 is a reduced product of the trabectedin, and the structures of the impurity ET-745 are respectively as follows:
Figure BDA0001178611340000021
WO0069441 discloses a sterile lyophilized formulation comprising ET-743, mannitol and phosphate buffer. The lyophilized formulation can be reconstituted and diluted for intravenous injection. However, the formulations are not stable for long term storage in the refrigerator and at room temperature and must be stored at-15 to-25 ℃ in the dark.
CN102018714A discloses a lyophilized ET-743 composition, mainly comprising ET-743 and a disaccharide, wherein the disaccharide is selected from lactose, trehalose, sucrose, and mixtures thereof. The invention achieves the aim of providing a stable dosage form of ET-743 by adding disaccharide to reduce the generation of impurity ET-701 in the freeze-drying process. However, some patients are tolerant to lactose in the practical application process, and the invention does not provide a new method for solving the problems of poor solubility and unstable property of ET-743 in water. Furthermore, these formulations still show a significant amount of active ingredient degradation when stored for 3 months at accelerated conditions of 40 ℃/70% RH, suggesting that the formulations are still not very stable if stored for a long period of time.
There is a need to provide a composition that addresses problems not achieved or completely solved by existing formulations and manufacturing methods, including stability and solubility of the trabectedin, embodiments of the trabectedin formulation should preferably exhibit advantageous freeze-drying characteristics.
The nano-emulsion is also called microemulsion, and is a transparent or semitransparent uniform dispersion system with the particle size of 1-100 nm formed spontaneously by a water phase, an oil phase, a surfactant, a cosurfactant and the like. Nanoemulsions have a number of advantages that are not comparable to other formulations: 1) the nano-emulsion is an isotropic transparent liquid and belongs to a thermodynamic stable system; 2) the nanoemulsion has a fat-soluble inner core, can encapsulate hydrophobic drugs, improves the solubility of the drugs, reduces the enzymolysis of the drugs in vivo, and also can improve the stability of the drugs in the storage process; 3) can improve the absorption of the medicine in the gastrointestinal tract, thereby improving the bioavailability of the medicine. Therefore, nanoemulsions have been extensively studied as a novel drug carrier.
Disclosure of Invention
The invention aims to provide a trabectedin nanoemulsion and a preparation method thereof. The invention also provides a composition containing the trabectedin nanoemulsion, and the composition can be in the form of emulsion or freeze-dried preparation.
The solubility of the trabectedin is improved by adopting a nanoemulsion technology, and meanwhile, because the fat-soluble inner core wraps the medicine, the contact between the trabectedin and water and the air is reduced, and the storage stability of the trabectedin the preparation process and after the medicine is prepared is improved.
In order to achieve the purpose, the technical scheme disclosed by the invention is as follows:
the invention provides a trabectedin nanoemulsion, which comprises a solubilizer and an emulsifier.
The solubilizing agent in the Trabetidine nanoemulsion provided by the invention is selected from one or more of natural oil and synthetic oil.
The emulsifier in the nanoemulsion of trabectedin provided by the invention can be selected from 15-hydroxystearic acid polyethylene glycol ester, tween 80, poloxamer 188, lecithin, sodium oleate, span 20, tween 20, polyoxyethylene monostearate, polyoxyethylene stearate, polyoxyethylene 40 stearate, polyoxyethylene 400 monolaurate, polyoxyethylene 400 monostearate, polyoxyethylene 400 monooleate, polyoxyethylene lauryl ether, polyoxyethylene cetyl ether, polyoxyethylene hydrogenated castor oil, polyoxyethylene alkylphenol, polyoxyethylene fatty alcohol ether, polyoxyethylene nonyl phenol ether, polyoxyethylene octyl phenyl ether, polyoxyethylene lauryl ether, preferably 15-hydroxystearic acid polyethylene glycol ester, lecithin, poloxamer 188, tween 20 and tween 80, more preferably 15-hydroxystearic acid polyethylene glycol ester and lecithin.
The invention provides the trastudine nanoemulsion, the mass ratio of the trastudine to the solubilizer is 1: 10000-1: 1,
preferably 1:1000 to 1:10, more preferably 1:500 to 1: 100.
