KR102142916B1 - Method for Manufacturing Nanosuspension Comprising Insoluble Drug Using Bottom-up Method, and Nanosuspension Made thereby - Google Patents

Abstract

The present invention relates to a method for preparing a nanosuspension comprising the step of dissolving a poorly soluble drug in a solvent, Transcutol, and a nanosuspension of a poorly soluble drug prepared therefrom, and has a low toxicity instead of a conventional organic solvent and By using the applicable Transcutol, it is possible to prepare a nano suspension without additionally removing the solvent, and the average diameter of the particle size of the nanosuspension is less than 300 nm, so long-term storage for more than 6 months is possible. It can be used usefully as a product.

Description

Method for Manufacturing Nanosuspension Comprising Insoluble Drug Using Bottom-up Method, and Nanosuspension Made thereby}

The present invention relates to a method for preparing a nanosuspension containing a poorly soluble drug through a bottom-up method, and to a nanosuspension prepared therefrom.

Resveratrol (3,5,4'-trihydroxystilbene) is a non-flavonoid polyphenolic substance found primarily in grapes, peanuts, mulberries, cranberries, blueberries and red wines, and is generally one of the dietary sources of people. It is consumed as a part. It has been reported that resveratrol can prevent coronary heart disease by inhibiting myocardial infarction, arrhythmia, high blood pressure, hypertrophy, fibrosis, atherosclerosis, and thrombosis. Has a beneficial biological effect. Resveratrol is considered a class II compound in the Biopharmaceutical Classification System (BCS) because it exhibits high permeability (Log P=3.1) and low water solubility. The physicochemical properties of resveratrol are as follows: molecular weight is 228.25 g/mol, melting point is 253 to 255°C, pK _a value is 9.22, it is insoluble in water and is soluble in methanol, ethanol and DMSO.

Due to its instability, low water solubility, short biological half-life, and rapid metabolism and elimination properties, the therapeutic application of resveratrol is very limited. In order to overcome these limitations of resveratrol, a number of strategies have been evaluated, such as the use of polymer nanoparticles, solid lipid nanoparticles, self-emulsifying drug delivery systems, nanoemulsions, liposomes, nanosuspensions and nanofibers.

Among these, nanosuspension refers to a colloidal dispersion of stabilized drug particles under conditions in which a polymer, a surfactant, or both are present. Nanosuspensions are used to deliver drug substances that exhibit low water solubility and lipid solubility. The small particles of the nanosuspension provide a very large drug surface area, increasing the dissolution rate of the insoluble drug. As a result, BCS class II and IV compounds exhibit improved bioavailability, rapid activity, and other desirable biopharmaceutical effects. Such nano-suspension technology can be broadly divided into a top-down method and a bottom-up method.

The top-down method reduces the particle size of the drug through milling (jet mill and ball mill) and homogenization under high pressure. However, this top-down method is very difficult to apply to materials exhibiting heat instability, because this method requires high energy to generate heat. In addition, large amounts of energy can also produce amorphous particles and deformed crystal structures.

The bottom-up method includes inducing precipitation of particles from the supersaturated drug solution. Various techniques such as solvent evaporation, supercritical fluid, anti-solvent precipitation, and chemical precipitation are applicable to the bottom-up method. These methods require lower energy pressure than top-down methods and can be applied to thermodynamically unstable materials. Therefore, in the present invention, the nanosuspension was prepared by a bottom-up method, but it can be said that it is important to control the residual solvent and particle growth by using this technique. Inadequate control of particle growth is due to incomplete understanding of formulation composition and manufacturing process. Therefore, the manufacturing process and formulation composition should be understood using a scientific and systematic method. In the present invention, quality by design (QbD) based on drug design was applied to develop a nano suspension.

The U.S. Food and Drug Administration (FDA) recommends applying the QbD principle in the areas of risk-based approaches and drug discovery, drug manufacturing and regulation. The FDA's focus on QbD is that the increase in test does not necessarily improve product quality. In the past few years, in the pharmaceutical sector, QbD has published ICH Q8 (pharmaceutical development), ICH Q9 (quality risk management), and ICH Q10 (pharmaceutical quality system). They provide high-level guidance on the scope and definition of QbD for the pharmaceutical industry.

In the ICH Q8 guidelines, QbD is defined as a systematic approach to development, starting with predefined goals, and based on sound science and quality risk management (ICH Q8, 2009), as well as an understanding of the product and process, as well as the process. Emphasize the control of In order to implement QbD, the quality target product profile (QTPP), and critical quality attributes (CQAs) must be defined. Critical material attributes (CMAs) and critical process variables (CPPs) and prior knowledge that affect CQAs based on risk assessments (RA) should also be identified. A design space should be established after the design of experiment (DoE) and risk analysis. The most important goal in product design and understanding is to develop a robust product that can deliver the desired QTPP during the product's life cycle. The product produced through QbD can improve stability, reduce production cost, increase patient efficacy, and minimize side effects.

Currently, the research, development, manufacturing, storage and clinical process development of nanosuspensions are still in infancy, so it is useful and necessary to apply QbD. The major barriers in the manufacture and clinical application of nanosuspensions include structural instability and incomplete understanding of the manufacturing process, so the development of a technology that can overcome these barriers is necessary.

Korean Patent Registration No. 10-1171375 (registered on July 31, 2012).

An object of the present invention is to provide a manufacturing method capable of efficiently preparing a nano-suspension of poorly soluble drugs while omitting the solvent removal process because an organic solvent is not used.

Another object of the present invention is to provide a nanosuspension of a poorly soluble drug that can be stored for a long time and has excellent bioavailability and drug release rate.

