Nanoemulsion delivery system compositionThe priority declares that:
this application claims priority based on the united states patent application No. 14/623, 150 filed on 16/2 2014 and united states provisional patent application No. 61/939,965 filed on 14/2 2014, and both of these united states patent applications are incorporated by reference in their entirety into this application.
Technical Field
The present invention relates to a stable, transparent, oil-in-water type nanoemulsion composition with an average diameter of less than 100nm, consisting of an oil phase (at least 10% w/v long chain triglycerides) and an aqueous phase without the addition of ethanol as a co-solvent. The preparation process of the composition can be used for preparing active ingredient carriers in pharmacy, foods, health products and cosmetics.
Background
The low solubility of the drug makes it difficult to develop drugs and health products, particularly intravenous and oral solutions, having biocompatibility and physical stability. A plurality of methods can be used for preparing the alkaline medicine intravenous injection and oral liquid composition which is difficult to dissolve in water.
These methods include: using surfactant to perform micelle solubilization or prepare a drug nanoparticle suspension; preparing a complex (hydroxypropyl-beta-cyclodextrin (HPBCD) and beta-sulfobutyl ether-beta-cyclodextrin (SBECD)) with cyclodextrin and a derivative thereof; various co-solvent systems are used; the strong acid salt is prepared using a low pH solution.
However, in the micellar system, the surfactant is liable to cause adverse reactions such as irritation, hemolysis, histamine, allergy, etc.; it has been reported that due to the higher contact area of the drug with the surfactant in the aqueous medium, there is a potential risk of catalytic degradation of the drug in nanosuspensions with polymers, surfactants. Taste masking and injection site pain are another problem caused by the higher concentration of free drug in the aqueous medium of the micelle/nanosuspension system;
it is well known that cosolvent systems are prone to precipitation, injection pain, and phlebitis; it has been reported that the use of cyclodextrins and their derivatives co-administered in combination with lipophilic drugs may induce potential renal toxicity and bradycardia, blood pressure reduction. The use of strong acids and weak base salts to make low pH solutions can cause drug-adjuvant, drug stability problems, and upon contact with blood at neutral pH, can cause the drug to precipitate as a free base, causing taste masking problems, irritation and pain at the injection site.
In summary, each approach has its inherent limitations, and neither of the intravenous, ophthalmic, nasal, topical, transdermal or oral formulations has the ability to incorporate low solubility drugs into biocompatible carriers to produce formulations with good stability, low side effects, and good pharmacokinetic profiles.
Oil-in-water type emulsion, which is composed of oil droplets dispersed in an aqueous continuous phase, is a special system capable of solving the problems of drug solubility and stability, and can be used for preparing a plurality of drugs, foods and cosmetics. One use of the emulsion is to deliver active pharmaceutical ingredients and active ingredients for topical, health, oral, nasal, and ophthalmic active ingredients and drugs. The active ingredient soluble in the oil phase may be dissolved/dispersed in the oil phase of the emulsion, or may be incorporated into the interfacial region of the emulsion if the active ingredient is poorly soluble in the oil or water phase.
Depending on the appearance and particle size of the emulsion, it can be divided into three types: macroemulsions, microemulsions and nanoemulsions. The coarse emulsion has an average particle size of greater than 100nm and appears cloudy and milky due to much of the interface scattering light passing through the emulsion. Microemulsions and nanoemulsions are two special emulsions that are optically transparent (translucent or transparent), with an average particle size of less than 100 nm. The optically transparent nature is due to scattering of light as it passes through droplets of 1/4 having a particle size larger than the wavelength of the incident light. When the average particle diameter of the droplets in the emulsion is less than 100nm, particularly less than 70nm, the light beam can pass through the emulsion directly without being scattered. Microemulsions are thermodynamically stable systems that form spontaneously by "dissolving" oil molecules in a mixture with surfactants, co-surfactants and co-solvents. However, nanoemulsions are thermodynamically metastable systems, the formation of which requires external energy to break down oil droplets below 100 nm.
Conventional oil-in-water emulsions, i.e., macroemulsions, are inherently unstable systems that do not form spontaneously. The formation of a coarse emulsion requires energy input in the form of mechanical agitation, homogenization, or sonication, etc., and the formed coarse emulsion has a tendency to return to a phase separation stable state (e.g., agglomeration and stratification). In addition to the instability of physical properties, due to the relatively large particle size of the droplets of the macroemulsion and the relatively low interfacial volume, the effective dissolution of a poorly soluble compound, which has low solubility in both the oil and aqueous phases, by the macroemulsion is limited; the opacity of the macroemulsion reduces the visual clarity when the ophthalmic delivery formulation is prepared. In addition, the release rate of active ingredients from oral macroemulsions comprising long chain triglycerides may be limited by the rate and extent of lipolysis. The digestibility of triglyceride emulsions in the gastrointestinal tract is a function of pH, lipase concentration, bile salts and emulsion surface area. The high surface area to volume ratio emulsion has a higher lipid breakdown rate than the low surface area to volume emulsion.