The mass ratio of the solubilizer to the emulsifier in the trimebutine nanoemulsion provided by the invention is 1: 0.02-1: 100, preferably 1: 0.1-1: 20, and more preferably 1: 1-1: 10.
When the natural oil is used in the trobestatin nanoemulsion, the natural oil can be selected from soybean oil, jojoba oil, olive oil, sweet almond oil, coconut oil, castor oil, sunflower seed oil and fish oil, and is preferably selected from soybean oil, coconut oil, olive oil and fish oil. When synthetic fats and oils are used, the synthetic fats and oils may be selected from medium chain glycerates, isooctyl palmitate, isopropyl palmitate, preferably medium chain glycerates.
The solubilizer in the trabectedin nanoemulsion provided by the invention is preferably a combination of natural oil and synthetic oil, such as soybean oil/medium chain glycerate, olive oil/medium chain glycerate, coconut oil/medium chain glycerate, fish oil/medium chain glycerate and the like for injection. In a preferred embodiment, the ratio of the natural oil to the synthetic oil is 1:100 to 1:0.02, preferably 1:20 to 1:0.2, and more preferably 1:10 to 1: 1.
The solubilizing agent and the emulsifier in the trobestatin nanoemulsion of the present invention can be selected from the following combinations: soybean oil/medium chain glycerate/15-hydroxystearic acid polyethylene glycol ester, olive oil/medium chain glycerate/lecithin E80, coconut oil/medium chain glycerate/poloxamer 188, fish oil/medium chain glycerate/Tween 80.
The particle size of the trabectedin nanoemulsion is 10-100 nm, preferably 10-50 nm, more preferably 20-50 nm, and most preferably 20-40 nm.
The nano-emulsion can be prepared by the following method:
1) taking an emulsifier and a solubilizer to form an oil phase;
2) mixing water and oil phase to form micelle;
3) adding the terbitidine into the micelle to obtain primary emulsion;
4) adding appropriate amount of water to the colostrum.
The trabectedin nanoemulsion disclosed by the invention can form a composition, and the composition also comprises a buffering agent, an osmotic pressure regulator and another excipient.
Another excipient in the composition disclosed by the invention can be selected from mannitol, sucrose, sorbitol, glucose, fructose, trehalose, lactose, maltose, dextran, hydroxyethyl starch, hydroxypropyl-beta cyclodextrin and the like, and mannitol and sucrose are preferred. The content of the excipient is selected from 0.01% to 50% (w/v), preferably 0.1% to 20% (w/v), and more preferably 1% to 10% (w/v).
The buffer in the disclosed compositions may be selected from the group consisting of phosphate buffers, carbonate buffers, citrate buffers, acetate buffers, glycine/hydrochloric acid buffers, phthalate buffers, borate buffers, lactate buffers, succinate buffers, tartrate buffers, preferably phosphate buffers. The content of the buffer salt is selected from 0.01-2% (w/v), preferably 0.05-1% (w/v), more preferably 0.1-0.5% (w/v).
The concentration of the trabectedin the composition disclosed by the invention is selected from 0.001 mg/mL-10 mg/mL, preferably 0.01 mg/mL-0.61 mg/mL, and more preferably 0.1 mg/mL-0.4 mg/mL.
The content of the solubilizer in the composition of the trabectedin disclosed by the invention is selected from 0.1-50% (w/v), preferably 0.2-20% (w/v), and more preferably 0.5-5% (w/v).
The content of the emulsifier in the composition of the trabectedin disclosed by the invention is selected from 0.01-50% (w/v), preferably 0.1-20% (w/v), and more preferably 1-10% (w/v).
The tonicity modifier in the disclosed compositions may be selected from the group consisting of sodium chloride, dextrose, sorbitol, glycerin, PEG, propylene glycol, preferably sodium chloride. The content of the osmotic pressure regulator is selected from 0.2-2% (w/v), preferably 0.4-1% (w/v), more preferably 0.5-0.8% (w/v).
The pH range of the composition disclosed by the invention is 6-9, preferably 6-8, and more preferably 7-8. The composition of the trabectedin disclosed by the invention can be prepared into a freeze-dried preparation which has good re-solubility, and the particle size range of the re-dissolved nano-emulsion is 10-100 nm, preferably 10-50 nm, more preferably 20-50 nm, and most preferably 20-40 nm.