In order to achieve the above object, the present invention provides a method for preparing a nano-suspension comprising the step of dissolving a poorly soluble drug in a solvent, Transcutol.

In order to achieve the above other object, the present invention provides a nanosuspension prepared by the above manufacturing method.

The present invention relates to a method for preparing a nanosuspension comprising the step of dissolving a poorly soluble drug in a solvent, Transcutol, and a nanosuspension of a poorly soluble drug prepared therefrom, and has low toxicity instead of a conventional organic solvent and By using the applicable Transcutol, it is possible to prepare a nano suspension without additionally removing the solvent, and the nanosuspension prepared in this way contains particles with an average diameter of less than 300 nm and is stored for a long time for more than 6 months. As this is possible, it can be usefully utilized as a drug product.

1 is a diagram illustrating a process of preparing a nanosuspension prepared by an anti-solvent precipitation method in an embodiment of the present invention.
Figure 2 shows the solubility of resveratrol in various solvents at 298.15K.
Figure 3 shows the solubility of resveratrol for the trans kyutol ^® HP and water mixture at 288.15 to 313.15K.
4 shows the value T _hm of ΔH° with respect to ΔG° Value (300.40 K), different transport kyutol ^® HP and the solubility of the resveratrol enthalpy of the water in the mixture-entropy correction (correction) shows the results of the analysis.
Figure 5 is the value of the ΔH TΔS ° ° for the T _hm Value (300.40 K), different transport kyutol ^® HP and the solubility of the resveratrol enthalpy of the water in the mixture-entropy correction (correction) shows the results of the analysis.
Figure 6 shows the inhibitory effect of povidone on the precipitation of resveratrol.
Figure 7 shows the inhibitory effect of HPMC and HPC on resveratrol precipitation.
8 shows a response curve plot showing the effect of various formulation parameters on response values Y ₁ , Y ₄ and Y ₉ (X ₁ : Concentration of PVP VA64 (mg/mL), X ₂ : Concentration of PVP K12) (mg/mL), SLS concentration (mg/mL), Y ₁ : particle size (z-average, day 1), day), Y ₄ : particle size (z-average, day 7), Y ₉ : Zeta Potential).
9 shows overlay plots for PVP VA64 (X ₁ ) and PVP K12 (X ₂ ) at the lowest (A), medium (B), or high (C) concentrations of SLS (X ₃ ).
10 is a target reaction value for reaction values Y ₁ -Y ₉ when the SLS concentration is 0.5 mg/mL (A), 1.0 mg/mL (B) and 1.5 mg/mL (C) by Monte Carlo simulation (Y ₁ : particle size (z-average, initial), Y ₂ : particle size (z-average, day 1), Y ₃ : particle size (z-average, day 3), Y ₄ : particle size (z-average, day 7), Y ₅ : particle size (d90, initial), Y ₆ : particle size (d90, day 1), Y ₇ : particle size (d90, day 3), Y ₈ : particle size (d90, day 7), Y ₉ : zeta potential).
11 is a probability map that satisfies the target response values for the response values Y ₁ -Y ₉ when the mixing speed is 500 rpm (A), 750 rpm (B) and 1,000 rpm (C) by Monte Carlo simulation. (Y ₁ : particle size (z-average, initial), Y ₂ : particle size (z-average, day 1), Y ₃ : particle size (z-average, day 3), Y ₄ : particle size ( z-average, day 7), Y ₅ : particle size (d90, initial), Y ₆ : particle size (d90, day 1), Y ₇ : particle size (d90, day 3), Y ₈ : particle size (d90 , Day 7), Y ₉ : Zeta potential).
12 shows the particle growth trend of the nanosuspension prepared by the complete factor experiment (the formulations and process conditions of Run-1 to Run-9 are as shown in Table 16).
13 shows the resveratrol profile dissolved from the optimal nanosuspension formulation and resveratrol raw material.
14 shows the plasma concentration-time profile of resveratrol in rats after oral administration of the resveratrol nanosuspension and resveratrol raw material according to the present invention (data are shown as mean±standard deviation [n=4]).
15 shows the TEM image and particle size distribution of the resveratrol nanosuspension (A: initial (day 1) TEM image), B: TEM image after 6 months, C: initial (day 1) particle size distribution, D: 6 Particle size distribution after month).
16 is a result of confirming the long-term stability of the optimized resveratrol nanosuspension (A: particle size [z-average and d90], B: drug content).

Hereinafter, the present invention will be described in more detail.

As shown in Figure 1, the inventors of the present invention show that a poorly soluble drug such as resveratrol or MHY498 exhibits high solubility in Transcutol, and when a combination of a specific surfactant and a water-soluble polymer is used as a surface stabilizer, the average particle is 300 The present invention was completed by confirming that it was possible to prepare a nanosuspension containing nanoparticles less than nm and retaining this shape for more than 6 months.

Accordingly, the present invention provides a method for preparing a nanosuspension comprising the step of dissolving a poorly soluble drug in a solvent, Transcutol.

The poorly soluble drugs are resveratrol, MHY498((Z)-5-(2,4-dihydroxybenzylidene)thiazolidine-2,4-dione), aprepitant, cyclosporine, celecoxib, coenzyme Q10, It may be selected from the group consisting of valsartan, dabigatran etexilate, sorafenib, and silymarin, but is not limited thereto.

In addition, the poorly soluble drug may be included in an amount of 1 to 100 parts by weight based on 100 parts by weight of the solvent.

In more detail, a nanosuspension of a poorly soluble drug can be prepared using a solvent/anti-solvent precipitation method, and transcutol as the solvent and water as an anti-solvent may be used, and in detail, the transcutol and water Silver may be used in a weight ratio of 1:5 to 1:100, and more specifically, may be used in a weight ratio of 1:10 to 1:20.