Emulsions with an average particle size of less than 100nm avoid the above-mentioned disadvantages, wherein the microemulsion is thermodynamically stable, and although the nanoemulsion is thermodynamically metastable in nature, the kinetic stability can be maintained for a long time due to the very small particle size. The formation of emulsions with droplet sizes of less than 100nm is of great benefit for the increase in relative interfacial area. The increased relative interfacial area allows for increased solubility of the poorly soluble active ingredient in the aqueous medium, with a faster rate of digestion of the lipid breakdown compared to a coarse emulsion, and thus a faster release rate of the active ingredient from the oil droplets. Since the droplet size is less than 100nm, microemulsions and nanoemulsions may help the active compound to penetrate the epithelial mucosal layer, such as the eye, skin, nose, lung, gastrointestinal tract, tumors, blood vessels, and blood-brain barrier.
The grain diameters of the micro-emulsion and the nano-emulsion are both less than 100nm, and the transmission modes of the active compounds are similar but different. Despite the thermodynamic stability of microemulsions, the surfactant concentration required for their systems is significantly higher than for the oil phase, normally several times higher than for nanoemulsions. Microemulsions are disadvantageous in the preparation of intravenous, ophthalmic and oral formulations compared to nanoemulsions due to the susceptibility of surfactants to a number of undesirable side effects and government regulations limiting the daily intake of many surfactants. In addition, many surfactants are bitter in taste, which tends to cause palatability problems when applied to foods/pharmaceuticals; the physical stability of microemulsion systems is often affected by dilution, heating, and changes in pH.
Although the nanoemulsion does not form spontaneously, only kinetic stability is maintained; the nano-scale particle size can be obtained by using water and less surfactant and breaking oil drops by mechanical shearing. From a toxicological and regulatory point of view, this is a more acceptable system for the human body. Similar to microemulsion, the nanoemulsion has the advantage of being translucent in appearance due to small particle size. Like microemulsions, nanoemulsions have the same high interfacial area to volume ratio, which facilitates the dissolution of poorly soluble compounds and the rapid digestion of the emulsion by fat. Unlike microemulsions, nanoemulsions remain physically stable upon dilution and/or changes in pH.
Despite the many advantages of nanoemulsions over macroemulsions and microemulsions, there is still the limitation of kinetic stabilization-particle size increases over time due to Ostwald ripening. This results in a decrease in clarity and a decrease in contact surface area. To prepare stable nanoemulsions with average particle size less than 100nm, low viscosity oil phases such as short chain triglycerides or medium chain triglycerides (e.g., miglyol) are often used, which have the disadvantage of having a tendency to Ostwald ripening due to their relatively small molecular weight, relatively high water solubility, and relatively low viscosity. The long-chain triglyceride with extremely low water solubility is used as the oil phase, so that the physical stability of the nano emulsion can be improved. However, due to the large molecular volume and high viscosity of the long chain triglycerides, it is difficult to rapidly form optically transparent (transparent or translucent) nanoemulsions when the content thereof is high. Therefore, to prepare optically clear, long chain triglyceride-containing nanoemulsions, it is generally necessary to add large amounts of low molecular weight organic solvents (ethanol), or larger amounts of toxic surfactants (Cremophor EL) relative to the oil phase, thereby reducing the surface tension of the oil phase. However, this may make it less acceptable to humans from a safety, toxicological and regulatory perspective. For example, because phosphatidylcholine (egg yolk or soy lecithin) is a natural non-toxic, biocompatible surfactant, the pharmaceutical industry is currently of great interest for lecithin-based emulsions. However, since phospholipids have a strong tendency to form liquid crystal structures at lower concentrations, especially in the aqueous phase, ethanol must be used as a co-solvent to reduce interfacial tension, thereby preparing a microemulsion/nanoemulsion based on lecithin and containing long-chain triglycerides. However, it is known that ethanol is susceptible to toxic side effects such as enzyme induction, drug-drug interactions or damage to the central nervous system.
Therefore, the nano-emulsion which is prepared by taking long-chain triglyceride as an oil phase as a base and has higher content of an emulsifiable solution phase, has an average particle size of less than 100nm (average strength), uses a biocompatible surfactant and less other surfactants, does not use ethanol as a cosolvent in a water phase, and has good stability and optical translucency for Ostwald ripening is also very challenging. The nanoemulsion plays an important role in improving the safety, curative effect, stability, tolerance and taste masking of the product once being successfully prepared.
Summary of The Invention
In order to solve the above technical drawbacks and problems, there is a need for a technique for preparing clear nanoemulsions comprising at least 10% w/v long chain triglycerides in the oil phase, droplet size less than 100nm and maintaining good stability towards Ostwald ripening, using biocompatible surfactants and less other surfactants (< 15%), without using ethanol as co-solvent in the aqueous phase.
In view of the above problems, the present invention provides an aqueous-based oil-in-water type nanoemulsion composition having an average particle size of less than 100nm, a nanoemulsion oil phase concentration of up to 50%, a small particle size distribution, optical transparency, and good stability to Ostwald ripening. The hydrophobic effective components for treatment and other adjuvants are prepared into nanoemulsion system, which can improve its solubility and stability in aqueous medium, and is helpful for the delivery of active components.
The invention aims to provide an oil-in-water type nano-emulsion with stable properties and optical transparency, which comprises an oil phase consisting of long-chain triglyceride and/or other oils, an ionic surfactant, a cosurfactant and a pH regulator, wherein ethanol is not used as a cosolvent in the water phase.
The oil phase in the emulsion is at least 0.5-50% w/v and the oil phase comprises at least 10% long chain triglycerides.