The invention discloses a preparation method of the trabectedin nano freeze-dried substance, which comprises the following steps:
1) heating the emulsifier and the solubilizer to 50-60 ℃, uniformly stirring and mixing, and cooling the water for injection to 40-45 ℃ for later use;
2) adding injection water with the temperature of 40-45 ℃ accounting for about 15% of the total volume into the oil phase, and stirring until micelles are formed; 3) wetting a certain amount of trabectedin a small amount of water, adding the wetted trabectedin a micellar solution, and stirring the mixture for 20 to 60 minutes at the temperature of between 40 and 45 ℃ to form primary emulsion;
4) adding water for injection to 90% of the total weight, adding osmotic pressure regulator and buffer, stirring to dissolve, and mixing. And adjusting the pH value of the solution to 7-8.
5) Filtering and sterilizing by adopting a 0.22 mu m PVDF millipore filter membrane.
6) Filling and half plugging, and freeze drying.
The invention discloses a freeze-drying process of a trabectedin nano-emulsion freeze-dried preparation, which comprises three stages of pre-freezing, primary drying and secondary sublimation. The pre-freezing temperature is-25 to-50 ℃, the primary drying temperature is-10 to-40 ℃, the secondary drying temperature is 15 to 30 ℃, and the vacuum degree is 0.1 to 0.5mbar in the primary drying and secondary drying processes. Nitrogen is introduced throughout the freeze-drying process, preferably 5-10 minutes before the end of freeze-drying.
The Trabectedin nano-emulsion freeze-dried preparation disclosed by the invention is white cake-shaped, has good stability and good re-solubility, overcomes the defects of poor water solubility and stability of the Trabectedin, and is beneficial to the application of the Trabectedin in clinical treatment.
The trabectedin and impurities thereof of the invention may be of semisynthetic or synthetic origin, including combinations of such sources, for example as described in WO0069862 and WO 0187895; on the other hand, the impurities of the trabectedin can also be obtained by outsourcing a reference substance or positioning the types of the impurities of the trabectedin according to a high performance liquid phase method.
Detailed Description
Example 1
The soybean oil for injection and the medium-chain glycerate are taken as solubilizer, 15-hydroxystearic acid-polyethylene glycol ester is taken as emulsifier, disodium hydrogen phosphate is taken as pH buffering agent, sodium chloride is taken as osmotic pressure regulator to prepare the trabectedin nanoemulsion, and mannitol is taken as another excipient to prepare the freeze-dried preparation, wherein the formula comprises the following components in percentage by weight:
Figure BDA0001178611340000051
the preparation method comprises the following steps:
1. adding 15-hydroxystearic acid-polyethylene glycol ester, medium-chain glyceride and soybean oil into a 7mL penicillin bottle, heating to 60 +/-5 ℃, and stirring at a certain stirring speed until the mixture is uniformly mixed.
2. 10mL of water for injection is taken and placed in beaker 2, and cooled to 45 ℃ for later use.
3. About 2.4mL (15%) of cooled water for injection was poured into penicillin bottle 1 and stirred at 400rpm for 10min until micelle formation, which was maintained at 45 ℃.
4. Placing 2mg of trabectedin powder into a 7mL penicillin bottle, adding 160 μ L of water for injection at 45 ℃ to wet API, adding into a beaker 1, adding 160 μ L of water for injection into the penicillin bottle 1, washing the beaker for 2 times, and stirring for 15 min.
5. 12mL of water for injection at room temperature is added into the beaker 1, mixed evenly, and added with sodium chloride and anhydrous disodium hydrogen phosphate until being completely dissolved. The pH was adjusted to 7.5. + -. 0.5 with 1N sodium hydroxide.
6. 0.8g of mannitol was added to the above nanoemulsion.
7. Filtering the nano emulsion solution by adopting a PVDF membrane with the diameter of 0.22 mu m, filling and half plugging, and then freeze-drying.
The particle size of the trabectedin nanoemulsion and the appearance, moisture, particle size after redissolution and pH value of the freeze-dried preparation thereof are examined (as shown in Table 1), and the change of the content of impurities after freeze-drying and the standing stability under the acceleration condition of 25 ℃ are detected (as shown in Table 2).