In one embodiment of the present invention, after the step of dissolving the poorly soluble drug, dissolving a water-soluble polymer in water to prepare an aqueous polymer solution; And injecting a solution in which the drug is dissolved in the aqueous polymer solution, wherein the water-soluble polymer forms a layer on the surface of the drug particle, thereby stabilizing the particles to prevent aggregation of the drug particles.

Specifically, the water-soluble polymer is selected from the group consisting of polyvinylpyrrolidone K12, polyvinylpyrrolidone K17, polyvinylpyrrolidone K25, polyvinylpyrrolidone K30, and polyvinylpyrrolidone vinyl acetate VA64. It may be one or more, and preferably, polyvinylpyrrolidone K12, polyvinylpyrrolidone vinyl acetate VA64, or a mixture thereof.

In one embodiment of the present invention, after the step of dissolving the poorly soluble drug, preparing an aqueous surfactant solution by dissolving a surfactant in water; And injecting a solution in which the drug is dissolved in the aqueous surfactant solution, wherein the surfactant is adsorbed at the solid-liquid interface to reduce hydrophobic interactions and coagulation, thereby inhibiting particle growth and aggregation. I can.

Specifically, the surfactant is sodium lauryl sulfate (SLS), sucrose fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene glycolated natural or hydrogenated castor oil, polyoxyethylene sorbitan fatty acid ester (e.g., Tween (Tweens), polyethylene glycol, polyoxyethylene stearate, polyoxyethylene-polyoxypropylene copolymers (e.g., poloxamer, poloxamine, etc.), polyethylene glycolated phospholipids, and alpha-tocopherol It may be selected from the group consisting of polyethylene glycol succinate, and preferably sodium lauryl sulfate.

The nanosuspension prepared by the above preparation method may contain poorly soluble drug nanoparticles having an average diameter of 1 nm to 300 nm, and the nanoparticles have a larger surface area and are more easily dissolved, thus maximizing bioavailability. .

Meanwhile, in all of the above processes, a process of mixing and mixing a poorly soluble drug with a solvent, a water-soluble polymer, or a surfactant may be included, and a nanosuspension may be prepared at a mixing speed of 500 rpm to 1,000 rpm.

In addition, the present invention prepares a nanosuspension prepared by the above manufacturing method.

Since the nanosuspension according to the present invention may contain poorly soluble drug nanoparticles having an average diameter of 1 nm to 300 nm, the bioavailability of poorly soluble drugs can be maximized.

Hereinafter, examples will be described in detail to help understanding of the present invention. However, the following examples are provided to more fully explain the present invention to those having average knowledge in the art, and are merely illustrative of the contents of the present invention, so the scope of the present invention is limited to the following examples. no.

< Example 1> Resveratrol Preparation of nano suspension

One. Resveratrol Research before formulation of nanosuspension ( Preformulation )

One) Of resveratrol variety Solubility in solvent

The solubility of resveratrol in various organic solvents was measured. A sample was prepared by adding an excess of resveratrol to a glass bottle containing 10 mL of an organic solvent. At this time, each experiment was repeated three times. The sample was vortexed for 5 minutes to mix well, and then sonicated in an ultrasonic bath (model 5800, Branson, Danbury, USA) for 1 hour. Thereafter, the glass bottle was placed in a shaking constant temperature water bath (BS-21, Jeiotech Co Ltd, Daejeon, Korea) at 24° C. for 24 hours. Thereafter, the saturated solution was filtered through a 45 μm RC syringe filter, transferred to a volumetric flask, and diluted with methanol. Next, the concentration of resveratrol is determined by UV/VIS detector SPD-20A, Communication Bus Module CBM-20A, AUTO Sampler SIL-20AC, liquid chromatograph LC-20AT, and degassing unit for data collection and overall system adjustment. It was measured using a Shimadzu HPLC system with DGU-20A 5R, and column oven CTO-20A (Shimadzu, Tokyo, Japan). Chromatographic separation was performed through a Gemini C18 reversed-phase column (Phenomenex, 150 mm Х 4.6 mm, 5 μm). The injection volume was 10 μL. The mobile phase was composed of water and acetone (60:40, v/v), the flow rate was 0.8 mL/min at 30°C, and the ultraviolet ray region was detected at a wavelength of 303 nm. A calibration curve was prepared by measuring the peak area of a known resveratrol reference at a concentration of 1 μg/mL to 100 μg/mL.

2) Of resveratrol menstruum Solubility in mixture

The mole fraction solubility of the resveratrol in the pure water (w = 0) and pure trans kyutol ^® HP (w = 1.0) trans kyutol ^® HP and co-solvent mixture of water (mass fraction w = 0.1-0.9) including measured. The solubility was measured using a shaking constant temperature water bath at a temperature of 288.15 K to 313.15 K. Excess of resveratrol was prepared sample was added to the vial trans kyutol ^® HP and water mixture containing 15 mL of pure solvent contained. At this time, each experiment was repeated four times. The sample was vortexed for 5 minutes to mix well, and then sonicated in an ultrasonic bath ((model 5800, Branson, Danbury, USA) for 1 hour. After that, the glass bottle was 288.15 K to 313.15 K in a shaking constant temperature water bath. It was shaken for 48 hours under the condition of temperature of. According to a preliminary experiment, this period of time was sufficient to reach full saturation The supernatant of the sample was filtered through a 0.45 μm RC syringe filter and the filtrate was methanol Finally, the concentration of resveratrol was analyzed by performing the HPLC method at 303 nm under the same conditions as described above.