The ionic surfactant is a biocompatible ionizable surfactant or derivative thereof (e.g. egg yolk or soy lecithin) in combination with a pharmaceutically acceptable co-surfactant at a total concentration of less than 25% w/v, wherein the ratio of surfactant to co-surfactant is in the range of 10:0.1-0.1:10, 10:1-1:5 or 5:1-1: 5; the total concentration of the two is less than 100% w/w of the oil phase.
The aqueous phase comprises water without the use of ethanol as a co-solvent.
The invention aims to provide an oil-in-water type nano-emulsion with stable properties and optical transparency, which comprises long-chain triglyceride, oil drops with the average particle size of less than 100nm, and no ethanol used as a cosolvent in a water phase; the preparation method of the nano-emulsion comprises the following steps: a) preparing an oil phase comprising long chain triglycerides; b) preparing an aqueous phase comprising water and a pH adjusting agent; c) incorporating a biocompatible surfactant and co-surfactant into the oil or water phase; d) dispersing the oil phase in the water phase to form a coarse emulsion; e) d, carrying out ultrasonic or high-pressure homogenization treatment on the crude emulsion obtained in the step d to obtain nano-emulsion; f) the pH value is adjusted.
It is another object of the present invention to provide a method for treating human or animal diseases using an optically clear nanoemulsion system composition comprising long-chain triglycerides, oil droplets having a particle size of less than 100nm, without using ethanol as an organic solvent co-solvent for therapeutic drugs, which can be used in pharmaceuticals, foods, cosmetics, and can be administered orally, intravenously, subcutaneously, intramuscularly, inhalatively, nasally, topically, ophthalmically, transdermally, etc.; the stability and purity of the combination meet the compendium for applicability, FDA and GMP requirements. The method comprises the following steps: a) preparing a liquid oil-in-water nanoemulsion composition by dispersing/dissolving a therapeutically active matrix or other matrix in an oily vehicle; b) preparing an aqueous phase comprising water and a pH adjusting agent; c) dispersing the oil phase in the water phase by ultrasonic or homogenization to form oil drops; d) the nanoemulsion is administered to human or animals.
The present invention is directed to a method of treating human or animal diseases using an optically clear nanoemulsion system composition comprising long chain triglycerides, oil droplets having a particle size of less than 100nm, without the use of ethanol as an organic solvent co-solvent for the therapeutic agent, which can be used in pharmaceuticals, foods, cosmetics, and also can be administered orally, intravenously, subcutaneously, intramuscularly, by inhalation, nasally, topically, ophthalmically, transdermally, etc.; the stability and purity of the combination meet the compendium for applicability, FDA and GMP requirements. The method comprises the following steps: a) dispersing the oil phase in the water phase by using an ultrasonic or homogenizing method to prepare a liquid oil-in-water type nano-emulsion composition; b) incorporating a therapeutically active agent or other drug into the oil-in-water nanoemulsion prepared in step a), mixing into the oil phase to dissolve it; c) the nanoemulsion is administered to human or animals.
The present invention is directed to a method for treating human or animal diseases using an optically clear nanoemulsion system composition comprising long chain triglycerides, oil droplets having a particle size of less than 100nm, without using ethanol as an organic solvent co-solvent for therapeutic drugs, which can be used in pharmaceuticals, foods, cosmetics, and can be administered orally, intravenously, subcutaneously, intramuscularly, by inhalation, nasally, topically, ophthalmically, transdermally, etc.; the stability and purity of the combination meet the compendium for applicability, FDA and GMP requirements. The method comprises the following steps: a) dispersing the oil phase in the water phase to prepare a liquid oil-in-water type coarse emulsion; b) incorporating a therapeutically active agent or other drug into the oil-in-water macroemulsion prepared in step a) and mixing into the oil phase to dissolve it; c) preparing liquid oil-in-water type nano-emulsion by ultrasonic or homogenization; d) the nanoemulsion is administered to human or animals.
Brief description of the drawings
FIG. 1 graph comparing particle size distribution of LCT/lecithin/Tween 80 nanoemulsion (example 1) and lecithin macroemulsion (example 7). The average particle size of the nanoemulsion was 47nm, and the average particle size of the macroemulsion was 177 nm.
Figure 2 transparent nanoemulsion of example 5
FIG. 3 particle size distribution plot of LCT/lecithin/Tween 80 nanoemulsion (example 1) stored for 5 months at 40 ℃/75% RH. The results show that the particle size distribution of the nanoemulsion before and after storage is substantially consistent.
Figure 4 particle size distribution of LCT/lecithin/tween 80/nanoemulsion/cyclosporine nanoemulsion (example 2) stored for 2 months, 5 months under 40 ℃/75% RH conditions. The particle size and the optical transparency of the nano-emulsion have no significant change. The average particle size of the nanoemulsion before storage was 38nm, and after storage at 40 ℃ for 5 months, it was 45 nm.
FIG. 5 particle size distribution of LCT/MCT/lecithin/Tween 80/nanoemulsion/cyclosporine nanoemulsion (example 3) stored for 2 months under 40 ℃/75% RH conditions and refrigerated conditions (-2-8 ℃). The results show that the particle size distribution of the particles before and after storage of the nanoemulsion is substantially consistent.