TABLE 1 Properties of nanoemulsion and its lyophilized formulation
Appearance of the product Moisture content pH value Particle size before lyophilization Particle size after reconstitution
White cake shape 0.624％ 7.41 29.07nm 29.06nm
TABLE 2 detection of impurity content and stability in standing
Figure BDA0001178611340000061
And (4) conclusion: the appearance and the moisture of the lobitedine nano-emulsion freeze-dried preparation are qualified, the pH value and the particle size after re-dissolution do not change greatly, and the impurity detection result of the freeze-dried preparation meets the limit requirement; the stability of the trobestatin nanoemulsion freeze-dried preparation is good, which shows that the stability of the trobestatin in the freeze-drying process and the long-term storage process can be improved by the nanoemulsion.
Example 2
The preparation method comprises the steps of preparing the trabectedin nanoemulsion by using olive oil for injection and medium-chain glycerate as solubilizers, lecithin as emulsifier, disodium hydrogen phosphate as buffer and sodium chloride as osmotic pressure regulator, and preparing the freeze-dried preparation by using sucrose as freeze-drying protective agent, wherein the formula comprises the following components in parts by weight:
Figure BDA0001178611340000071
the preparation method comprises the following steps:
1. adding lecithin E80, medium chain glyceride and olive oil into a 7mL penicillin bottle, heating to 60 + -5 deg.C, and stirring at a certain stirring speed until uniformly mixed.
2. 10mL of water for injection is taken and placed in beaker 2, and cooled to 45 ℃ for later use.
3. About 2.4mL (15%) of cooled water for injection was poured into penicillin bottle 1 and stirred at 400rpm for 10min until micelle formation, which was maintained at 45 ℃.
4. Placing 2mg of trabectedin powder into a 7mL penicillin bottle, adding 160 μ L of 45 ℃ water for injection to wet API, adding into a beaker 1, adding 160 μ L of water for injection into the penicillin bottle 1, washing the beaker for 2 times, and stirring for 15 min.
5. 12mL of water for injection at room temperature is added into the beaker 1, mixed evenly, and added with sodium chloride and anhydrous disodium hydrogen phosphate until being completely dissolved. The pH was adjusted to 7.5. + -. 0.5 with 1N sodium hydroxide.
6. 0.8g of sucrose was added to the above nanoemulsion.
7. Filtering the nano emulsion by adopting a PVDF membrane with the diameter of 0.22 mu m, filling and half plugging, and then freeze-drying.
The particle size of the trabectedin nanoemulsion and the appearance, moisture, particle size after redissolution and pH value of the freeze-dried preparation thereof are examined (as shown in Table 3), and the change of the content of impurities after freeze-drying and the standing stability under the acceleration condition of 25 ℃ are detected (as shown in Table 4).
TABLE 3 Properties of the nanoemulsion and its lyophilized formulation
Appearance of the product Moisture content pH value Particle size before lyophilization Particle size after reconstitution
White cake shape 1.146％ 7.40 28.82nm 28.86nm
TABLE 4 detection of impurity content and stability in standing
Figure BDA0001178611340000072
Figure BDA0001178611340000081
And (4) conclusion: the preparation method has the advantages that the trobestatin nano-emulsion freeze-dried preparation prepared by taking lecithin as an emulsifier is qualified in appearance and moisture, the pH value and the particle size after redissolution are not greatly changed, and the impurity detection result of the freeze-dried preparation meets the limit requirement; the stability of the trobestatin nanoemulsion freeze-dried preparation is good, which shows that the stability of the trobestatin in the freeze-drying process and the long-term storage process can be improved by the nanoemulsion.
Example 3
Coconut oil and medium-chain glyceride are taken as solubilizers, poloxamer 188 is taken as an emulsifier, disodium hydrogen phosphate is taken as a buffering agent, sodium chloride is taken as an osmotic pressure regulator to prepare the trabectedin nanoemulsion, and sucrose is taken as another excipient to prepare a freeze-dried preparation of the trabectedin nanoemulsion, wherein the formula comprises the following components in dosage:
Figure BDA0001178611340000082
the preparation method comprises the following steps:
1. adding poloxamer 188, medium-chain glyceride and coconut oil into a 7mL penicillin bottle, heating to 60 +/-5 ℃, and stirring at a certain stirring speed until the components are uniformly mixed.