Next, the mole fraction solubility ( χ _e ) of the experimental resveratrol was calculated using the following equations (1) and (2):

In the above formula 1 and 2, m ₁ is the resveratrol mass in a saturated solution, m ₂ is the mass of the transformer kyutol ^® HP, m ₃ indicates a mass of water, M _1, M ₂ and M ₃ are each resveratrol, trans Cutol ^® means the molecular weight of HP and water. Formula (1) of the χ _e in pure solvent (trans kyutol ^® HP and water) Values, equation (2) of trans-resveratrol from kyutol ^® HP and mixtures χ _e It was applied to calculate the value.

In addition, to measure the correlation between physicochemical properties, the Jouyban-Acree model was applied to a mixed solvent system. The general equation for predicting solute solubility in water co-solvent mixtures at various temperatures is as follows:

,

, 및

은 각각 온도 T(켈빈)의 혼합 용매, 트랜스큐톨^® HP 및 물에서의 레스베라트롤의 몰분율이며, w ₁ 및 w ₂는 트랜스큐톨^® HP 및 물의 순수한 질량 분율이고, J _i 는 다음 식

을 다음 식(

,

) 및(

)에 대하여 회귀함으로써 계산된 상수이다.In the above formula,

,

, And

Is the mole fraction of resveratrol in a solvent mixture of trans kyutol ^® HP and water of temperature T (Kelvin), respectively, w ₁ and w ₂ are trans kyutol ^® HP and pure water mass fractions, J _i is obtained using the equation

Then the expression (

,

) And(

) Is a constant calculated by regression.

Regarding the logarithm of the molar fraction of resveratrol versus temperature, the Vanthoff equation [Equation (4)], an inverse linear function of the absolute temperature T, was used:

은 레스베라트롤의 몰 분율이고, 반트호프 모델 패러미터(A 및 B)는 최소제곱법분석(least squares analysis)을 통해 계산되었다. 사실 대부분의 진짜 해들(real solutions)은 비이상적인 거동을 나타내었다. 그러므로, 표준 엔탈피 및 엔트로피가 용액 과정에서 고려되어야 한다. 결과적으로, 하기 식 (5)가 얻어졌다:In the above formula,

Is the molar fraction of resveratrol, and the VanThof model parameters (A and B) were calculated through least squares analysis. In fact, most of the real solutions exhibited unusual behavior. Therefore, standard enthalpy and entropy must be considered in the solution process. As a result, the following formula (5) was obtained:

In the above equation, Δ H° and Δ S° are the enthalpy and entropy of the solution, respectively.

Applying a large amount of experimental molar fraction data to the Jouyban-Acree model can provide a more predictable model. Therefore, the formula combined in this way is as the following formula (6):

In the above equation, A ₁ , B ₁ , A ₂ , and B ₂ are constants of the Vant Hoff equation calculated by regression analysis of the mole fraction of resveratrol in a pure solvent at various temperatures.

The mean relative deviation (MRD) was used to confirm the accuracy of the predictive model of the method, and was calculated by the following equation (7):

In the above equation, N denotes the number of data points in each set.

The thermodynamic properties of resveratrol dissolved in the mixed solution were determined through analysis of solubility enthalpy (△ H° ), Gibbs free energy (△ G° ), and dissolution entropy (△ S° ). The thermodynamic change was calculated according to the following equation (8) at the mean harmonic temperature ( T _hm ):

In the above formula, n is the number of temperatures analyzed. Δ H° was calculated using the modified VanThof equation as shown in Equation (9):

In the above formula, X is the molar fraction solubility in the mixed solution, T is the absolute temperature (K), and R is the universal ideal gas constant (8.314 J · K ^-1 · mol ^{- 1} ). The derived enthalpy value (Δ H° ) was calculated using the Vant Hoff equation as shown in Equation (10) below.

Gibbs energy △ G° of resveratrol generated during the dissolution process is T _hm It was calculated through the following equation (11).

Δ S° of resveratrol for the dissolution process was calculated as shown in the following equations (12) and (13) by combining the values of Δ H° and Δ G° .

The relative contribution of enthalpy to Gibbs energy during the dissolution process of resveratrol in the mixed solution (%ζ _H ) and the relative contribution of entropy (%ζ _TS ) were calculated as shown in Equations (14) and (15) below.

3) Polymeric Resveratrol Confirmation of sedimentation inhibition effect

Precipitation of resveratrol in 500 mL of distilled water (including 0.5% (w/v) polymer) was performed at 37°C and 50 rpm using a USP rotating paddle apparatus (Electrolab, Mumbai, India). Was measured. Samples (3 mL) were removed at a specific time, filtered using a 0.45 μm RC syringe filter, and then diluted with methanol. Thereafter, the concentration of resveratrol was analyzed using HPLC in the same manner as described above.

2. Resveratrol Preparation of nanosuspension

Resveratrol nanosuspension was prepared using an anti-solvent precipitation method. In other words, the trans-resveratrol was dissolved kyutol ^® HP. In addition, an aqueous solution was prepared by dispersing a specific concentration of a polymer and a surfactant in distilled water. Then, the obtained trans kyutol ^® HP solution was quickly added to an aqueous solution used as a solvent and stirred to self 750 rpm. Then, a blue suspension was immediately produced, and a resveratrol nanosuspension was produced through stirring.

3. Resveratrol Experimental design for preparation of nanosuspension

One) Formulation Parametric Box- for optimization Behnken design

The Box-Behnken design (BBD) and response surface methodology were used to optimize the composition of the resveratrol nanosuspension. In this design, the experimental area was assumed to be a cube. The experiment was performed at the point corresponding to the center point of each axis, and the repeated experiment was performed at the center of a multidimensional cube. A total of 17 experiments were conducted at three levels of three elements. The central point was verified 5 iterations. Formulation parameters and parameter levels were selected based on the preliminary investigation. That is, PVP VA64 (X ₁ ), PVP K12 (X ₂ ), and SLS (X ₃ ) were selected as formulation parameters (elements) at three levels (-1, 0, +1). The particle size (z-average, d90) and the zeta potential (ζ) were chosen as the response. Formulation parameters and reaction values were as shown in Table 1 below.