FIG. 6 particle size distribution of LCT/lecithin/Tween 80/nanoemulsion/cyclosporin nanoemulsion (example 5) stored for 12 months at room temperature and refrigerated (-2-8 ℃ C.). The particle size and the optical transparency of the nano-emulsion have no significant change. The average particle size of the nanoemulsion before storage was 38nm, and after storage at 40 ℃ for 5 months, it was 45 nm.
Detailed Description
The term "emulsion" refers to a system of liquids dispersed in another immiscible liquid (e.g. fat dispersed in milk) in the form of droplets with a particle size larger than that of the micelles, with or without the aid of emulsifiers.
The term "oil-in-water emulsion" refers to an emulsion system formed by oil droplets dispersed in an aqueous continuous phase. In the present invention, the term "emulsion" refers to all oil-in-water type emulsions.
The term "microemulsion" refers to a dispersion of water, oil and surfactant (S), which is an isotropic thermodynamically stable system having a particle size distribution of about 1 to 100nm, usually 10 to 50 nm. The droplet size is obtained by measuring the average particle size or the intensity-uniform average particle size by dynamic light scattering. In the present invention, the term "microemulsion" refers to all oil-in-water emulsions.
The term "nanoemulsion" refers to a dispersion of water, oil and surfactant (S), a thermodynamically metastable system with a particle size distribution of about 1-100 nm, usually 10-50 nm. The droplet size is obtained by measuring the average particle size or the intensity-uniform average particle size by dynamic light scattering. In the present invention, the term "nanoemulsion" refers to all oil-in-water emulsions.
The term "medium chain triglyceride" refers to a medium chain (6 to 12 carbon atoms) glycerol fatty acid ester.
The term "long chain triglyceride" refers to long chain (greater than 12 carbon atoms) glycerol fatty acid esters.
The term "surfactant" generally refers to an amphiphilic organic compound having both hydrophobic and hydrophilic groups.
The term "ionic surfactant" generally refers to an amphiphilic organic compound having both hydrophobic and hydrophilic groups, the terminal groups of which can ionize at physiological conditions below pH 10.
The term "co-surfactant" generally refers to another surfactant that, when added, further lowers the surface tension of a liquid.
The term "co-solvent" refers to an organic solvent that, when added to another solvent, further reduces the surface tension of the liquid.
The term "transparent" refers to the physical property that light passes through a material without being scattered. It follows Snell's law; in other words, a transparent medium allows light transmission and image formation.
The term "translucent" refers to a transparent superset: it is transparent to light, but does not necessarily follow Snell's law; in other words, a translucent medium allows transmission of light but does not allow image formation.
The term "optically transparent" as used herein refers to transparent or translucent.
The invention provides an oil-in-water type nano-emulsion which takes a water phase as a matrix, has an average droplet size (intensity-average, nm) of less than 100nm, and consists of an oil phase, a mixture of an ionic surfactant and a cosurfactant and a water phase liquid carrier. The prescription of the invention comprises:
a) oil phase of long chain triglycerides and/or other oils
b) Mixtures of ionic surfactants and cosurfactants
c) Water phase comprising water, pH regulator, and ethanol-free cosolvent
Alternatively, the emulsion formulation may further contain an antioxidant, a chelating agent, a penetrating agent, a suspending agent, a preservative and a buffer agent, which are active ingredients (S) in pharmaceutical products, health products, foods and cosmetics. In some embodiments, the formulation further comprises a solubilizing agent, flavoring agent, sweetener, viscosity increasing agent, electrolyte, other therapeutic agent, or a combination thereof.
The present invention may provide various embodiments in combination with the combination of different ingredients in the nanoemulsion.
The nanoemulsion of one embodiment of the invention consists of:
a) at least 0.5-50% w/w of an oil phase comprising long chain triglycerides,
b) 0.01-30% w/w of an ionic surfactant
c) 0.01-30% w/w of a co-surfactant; and
d) 50-99% w/w of an aqueous phase
Wherein the average particle size of the oil droplets is less than 100nm, the ratio of the ionic surfactant to the co-surfactant is from 10:0.1 to 0.1:10, 10:1 to 1:5 or 5:1 to 1:5, and the ratio of the surfactant/co-surfactant mixture to the oil is less than 1: 1.
In a preferred embodiment, the oil-in-water nanoemulsion comprises 0.5-50 w/v% of an oil phase (containing at least 10% w/w of long chain triglycerides in the oil phase), 0.01-30% of an ionic surfactant, 0.01-30% of a co-surfactant, and an aqueous phase without ethanol as a co-solvent.
The oil phase in the emulsion may be liquid or solid fats from animals, vegetables, algae or artificial synthetics. Oils or fats of animal origin include fish oil, cod liver oil, whale oil, lard, tallow, goose oil and milk fat. Oils of vegetable origin include rapeseed oil, castor oil, cocoa butter, coconut oil, coffee seed oil, corn oil, cottonseed oil, evening primrose oil, grape seed oil, linseed oil, menhaden oil, mustard oil, olive oil, palm kernel oil, peanut oil, poppy seed oil, rapeseed oil, rice bran oil, safflower oil, sesame oil, soybean oil, sunflower seed oil, palm kernel oil, hazelnut oil, sesame oil and wheat germ oil. Synthetic oils include synthetic triglycerides, fractionated triglycerides, modified triglycerides, hydrogenated triglycerides or mixtures of partially hydrogenated and triglycerides.