2. 10mL of water for injection is taken and placed in beaker 2, and cooled to 45 ℃ for later use.
3. About 2.4mL (15%) of cooled water for injection was poured into penicillin bottle 1 and stirred at 400rpm for 10min until micelle formation, which was maintained at 45 ℃.
4. Placing 2mg of trabectedin powder into a 7mL penicillin bottle, adding 160 μ L of 45 ℃ water for injection to wet API, adding into a beaker 1, adding 160 μ L of water for injection into the penicillin bottle 1, washing the beaker for 2 times, and stirring for 15 min.
5. 12mL of water for injection at room temperature is added into the beaker 1, mixed evenly, and added with sodium chloride and anhydrous disodium hydrogen phosphate until being completely dissolved. The pH was adjusted to 7.5. + -. 0.5 with 1N sodium hydroxide.
6. 0.8g of sucrose was added to the nanoemulsion injection.
7. Filtering the nano emulsion solution by adopting a PVDF membrane with the diameter of 0.22 mu m, filling and half plugging, and then freeze-drying.
The particle size of the trabectedin nanoemulsion and the appearance, moisture, particle size after redissolution and pH value of the freeze-dried preparation thereof were examined (as shown in Table 5), and the change of the content of impurities after freeze-drying and the stability in standing under an accelerated condition at 25 ℃ were examined (as shown in Table 6).
TABLE 5 properties of nanoemulsion and its lyophilized formulation
Appearance of the product Moisture content pH value Particle size before lyophilization Particle size after reconstitution
White cake shape 1.062％ 7.39 28.94nm 28.95nm
TABLE 6 detection of impurity content and stability in standing
Figure BDA0001178611340000091
And (4) conclusion: the preparation method has the advantages that the trobestatin nano-emulsion freeze-dried preparation prepared by taking poloxamer 188 as an emulsifier is qualified in appearance and moisture, the pH value and the particle size after redissolution are not greatly changed, and the impurity detection result of the freeze-dried preparation meets the limit requirement; the stability of the trobestatin nanoemulsion freeze-dried preparation is good, which shows that the stability of the trobestatin in the freeze-drying process and the long-term storage process can be improved by the nanoemulsion.
Example 4
The preparation method comprises the steps of preparing the trabectedin nanoemulsion by using fish oil for injection and medium-chain glyceride as solubilizers, tween 80 as an emulsifier, disodium hydrogen phosphate as a pH buffering agent and sodium chloride as an osmotic pressure regulator, and preparing the freeze-dried preparation by using trehalose as a freeze-drying protective agent, wherein the formula comprises the following components in percentage by weight:
Figure BDA0001178611340000092
Figure BDA0001178611340000101
the preparation method comprises the following steps:
1. adding tween 80, the medium-chain glyceride and the fish oil into a 7mL penicillin bottle, heating to 60 +/-5 ℃, and stirring at a certain stirring speed until the mixture is uniformly mixed.
2. 10mL of water for injection is taken and placed in beaker 2, and cooled to 45 ℃ for later use.
3. About 2.4mL (15%) of cooled water for injection was poured into penicillin bottle 1 and stirred at 400rpm for 10min until micelle formation, which was maintained at 45 ℃.
4. Putting 4mg of trabectedin powder into a 7mL penicillin bottle, adding 160 mu L of 45 ℃ water for injection to wet API, adding the mixture into a beaker 1, adding 160 mu L of water for injection into the penicillin bottle 1, washing the beaker for 2 times, and stirring for 15 min.
5. 12mL of water for injection at room temperature is added into the beaker 1, mixed evenly, and added with sodium chloride and anhydrous disodium hydrogen phosphate until being completely dissolved. The pH was adjusted to 7.5. + -. 0.5 with 1N sodium hydroxide.
6. 0.8g of trehalose was added to the above nanoemulsion.
7. Filtering the nano emulsion solution by adopting a PVDF membrane with the diameter of 0.22 mu m, filling and half plugging, and then freeze-drying.