These experimental designs were generated and evaluated using Design Expert ^® 11.0 (Stat-Ease, Inc. Minneapolis, USA).

2) process variables Robustness Full factorial method to establish design; FD )

In this experiment, the perfect factor placement method was performed to ensure the robustness of the manufacturing process with an optimized configuration. A total of 9 experiments were performed with 3 elements at two levels, and the center point was performed once. The injection rate, temperature, and mixing speed were selected as process parameters at two levels (-1, +1), and particle size (z-average, d90) and zeta potential (ζ) were used as reaction values. Process variables and reaction values were as shown in Table 2 below.

These experimental designs were generated and evaluated using Design Expert ^® 11.0 (Stat-Ease, Inc. Minneapolis, USA).

< Example 2> Resveratrol Nanosuspension Specialization

1. Particle size and zeta Potential Measure

For characterization of the particle surface, the particle size distribution and zeta potential of the resveratrol nanosuspension were measured at 25° C. using a dynamic light scattering (DLS) device (ELSZ-1000, Photal Otsuka Electronics, Osaka, Japan). The particle size was specified three times and expressed as the z-average diameter. In addition, the hydrodynamic diameter and particle distribution of the nanoparticles in the suspension were determined using DLS. DLS is a fast and sensitive method, especially in the small nanometer range, requiring very small amounts of particles. Therefore, when very few active pharmaceutical ingredients are used, it is a very suitable method for routine measurements and confirming early formation development. Zeta potential is a measure of the electrical charge on the surface of a particle and indicates the physical stability of a colloidal system. Zeta potential was measured using the Laser-Doppler method.

2. Confirmation of morphological features

Morphological analysis was measured at 80 kV using a transmission electron microscopy (TEM, Hitachi H-7600, Tokyo, Japan). First, the suspension was dropped on a carbon film coated with a copper grid, dried in a fume hood at room temperature for 24 hours, and then observed by TEM.

3. In vitro dissolution assay

Dissolution was performed in 500 mL of distilled water at 37±0.5°C using USP Dissolution Apparatus II (paddle method, Electrolab, Mumbai, India) at a paddle speed of 50 rpm. Resveratrol suspension (5 mg/mL, 20 mL) and resveratrol raw material (100 mg) were placed in a dissolution vessel. Thereafter, samples (5 mL) were taken at different time intervals and replaced with fresh dissolution medium maintained at 37±0.5° C. to maintain a constant volume. The extracted samples were filtered using a 0.45 μm RC syringe filter, and then diluted with methanol. The final concentration of resveratrol was determined using HPLC as described above.

4. In rats drug Dynamics Research

Eight male Sprague-Dawley rats (200 ± 10 g; Orient Bio Inc., Seongnam, Korea) were divided into two treatment groups, each of 4 mice. Prior to the experiment, rats were fasted for 18 hours. Resveratrol raw material or resveratrol suspension was orally administered to the two experimental groups so that the dose of resveratrol was 10 mg/kg. The resveratrol raw material was immediately dispersed in 1 mL of water immediately before oral administration. Thereafter, a blood sample (approximately 0.25 mL) was drawn from the rat's neck vein after 0.25, 0.5, 0.75, 1, 1.5, 2, 4, 6, 8, and 12 hours after the administration and collected in heparinized tubes. Thereafter, the blood sample was centrifuged at 8,000 rpm for 10 minutes at 4°C. The resulting plasma was transferred to individual centrifuge tubes and stored at -70°C. The amount of resveratrol was determined by HPLC using a method disclosed in a known paper (Das et al., 2011, Journal of Liquid Chromatography & Related Technologies. 2011;34:1399-1414.).

A resveratrol stock solution (1 mg/mL) and an internal standard carbamazepine (1 mg/mL) were dissolved in methanol to prepare. A standard working solution of resveratrol (50 μg/mL) and carbamazepine (50 μg/mL) in a mixture of water and methanol (1:1) were prepared just before the experiment. The reference working solution (50 μg/mL) of resveratrol was serially diluted with a water-methanol (1:1) mixture, and a known amount (10 μL) of the solution was added to the blank plasma (100 μL) as a reference working solution inside gypsum. (50 μg/mL) 5 μL was added. Plasma samples were then extracted through a liquid-liquid extraction method as follows. Seven calibration solutions were prepared to obtain final concentrations (5 ng/mL to 5,000 ng/mL). Resveratrol and carbamazepine (internal reference) were extracted from plasma using a liquid-liquid extraction method. That is, 40 μL of phosphate-buffered saline (PBS; 30 mM, pH 6) was added to the plasma sample in a 2 mL centrifuge tube, and the contents were well mixed through vortexing for 15 seconds. Finally, ethyl acetate (300 μL) was added and mixed for 30 seconds. After ethyl acetate extraction, the sample was centrifuged at 8,000 rpm for 10 minutes, and the upper organic layer was transferred to a clean tube. This extraction process was performed twice more, and in order to dry the mixed organic layer, it was evaporated using nitrogen gas and a heating block (Eyela MG-2200, Tokyo, Japan) at 35°C. The residue was restored to its original state with 75 μL of the mobile phase, and centrifuged at 13,000 rpm for 10 minutes. Thereafter, the supernatant was transferred to a glass insert previously installed in a glass bottle for an auto sampler. During each assay, 20 μL of the supernatant was injected into the HPLC system.