The preferred oil phase in the emulsion is a pharmaceutical grade oil, particularly triglycerides, but is not limited to soybean oil, safflower oil, olive oil, cottonseed oil, sunflower oil, fish oil (containing the omega-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)), castor oil, sesame oil, peanut oil, corn oil, medium chain triglycerides (such as Miglyol812 or 810) and short chain triglycerides. Surfactants and/or co-surfactants may also be included in the oil phase, such as egg yolk lecithin, soy lecithin and other phospholipids, propylene glycol diesters, oleic acid, or monoglycerides (e.g., acetyl monoglycerides). The oil phase may also be a mixture of the above ingredients.
Preferred lipid phases are soybean oil, Medium Chain Triglycerides (MCT), olive oil and fish oil, one or more mixtures thereof.
The most preferred oil phase is soybean oil. The preferred range for oil carriers is 0.5 to 50%, most preferably 5 to 20%.
The surfactant can be selected from any pharmaceutically acceptable ionic surfactant, preferably lecithin extracted from egg yolk or soybean, synthetic lecithin or lecithin extracted from plant. Hydrogenated derivatives, such as hydrogenated lecithin (egg yolk) and hydrogenated lecithin (soy) may also be used.
The nanoemulsion may also include co-surfactants that act synergistically on the ionic surfactants, which can alter the surface tension to allow the nanoemulsion to form.
The cosurfactant can be selected from any pharmaceutically acceptable surfactant, and is not limited to nonionic surfactants such as poloxamers (e.g., poloxamers 188 and 407), poloxamines, polyoxyethylene stearates, polyoxyethylene sorbitan fatty acid esters, or sorbitan fatty acid esters, but can also be derivatized with ionic surfactants such as cholic and deoxycholic acids or surface active derivatives and salts thereof. The external aqueous phase does not contain ethanol as a co-surfactant or co-solvent. The proportion of the cosurfactant in the emulsion of the invention is 0.01-30 w/v% of the composition. The ratio of the surfactant to the co-surfactant is 10:0.1-0.1:10, 10:1-1:5 or 5:1-1: 5.
The preferred range of the aqueous phase is 50-99%.
The formulation of the emulsion may contain active ingredients (S) in pharmaceuticals, nutraceuticals, foods and cosmetics, antioxidants, chelating agents, penetrants, suspending agents, preservatives and buffers. In some embodiments, the emulsion may also contain solubilizers (excluding ethanol), chelating agents, preservatives, antioxidants, stabilizers, suspending agents, pH adjusters or elasticity modifiers, such as glycerin, polymers as suspending agents, sweeteners, and the like. The stabilizer may act as a pH adjuster, an anti-emulsifier or an anti-foaming agent, or an additive to stabilize the nanoemulsion.
The active ingredient in the nanoemulsion may account for 0-50%.
The proportion of other ingredients than the active ingredient in the nanoemulsion may be 0-50% w/w.
The emulsion is ideally an optically clear (transparent or translucent) stable system with an average droplet size of less than 100 nm. The average droplet size of the emulsion is preferably less than 100 nm; most preferably less than 175 nm.
The preferred pH range for the finished and stored emulsion is below 10. The pH adjusting agent may be a buffer or sodium hydroxide or other pH adjusting agent, or a combination thereof.
The emulsion of the invention can be prepared by the following steps: water phase, placing water in a container, heating to about 40-80 deg.C and adjusting pH to 1-10. Oil phase, placing the oil in another container, and heating to about 40-80 deg.C. The surfactant and co-surfactant are added to the oil phase and heated to about 40-80 deg.C. Alternatively, surfactants and co-surfactants may be added to the aqueous phase. The aqueous phase and the oil phase are mixed by a high shear mixer to form a coarse emulsion. Homogenizing by an ultrasonic or high-pressure homogenizer or a microfluidization device (with the pressure of 5000-. The nanoemulsion is pH adjusted using a pH adjuster (e.g., sodium hydroxide) to obtain the final product. The sample is filtered, dispensed into containers, covered and capped, typically with nitrogen. The product can be produced by aseptic processing or terminal sterilization. The preferred product is a sterile, stable emulsion obtained by autoclaving. In one embodiment, the emulsion is autoclaved at 121 ℃ for 15-20 minutes. In another embodiment, the emulsion is prepared under aseptic conditions without autoclaving.
One embodiment of the present invention is a method for preparing a nanoemulsion (containing an active ingredient) for human or animal therapy, comprising the steps of: a) an oil-in-water liquid nanoemulsion composition is prepared by i) adding the therapeutically active ingredient or other ingredients to the oil phase, mixing well until completely dissolved; ii) dispersing the oil phase containing the active ingredients in the water phase by ultrasonic or homogeneous dispersion to form a nano-emulsion; b) the nanoemulsion is administered to a human or animal.
One embodiment of the present invention is a method for preparing a nanoemulsion (containing an active ingredient) for human or animal therapy, comprising the steps of: a) preparing an oil-in-water liquid nanoemulsion composition by i) ultrasonically or homogenously dispersing an oil phase in an aqueous phase to form a nanoemulsion; ii) adding the therapeutically active ingredient or other ingredients to the oil-in-water nanoemulsion; iii) mixing thoroughly to dissolve the active ingredient or other ingredients in the oil phase; b) the nanoemulsion is administered to a human or animal.