The particle size of the trabectedin nanoemulsion and the appearance, moisture, particle size after redissolution and pH value of the freeze-dried preparation thereof were examined (as shown in Table 7), and the change of the content of impurities after freeze-drying and the stability in standing under an accelerated condition at 25 ℃ were examined (as shown in Table 8).
TABLE 7 Properties of nanoemulsion and its lyophilized formulation
Appearance of the product Moisture content pH value Particle size before lyophilization Particle size after reconstitution
White cake shape 0.0742％ 7.41 28.34nm 28.37nm
TABLE 8 detection of impurity content and stability in standing
Figure BDA0001178611340000102
Figure BDA0001178611340000111
And (4) conclusion: the trabectedin nano-emulsion freeze-dried preparation prepared by taking the Tween 80 as the emulsifier has qualified appearance and moisture, the pH value and the particle size after redissolution do not change greatly, and the impurity detection result of the freeze-dried preparation meets the limit requirement; the stability of the trobestatin nanoemulsion freeze-dried preparation is good, which shows that the stability of the trobestatin in the freeze-drying process and the long-term storage process can be improved by the nanoemulsion.

Claims (40)

1. The trabectedin nanoemulsion comprises a solubilizer and an emulsifier, and is characterized in that the solubilizer is a combination of natural oil and synthetic oil, wherein the natural oil is selected from soybean oil, coconut oil, olive oil and fish oil, the synthetic oil is selected from medium-chain glycerate, isooctyl palmitate and isopropyl palmitate, the emulsifier is selected from 15-hydroxystearic acid polyethylene glycol ester, phospholipid E80, Tween 80 and poloxamer 188, the mass ratio of the trabectedin to the solubilizer is 1: 1000-1: 10, and the mass ratio of the solubilizer to the emulsifier is 1: 1-1: 10.
2. The nanoemulsion according to claim 1, characterized in that the mass ratio of trabectedin to the solubilizer is 1: 500-1: 100.
3. The nanoemulsion of claim 1, wherein the solubilizer is a mixture of natural oil and synthetic oil, and the mass ratio of the natural oil to the synthetic oil is 1: 100-1: 0.02.
4. The nanoemulsion of claim 1, wherein the solubilizer is a mixture of natural oil and synthetic oil, and the mass ratio of the natural oil to the synthetic oil is 1: 20-1: 0.2.
5. The nanoemulsion of claim 1, wherein the solubilizer is a mixture of natural oil and synthetic oil, and the mass ratio of the natural oil to the synthetic oil is 1: 10-1: 1.
6. The nanoemulsion of claim 1, wherein the synthetic oil is a medium chain glycerate.
7. The nanoemulsion of claim 1, characterised in that the solubilising agent is selected from the group consisting of soybean oil/medium chain glycerate, olive oil/medium chain glycerate, coconut oil/medium chain glycerate, fish oil/medium chain glycerate.
8. The nanoemulsion of claim 1, characterized in that the solubilizer and the emulsion in the nanoemulsion are in a combination selected from the group consisting of: soybean oil/medium chain glycerate/15-hydroxystearic acid polyethylene glycol ester, olive oil/medium chain glycerate/lecithin E80, coconut oil/medium chain glycerate/poloxamer 188, fish oil/medium chain glycerate/Tween 80.
9. The nanoemulsion according to any of claims 1-8, characterized in that the particle size of the nanoemulsion ranges from 10 to 100 nm.
10. The nanoemulsion according to claim 9, wherein the particle size of the nanoemulsion ranges from 10 to 50 nm.
11. The nanoemulsion according to claim 9, characterized in that the particle size of the nanoemulsion is between 20 and 50 nm.
12. The nanoemulsion according to claim 9, characterized in that the particle size of the nanoemulsion is between 20 and 40 nm.
13. A process for preparing a nanoemulsion according to any of claims 1 to 8, 10 to 12, characterized in that it comprises the following steps:
1) taking an emulsifier and a solubilizer to form an oil phase;
2) mixing water and oil phase to form micelle;
3) adding the terbitidine into the micelle to obtain primary emulsion;
4) adding appropriate amount of water to the colostrum.
14. A composition comprising a nanoemulsion according to any one of claims 1 to 8, 10 to 12, further comprising another excipient.