All of the above processes were performed in the absence of direct light. Chromatographic separation was done through a reversed-phase HPLC column (Phenomenex, EVO 5 μm C18 250 Х 4.6 mm, 100 A), which was protected with a guard column (Phenomenex, Gemini C18, 4 Х 3.0 mm). The assay was carried out by isocratic elution of a mobile phase consisting of acetonitrile and 30 mM PBS (pH 7.0; 30:70 v/v) at a flow rate of 1 mL/min and a temperature of 35°C. Thereafter, the mobile phase was filtered through a 0.2 mm nylon membrane, and all gases were removed through sonication at 25° C. for 20 minutes before use. In addition, the UV absorbance at 306 nm was recorded.

5. Stabilization study

Stabilization studies of resveratrol nanosuspension were conducted at room temperature (25 ± 5 °C, 60% RH) for a period of 6 months. Every month, the resveratrol nanosuspension was characterized in terms of its particle size and drug content.

< Experimental Example 1> Resveratrol Research before formulation of nanosuspension

1. in various solvents Of resveratrol Solubility

The results of confirming the solubility of resveratrol in various solvents are shown in FIG. 2. According to Figure 2, the solubility in tetrahydrofuran and trans kyutol ^® HP were respectively 412.7 mg / g and 332.9 mg / g. Despite the high solubility of resveratrol in tetrahydrofuran, rapid particle agglomeration occurred during preparation of the resveratrol nanosuspension using tetrahydrofuran. Therefore, tetrahydrofuran was not a suitable solvent for preparing nanosuspensions. Transcutol ^® HP has low toxicity and high solubilizing effect. In addition, the trans kyutol ^® HP has the advantage that it does not require a solvent removing process in the manufacturing process of nano-suspensions because it can be absorbed by the small amount. Thus, the selected the trans kyutol ^® HP to the best solvents of the present invention.

2. Transcutol ^® In a mixed solvent of HP and water Of resveratrol Solubility

At a temperature of 288.15 K to 313.15 K, χ _e value of resveratrol in the trans kyutol ^® HP and water mixtures were as shown in Table 3 and Fig.

(w is the weight fraction of the trans kyutol ^® HP)

In other words, the mole fraction solubility of the trans kyutol ^® HP and water mixture was increased with increasing the temperature of the mixture, or trans kyutol ^® HP is increased fraction of the. The highest molar fraction solubility of resveratrol (1.62 Х 10 ^-1 ) was observed in pure Transcutol ^® HP of 313.15 K, and the lowest value (1.29 Х 10 ^-6 ) was observed in pure water of 288.15 K. Since the molar fraction solubility depends on many factors including chemical structure, permittivity, solvent polarity, temperature and cosolvent ratio, the high χ _e value of resveratrol is a low transcutol ^® HP dielectric constant compared to water. It is believed to be due to polarity. Judging from these results, trans kyutol ^® HP can be used as a potential co-solvent compatible with that can improve the solubility of resveratrol water, physiological. Thus, it was confirmed that the nano-suspensions from trans kyutol ^® HP (mass fraction 0.1-0.2) and water mixture can be prepared effectively.

Then, in order to confirm the molar solubility relationship of the saturated solution of resveratrol, the experimental data were applied to the Jouyban-Acree model. As a result, the molar fraction data obtained from the above experiment fit well into the Jouyban-Acree model, and the trained model was as shown in the following equations (16) and (17):

[Equation 16]

=> R ² = 0.986 → F = 4632.3 → p <0.0005 → MRD = 2.12%.

및

의 실험적인 값은 제한되었고, 추가적인 실험 수행이 요구되었다. 이러한 한계를 다루기 위해, Jouyban-Acree 및 반트호프 모델을 조합하여 사용하였다. 결과적으로 다음과 같은 예측 모델이 얻어졌다:The above equation was used to predict the experimental molar fraction with an MRD of 2.12%.

And

The experimental value of was limited, and additional experiments were required. To address these limitations, Jouyban-Acree and VanThof models were used in combination. As a result, the following predictive model was obtained:

[Equation 17]

=> R ² = 0.986 → F = 4631 → p <0.0005 → MRD = 2.12%.

Like the trained equation, the equation inverted the remaining molar fraction value as MRD 2.12% (N=66).

On the other hand, Table 4 shows a summary of the thermodynamic parameters of the study trans kyutol ^® HP + water mixture. Transcutol ^® HP + water mixture dissolution enthalpy (△ H° ) , Gibbs free energy (△ G° ), and dissolution entropy (△ S° ) as thermodynamic parameters during resveratrol dissolution, the equation (8) disclosed in Example 1 above. ) It was calculated as shown in (13). In order to confirm the contribution related to the solubility of enthalpy and entropy, ζ _H (relative contribution of enthalpy) and ζ _TS (relative contribution of entropy) as shown in Equations (14) and (15) disclosed in Example 1, respectively. Calculated.

Referring to Table 4, △ H ° of the transformer kyutol ^® HP and water mixture was a positive value. These results mean that the dissolution of resveratrol in a solvent is an endothermic reaction in the temperature range of the experimental conditions. That is, the interaction between resveratrol and solvent molecules is stronger than that between solvent-solvent molecules and resveratrol-resveratrol molecules.

In the pure trans kyutol ^® HP, dissolution of resveratrol for the pure water and trans kyutol ^® HP and the mixture of water, △ G ° is a positive value was shown, which means that the naturally-occurring dissolved in resveratrol.