Another embodiment of the present invention is a method for preparing a nanoemulsion (containing an active ingredient) for human or animal therapy, comprising the steps of: a) preparing an oil-in-water liquid macroemulsion composition by i) dispersing an oil phase in an aqueous phase to form a macroemulsion; ii) adding the therapeutically active ingredient or other ingredients to the oil-in-water macroemulsion of i); iii) mixing thoroughly to dissolve/disperse the active ingredient or other ingredients in the oil phase; iv) preparing a liquid oil-in-water nanoemulsion by ultrasound or homogenization; b) the nanoemulsion is administered to a human or animal.
The invention is described by way of non-limiting examples, including combinations of the examples and aspects detailed. Thus, the present invention also includes combinations and subcombinations of the various elements of the embodiments or aspects described therein. Other features, advantages and embodiments of the invention will be apparent to those skilled in the art from the following description and accompanying examples. The present invention is directed to improvements and modifications in the elements and methods well known to those skilled in the art. Furthermore, the identification and illustration of the embodiments are for illustrative purposes only and are not meant to be exclusive or limited to what is described herein. Various modifications and alterations of this invention will become apparent to those skilled in the art without departing from the spirit of this invention.
Examples
Example 1 preparation of active ingredient free nanoemulsion Using Soybean oil
| Prescription | Quality of |
| g |
| Soybean oil | 20.0 |
| Egg yolk lecithin | 7.2 |
| Tween 80 | 7.2 |
| Sodium hydroxide | Proper amount to pH 6-9 |
| Water for injection | To 100g |
All preparation procedures were completed under nitrogen atmosphere.
The oil-in-water type aqueous nano-emulsion is prepared as follows:
1. the aqueous phase was prepared with water for injection, stirred and heated to about 60 ℃.
2. The aqueous phase was filtered through a 0.22 micron filter and added to the mixing vessel.
3. In addition, an oil phase was prepared using soybean oil filtered through a 0.22 micron filter membrane, tween 80 and egg yolk lecithin, and the mixture was stirred at about 60 ℃ until all dissolved.
4. The oil phase was added to the aqueous phase.
5. Mixing with high shear mixer (Polytron PT3100) at 10,000rpm for 5min to obtain coarse emulsion with large particles, and adjusting pH to 6-9.
6. The mixture was homogenized using a high pressure homogenizer (APV 2000) at a pressure of 5,000-30,000psi until the target particle size was reached.
7. The obtained oil-in-water type nanoemulsion is cooled, adjusted to pH 6-9 if necessary, and transferred to a filling container.
8. The nanoemulsion was filtered through a 0.22 micron filter membrane and transferred to a vessel under nitrogen.
Example 2 preparation of a nanoemulsion containing cyclosporine as an active ingredient Using Soybean oil
All preparation procedures were completed under nitrogen atmosphere.
The oil-in-water type aqueous nano-emulsion is prepared as follows:
1. the aqueous phase was prepared with glycerol and water for injection, stirred and heated to about 60 ℃.
2. The aqueous phase was filtered through a 0.22 micron filter and added to the mixing vessel.
3. In addition, an oil phase was prepared using soybean oil, cyclosporine, tween 80 and egg yolk lecithin filtered through a 0.22 micron filter membrane, and the mixture was stirred at about 60 ℃ until all dissolved.
4. The oil phase was added to the aqueous phase.
5. The crude emulsion was obtained by mixing with a high shear mixer (Polytron PT3100) at 10,000rpm for 5min, and the pH was adjusted to 6-9.
6. The mixture was homogenized using a high pressure homogenizer (APV 2000) at a pressure of 5,000-30,000psi until the target particle size was reached.
7. The obtained oil-in-water type nanoemulsion is cooled, adjusted to pH 6-9 if necessary, and transferred to a filling container.
8. The nanoemulsion was filtered through a 0.22 micron filter membrane and transferred to a vessel under nitrogen.
Dry eye syndrome (KCS) is a disease caused by dry eye, and may be caused by decreased tear secretion or increased tear film evaporation. Commonly found in humans and certain animals. KCS is the most common ophthalmological disease with a prevalence of 5-6%, in post-menopausal women up to 6-9.8%, and in elderly up to 34%. Inflammation due to the feedback of the high permeability of the tear film may be achieved using a topical immunosuppressive agent, such as cyclosporine. Thus, the formulations described in this and the following examples are useful for increasing tear fluid in patients with KCS-associated ocular inflammation.
Example 3 preparation of a nanoemulsion containing cyclosporine as an active ingredient using soybean oil/medium chain triglycerides
All preparation procedures were completed under nitrogen atmosphere.
The oil-in-water type aqueous nano-emulsion is prepared as follows:
1. the aqueous phase was prepared with glycerol and water for injection, stirred and heated to about 60 ℃.
2. The aqueous phase was filtered through a 0.22 micron filter and added to the mixing vessel.
3. In addition, an oil phase was prepared using soybean oil and medium chain triglyceride (miglyoyl 812), cyclosporine, tween 80 and egg yolk lecithin filtered through a 0.22 micron filter membrane, and the mixture was stirred at about 60 ℃ until all dissolved.