15. The composition of claim 14, further comprising a buffering agent.
16. The composition of claim 14, further comprising a buffer selected from the group consisting of phosphate buffers, carbonate buffers, citrate buffers, acetate buffers, glycine/hydrochloric acid buffers, phthalate buffers, borate buffers, lactate buffers, succinate buffers, and tartrate buffers.
17. The composition of claim 14, further comprising a phosphate buffer.
18. A composition comprising a nanoemulsion according to any of claims 1 to 8 and 10 to 12, further comprising another excipient selected from mannitol, sucrose, sorbitol, glucose, fructose, trehalose, lactose, maltose, dextran, hydroxyethyl starch, hydroxypropyl-beta cyclodextrin.
19. A composition comprising a nanoemulsion according to any of claims 1 to 8 and 10 to 12, further comprising another excipient selected from mannitol and sucrose.
20. The composition according to claim 15, wherein the content of the solubilizer is selected from the group consisting of 0.1% to 50% (w/v).
21. The composition according to claim 15, wherein the content of the solubilizer is selected from the group consisting of 0.2% to 20% (w/v).
22. The composition according to claim 15, wherein the content of the solubilizer is selected from the group consisting of 0.5% to 5% (w/v).
23. The composition according to claim 15, wherein the emulsifier is present in an amount selected from the group consisting of 0.01% to 50% (w/v).
24. The composition according to claim 15, wherein the emulsifier is present in an amount selected from the group consisting of 0.1% to 20% (w/v).
25. The composition according to claim 15, characterized in that the content of said emulsifier is selected from 1% to 10% (w/v).
26. The composition according to claim 15, wherein the excipient is present in an amount selected from the group consisting of 0.01% to 50% (w/v).
27. The composition according to claim 15, characterized in that the content of said excipient is selected from 0.1% to 20% (w/v).
28. The composition according to claim 15, characterized in that the content of said excipient is selected from 1% to 10% (w/v).
29. The composition of claim 15, wherein the buffer is present in an amount selected from the group consisting of 0.01% to 2% (w/v).
30. The composition of claim 15, wherein the buffer is present in an amount selected from the group consisting of 0.05% to 1% (w/v).
31. The composition of claim 15, wherein the buffer is present in an amount selected from the group consisting of 0.1% to 0.5% (w/v).
32. The composition according to any one of claims 15-17, 20-31, wherein the pH of the composition is selected from the range of 6 to 9.
33. The composition according to any one of claims 15-17, 20-31, wherein the pH of the composition is selected from the range of 6 to 8.
34. The composition according to any one of claims 15-17, 20-31, wherein the pH of the composition is selected from the range of 7 to 8.
35. A lyophilized formulation obtained by lyophilizing the composition of any one of claims 15-17 and 20-31.
36. An emulsion obtained by reconstituting the lyophilized preparation of claim 35, having a particle size in the range of 10 to 100 nm.
37. An emulsion obtained by reconstituting the lyophilized preparation of claim 35, having a particle size in the range of 10 to 50 nm.
38. An emulsion obtained by reconstituting the lyophilized preparation of claim 35, having a particle size in the range of 20 to 50 nm.
39. An emulsion obtained by reconstituting the lyophilized preparation of claim 35, having a particle size in the range of 20 to 40 nm.
40. A method of preparing the lyophilized formulation of claim 35, wherein: a nanoemulsion prepared by the method of claim 18, incorporating the excipient of claim 14, and further comprising the step of lyophilisation.
CN201611144699.4A 2015-12-15 2016-12-13 Trabectedin nanoemulsion and preparation method thereof Withdrawn - After Issue CN106880592B (en)

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1823794A (en) * 2004-10-29 2006-08-30 法马马私人股份有限公司 Formulations
WO2008013785A2 (en) * 2006-07-24 2008-01-31 Singh-Broemer And Company, Inc. Solid nanoparticle formulation of water insoluble pharmaceutical substances with reduced ostwald ripening

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1823794A (en) * 2004-10-29 2006-08-30 法马马私人股份有限公司 Formulations
WO2008013785A2 (en) * 2006-07-24 2008-01-31 Singh-Broemer And Company, Inc. Solid nanoparticle formulation of water insoluble pharmaceutical substances with reduced ostwald ripening

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