In addition, the mass fraction of the trans kyutol ^® HP was observed an amount △ S ° value of from 0.6 trans kyutol ^® HP and the mixture of water or less. A positive Δ S° value means that the dissolution of resveratrol is due to entropy. These results show that the solubilization of resveratrol in a mixed solvent is favorable entropy.

In contrast, when the mass fraction of the trans kyutol ^® HP is larger than 0.7, the △ S ° negative value was observed. These results can be described as being in a trans kyutol ^® HP, resveratrol is rapidly dissolved without the need for substantial heat absorption. In other words, since the solubility of resveratrol in Transcutol ^® HP is high, the Δ S° value decreases as the fraction of Transcutol ^® HP increases.

Referring to Table 4, in all cases, the %ζ _TS value was less than 41.4%, and the %ζ _H value was greater than 58.6%. This means that enthalpy is a major contributor to the Δ G° value, and is consistent with the above results that the dissolution of resveratrol in the selected solvent is caused by enthalpy.

An entropy compensation analyzed - enthalpy to determine the behavior for plum of resveratrol in the trans kyutol ^® HP and water mixture that contains a pure solvent (solvation behavior) and a co-solvent activation (co-solvent action). The results of this analysis are shown in FIGS. 4 and 5.

4 and 5, in all mixed solutions, resveratrol is between △ H° and △ G° or △ S° with an R ² value of 0.9961 (both both) and a positive slope (> 1.0). Showed a linear relationship. Therefore, it can be suggested that the mechanism causing the solvation of resveratrol is due to enthalpy. This is believed to be due to the very high degree of solubility in the Transcutol ^® HP molecule when compared to the thermodynamic solvation behavior of resveratrol in water molecules.

3) Resveratrol Inhibition of stabilizers against precipitation Check effect

The effect of inhibiting precipitation of polyvinylpyrrolidone vinyl acetate (PVP VA64), polyvinylpyrrolidone (PVP), hydroxypropyl methyl cellulose (HPMC), and hydroxypropyl cellulose (HPC) is shown in FIGS. 6 and 7. . In the absence of a stabilizer, the precipitation of resveratrol took place at a very rapid rate. PVP and PVP VA64 showed superior precipitation inhibitory activity compared to HPMC and HPC. For PVP K12, PVP K17 and PVP VA64, concentrations above 330 μg/mL were maintained for 120 minutes. On the other hand, when HPMC and HPC were used, particle agglomeration occurred gradually with time. Therefore, it was confirmed that PVP and PVP VA64 were more effective than HPMC and HPC in controlling particle growth when preparing nanosuspensions.

< Experimental Example 2> Resveratrol Nano Suspension Optimization Study

1. Formulation Parametric Box- for optimization Behnken design( BBD )

In order to determine the optimal composition of the resveratrol nanosuspension, according to the matrix produced by the BBD, 17 resveratrol nanosuspensions of different concentrations of PVP VA64 (X ₁ ), PVP K12 (X ₂ ), and SLS (X ₃ ) It was prepared using. The results obtained for the particle size and zeta potential are shown in Table 5 below.

(In Table 5, X ₁ : concentration of PVP VA64 (mg/mL), X ₂ : concentration of PVP K12 (mg/mL), X ₃ : concentration of SLS (mg/mL))

Referring to Table 5, in all of the prepared nanosuspensions, the particle size (z-average, initial) was less than 200 nm, and the particles grew over time. One, all zeta potential values were -30 mV or less, which was a value that satisfies the target value of the response value.

On the other hand, regression analysis is used to identify the functional relationship between the formulation composition variable and the response value, so that not only a specific pattern, but also most suitable regression functions can be found. The constants in the regression analysis were estimated based on the least squares method. The main effects, interactions, and curvature effects of each factor were confirmed through analysis of variance (ANOVA).

In ANOVA, data were analyzed based on F-values to determine whether elements were significantly related to response values. Only response values with a p-value of 0.05 or less were considered statistically significant.

(In Table 6, Y ₁ : particle size (z-average, initial), Y ₂ : particle size (z-average, day 1), Y ₃ : particle size (z-average, day 3), Y ₄ : Particle size (z-average, day 7), Y ₅ : Particle size (d90, initial), Y ₆ : Particle size (d90, day 1), Y ₇ : Particle size (d90, day 3), Y ₈ : Particle Size (d90, 7th day), Y ₉ : Zeta potential)

Table 6 shows the regression analysis and ANOVA results for the response values. The p-values of all models were less than 0.05, indicating significant significance. The p-value of the lack of fit test exceeded 0.05, meaning that there was no significance. In the Y ₁ -Y ₉ model, the regression constant (R ² ) ranged from 0.9959 to 0.8629. In general, a regression constant of 0.8 or higher is considered to be a good model.

Positive coefficients mean beneficial effects in the optimization analysis, whereas negative values mean there is an inverse correlation between the factor and the response. As shown in the <Table 6>, the concentration of PVP VA64 (X ₁ ) and the concentration of SLS (X ₃ ) showed a negative effect on the particle size (Y ₁ -Y ₈ ), whereas PVP K12 (X The concentration of ₂ ) showed a positive effect. These results mean that the particle size increases as the concentration of PVP K12 (X ₂ ) increases or the concentration of PVP VA64 (X ₁ ) and SLS (X ₃ ) decrease. Specifically, the concentration effect of PVP K12 (X ₂ ) on the particle size (z-average, Y ₁ -Y ₈ ) increased with time. In the early stages of production, the concentration of PVP VA64 (X ₁ ) and the concentration of SLS (X ₃ ) had a greater effect than that of PVP K12 (X ₂ ). However, in the step of maintaining the particle size, all formulation composition parameters showed a significant effect on the particle size.