4. The oil phase was added to the aqueous phase.
5. Mixing with a high shear mixer (Polytron PT3100) at 10,000rpm for 5min to obtain a crude emulsion, and adjusting the pH to 6-9 if necessary.
6. The mixture was homogenized using a high pressure homogenizer (APV 2000) at a pressure of 5,000-30,000psi until the target particle size was reached.
7. The obtained oil-in-water type nanoemulsion is cooled, adjusted to pH 6-9 if necessary, and transferred to a filling container.
8. The nanoemulsion was filtered through a 0.22 micron filter membrane and transferred to a vessel under nitrogen.
Example 4 dilution example 1 preparation of nanoemulsion
| Prescription | Quality of |
| g |
| Soybean oil | 5.0 |
| Egg yolk lecithin | 1.8 |
| Tween 80 | 1.8 |
| Sodium hydroxide | Proper amount to pH 6-9 |
| Water for injection | To 100g |
All preparation procedures were completed under nitrogen atmosphere.
The oil-in-water type aqueous nano-emulsion is prepared as follows:
1. obtaining a nanoemulsion sample of example 1
2. Diluting the nanoemulsion (1:3v/v) with purified water, and mixing well
3. Adjusting pH to 6-9 if necessary, and mixing
4. The nanoemulsion was filtered through a 0.22 micron filter membrane and transferred to a vessel under nitrogen.
Example 5 dilution example 2 preparation of nanoemulsion
| Prescription | Quality of |
| g |
| Cyclosporin | 0.05 |
| Soybean oil | 5 |
| Egg yolk lecithin | 1.8 |
| Tween 80 | 1.8 |
| Glycerol | 2.25 |
| Sodium hydroxide | q.s.to pH 6-9 |
| Water for injection | to 100g |
All preparation procedures were completed under nitrogen atmosphere.
The oil-in-water type aqueous nano-emulsion is prepared as follows:
1. obtaining a nanoemulsion sample of example 2
2. Diluting the nanoemulsion (1:3v/v) with purified water containing 2.25% glycerol, and mixing well
3. Adjusting pH to 6-9 if necessary, and mixing
4. The nanoemulsion was filtered through a 0.22 micron filter membrane and transferred to a vessel under nitrogen.
Example 6 dilution example 3 preparation of nanoemulsion
All preparation procedures were completed under nitrogen atmosphere.
The oil-in-water type aqueous nano-emulsion is prepared as follows:
1. obtaining a nanoemulsion sample of example 2
2. Diluting the nanoemulsion (1:3v/v) with purified water containing 2.25% glycerol, and mixing well
3. Adjusting pH to 6-9 if necessary, and mixing
4. The nanoemulsion was filtered through a 0.22 micron filter membrane and transferred to a vessel under nitrogen.
Example 7 preparation of an emulsion using Soybean oil and lecithin (control group)
| Prescription | Quality of |
| g |
| Soybean oil | 20.0 |
| Egg yolk lecithin | 12 |
| Glycerol | 2.25 |
| Sodium hydroxide | Proper amount to pH 6-9 |
| Water for injection | To 100g |
All preparation procedures were completed under nitrogen atmosphere.
The oil-in-water type aqueous nano-emulsion is prepared as follows:
1. the aqueous phase was prepared with glycerol and water for injection, stirred and heated to about 60 ℃.
2. The aqueous phase was filtered through a 0.22 micron filter and added to the mixing vessel.
3. In addition, the oil phase was prepared using soybean oil and egg yolk lecithin filtered through a 0.22 micron filter and the mixture was stirred at about 60 ℃ until all dissolved.
4. The oil phase was added to the aqueous phase.
5. Mixing with high shear mixer (Polytron PT3100) at 10,000rpm for 5min to obtain large emulsion, and adjusting pH to 6-9.
6. The mixture was homogenized using a high pressure homogenizer (APV 2000) at a high pressure of 5,000-30,000psi until no further reduction in particle size was achieved.
7. The obtained oil-in-water type nanoemulsion is cooled, adjusted to pH 6-9 if necessary, and transferred to a filling container.
8. The nanoemulsion was filtered through a 0.45 micron filter membrane and transferred to a vessel under nitrogen.
Example 8 preparation of an emulsion Using Soybean oil, lecithin and Tween 80
| Prescription | Quality of |
| g |
| Soybean oil | 20 |
| Egg yolk lecithin | 12 |
| Tween 80 | 2.4 |
| Sodium hydroxide | Proper amount to pH 6-9 |
| Water for injection | To 100g |
All preparation procedures were completed under nitrogen atmosphere.
An oil-in-water type aqueous emulsion was prepared by the method of example 1, and the intensity-uniform average particle size of the emulsion in the above invention was 68nm (D50) as measured by a dynamic light scattering apparatus.
Example 9 preparation of an emulsion Using Soybean oil, lecithin and Tween 80
| Prescription | Quality of |
| g |
| Soybean oil | 35.0 |
| Egg yolk lecithin | 2.52 |
| Tween 80 | 12.6 |
| Sodium hydroxide | Proper amount to pH 6-9 |
| Water for injection | To 100g |
All preparation procedures were completed under nitrogen atmosphere.
An oil-in-water type aqueous emulsion was prepared by the method of example 1, and the intensity-uniform average particle size of the emulsion in the above invention was 99nm (D50) as measured by a dynamic light scattering apparatus.
Example 10 preparation of an emulsion Using Soybean oil, lecithin and Poloxamer F68
| Prescription | Quality of |
| g |
| Soybean oil | 13.3 |
| Egg yolk lecithin | 4.8 |
| Poloxamer F68 | 4.8 |
| Glycerol | 6.7 |
| Sodium hydroxide | Proper amount to pH 6-9 |
| Water for injection | To 100g |
All processing stages are carried out under nitrogen.
An oil-in-water type aqueous emulsion was prepared by the method of example 1, and the intensity-uniform average particle size of the emulsion in the above invention was 67nm (D50) as measured by a dynamic light scattering apparatus.
Example 11 preparation of an emulsion Using Soybean oil, lecithin and Tween 80
| Prescription | Quality of |
| g |
| Soybean oil | 5.0 |
| Egg yolk lecithin | 3.0 |
| Tween 80 | 0.6 |
| Glycerol | 2.25 |
| Sodium hydroxide | Proper amount to pH 6-9 |
| Water for injection | To 100g |
All preparation procedures were completed under nitrogen atmosphere.
The nanoemulsion of example 8 was diluted with glycerol solution to prepare an oil-in-water type aqueous emulsion in which the intensity uniformity of the emulsion in the above invention was 67nm (D50) as measured by dynamic light scattering.
Example 12 preparation of an emulsion Using Soybean oil, lecithin and Poloxamer F68
| Prescription | Quality of |
| g |
| Soybean oil | 20.0 |
| Egg yolk lecithin | 7.2 |
| Poloxamer F68 | 1.27 |
| Glycerol | 10 |
| Sodium hydroxide | Proper amount to pH 6-9 |
| Water for injection | To 100g |
All preparation procedures were completed under nitrogen atmosphere.
An oil-in-water type aqueous emulsion was prepared by the method of example 1, and the intensity-uniform average particle size of the emulsion in the above invention was 89nm (D50) as measured by a dynamic light scattering apparatus.
Example 13 preparation of an emulsion Using Soybean oil, lecithin and Poloxamer F68
| Prescription | Quality of |
| g |
| Soybean oil | 20.0 |
| Egg yolk lecithin | 7.2 |
| Poloxamer F68 | 4 |
| Glycerol | 10 |
| Sodium hydroxide | Proper amount to pH 6-9 |
| Water for injection | To 100g |
All preparation procedures were completed under nitrogen atmosphere.
An oil-in-water type emulsion in water was prepared by the method of example 1, and the intensity-uniform average particle size of the emulsion in the above invention was 75nm (D50) as measured by a dynamic light scattering apparatus.
Example 14 characterization of the particle size distribution of the nanoemulsion Using a Malvern NanoZetasizer
Comparing the emulsion prepared using egg yolk lecithin as a single surfactant (example 7) with the emulsion of the present invention (example 1), it was found that the emulsion of the present invention had an intensity-uniform average particle size of-47 nm (D50), was optically clearly visible and was transparent (FIG. 2), whereas the emulsion prepared using egg yolk lecithin as a single surfactant (example 7) was milky and had an intensity-uniform average particle size of-177 nm (D50).
Example 15 stability of nanoemulsion prepared with LCT/egg lecithin and Tween 80
The emulsion prepared with lecithin/tween 80 (example 1) was stored at 40 ℃/75% RH for 5 months and the particle size distribution was examined (figure 3) and found to be unchanged in particle size and to remain optically clear.
Example 16 stability of nanoemulsion prepared with LCT/egg lecithin and Tween 80 with active ingredients
The cyclosporine emulsion prepared with lecithin/tween 80 (example 2) was stored at 40 ℃/75% RH for 5 months and the particle size distribution was examined (figure 4) and no significant change in particle size was found. The primary average particle size was 38nm and 45nm after standing at 40 ℃ for 5 months. The emulsion is optically transparent, and the chemical stability of cyclosporine is not different.
| Initial | 3 months old | For 5 months |
| Content (mg/mL) | 1.7 | 1.7 | 1.8 |
| D50(nm) | 38 | 37 | 45 |
Example 17 stability of nanoemulsion prepared with Mixed oil, egg yolk lecithin and Tween 80 with active ingredients
An emulsion prepared with LCT/MCT/lecithin/tween 80 and the active ingredient (example 3) was stored at 40 ℃/75% RH for 5 months at 2-8 ℃ and the particle size distribution was examined (fig. 5), and it was found that there was no change in particle size and the emulsion was optically clear.
Example 18 stability of nanoemulsion prepared with Mixed oil, egg yolk lecithin and Tween 80 with active ingredients
The emulsion (example 5) prepared using LCT/MCT/lecithin/Tween 80 and cyclosporin, an active ingredient, was stored at room temperature (-25 ℃) and refrigerated conditions (-2-8 ℃) for 14 months, and the particle size distribution thereof was examined (FIG. 6), and it was found that the average particle sizes were all below 50 nm. The emulsion is optically transparent, and the chemical stability of cyclosporine is not different.
| Initial | 14 months @5 deg.C | 14 months @25 deg.C |
| Content (mg/mL) | 0.45 | -- | 0.48 |
| D50(nm) | 38 | 39 | 49 |