In the zeta potential (Y ₉ ), the PVP VA64 concentration (X ₁ ) and the PVP K12 concentration (X ₂ ) showed a positive effect, and the SLS concentration (X ₃ ) showed a negative effect. PVP VA64 and PVP K12 are nonionic polymers and SLS are anionic surfactants. As the concentration of SLS increases or the concentration of polymer decreases, the charge on the particle surface becomes negative. When the surface is negatively charged, an ionic barrier is formed between the particles to inhibit the growth of the particles.

Meanwhile, the manufacturing process of the nanosuspension is thermodynamically unstable. Therefore, a stabilizer such as a polymer or a surfactant is essential to maintain a physically stable state. It is known that PVP, which is a nonionic polymer, is immobilized on the surface of drug particles to occupy an adsorption site and prevents drug molecules from being bonded to the crystal lattice in a solution, thereby providing a mechanical barrier to crystallization. However, if the polymer concentration is not sufficient, crystals grow rapidly and aggregate. As the concentration of the polymer increases continuously, the size of the particles increases together due to the thick layer on the surface of the particles, and diffusion between the solvent and the anti-solvent is inhibited during the aggregation process. In addition, as the concentration of the polymer increases, the osmotic pressure increases, resulting in an increase in the attractive force between the colloidal particles. This leads to particle growth.

Surfactants are adsorbed at the solid-liquid interface, reducing the surface tension of the interface and increasing the nucleation rate, which results in an initial reduction in particle size. Moreover, adsorption of the surfactant reduces hydrophobic interactions and coagulation, making the resveratrol particles less hydrophobic and reducing particle growth. Particularly, when SLS, which is an ionic surfactant, is adsorbed on the particle surface, the particle surface has a negative charge. This increases the energy barrier by increasing the repulsion between the particles, thereby preventing particle growth and agglomeration. As a result, the addition of an appropriate concentration of a stabilizing agent can lower the excessively high surface energy of the nanoparticles to be produced.

Next, the relationship between the formulation composition variable and the response value variables was determined through a response curve plot. To make a response curve plot, only two formulation composition variables were indicated at a time; Accordingly, one formulation composition parameter must be fixed. Referring to FIG. 8, it can be seen that the particle size decreases as the concentration of PVP VA64 and SLS increases. After 7 days, the PVP K12 concentration decreased or the particle size decreased as the SLS concentration increased. In addition, a similar trend was observed in the response value variable Y ₅ -Y ₈ (results not shown), that is, as the concentrations of PVP VA64 and PVP K12 decreased, the zeta potential also decreased. These results are consistent with the negative contribution of SLS and the positive contribution of PVP VA64 and PVP K12 shown in Tables 6 and 8 above.

2. Establishment of design space

ICH Guideline Q8 defines the design space as "a multidimensional combination and interaction of input variables (eg material properties) and process variables that have been proven to provide quality correction." Work in this design space does not take into account changes and a flexible and robust process can be established.

In the present invention, the target values were set to 250 nm or less, 1 μm (Y ₅ -Y ₈ ) or less, and -20 mV (Y ₉ ), and a 95% confidence interval was applied. An overlay plot consisting of all response values is shown in FIG. 9. The yellow part of the plot indicates the area that meets the target value. Established yellow areas met the target values in all areas when the concentrations of SLS were 1.0 mg/mL and 1.5 mg/mL. However, when the concentration of SLS was 0.5 mg/mL, the yellow area decreased.

10 shows a probability map drawn using MODDE software based on a calculated particle size and zeta potential model and Monte Carlo simulation for risk analysis. Monte Carlo simulation was performed to propagate the error of the function related from the independent variable to the dependent variable, which represents the distribution of dependent variables (ie, resolution) for each operating condition of the RSM.

The design space established by Monte Carlo simulation was expected to be significantly smaller than that of the overlay plot. However, the region obtained from the Monte Carlo simulation was a robust region that satisfies the set target value with few errors. In this experiment, the design space is represented by the probability of failure for each combination of parameters. A probability of failure of 1% was chosen, which identifies the design space as a region that satisfies a defined factor of 99% or more probability in the predicted CQA. For each combination, 10,000 simulations were performed, and a 95% confidence interval was applied to explain the variability of the Monte Carlo simulation. As a result, the design space of the green area shown in FIG. 10 was a robust area where reliable experimental values are expected. The established green area satisfied the target response values in all areas when the SLS concentration was 1.0 mg/mL. In other words, this area was guaranteed to be robust. Therefore, the highest point was identified in the area where the concentration of SLS was 1.0 mg/mL. Based on the expected function, the optimal values of the parameters were confirmed through schematic and numerical analysis using Design Expert ^® 11.0, and a formulation capable of manufacturing a robust product with target high quality characteristics was determined. That is, a mixed formulation (PVP VA64, PVP K12 and SLS) that maximizes the expected function was identified (10%/5%/1%, w/v). This combination is expected to exhibit the smallest particle size and zeta potential.

3. Robustness of process variables in an optimized configuration

In the formulation of the optimized resveratrol nanosuspension, in order to establish the robustness of the process parameters, nine experiments were performed using FD, and the results are shown in Table 7 below.

Referring to <Table 7>, the measured particle size distribution and zeta potential satisfied target values. The probability map obtained by Monte Carlo simulation showed a defect probability of 1% or less in all areas, as shown in FIG. 11. Therefore, it could be confirmed that the robustness of the process parameters for the optimized formulation was established.

4. Validation of the model

As shown in Table 8 below, from the regression model generated in the present invention for the optimization formulation, a prediction interval of 95% for the response value was calculated. The BBD, which was used for optimizing the formulation composition variable, was used to calculate the system suitability value of the selected response value.

(In Table 8, Y ₁ : particle size (z-average, initial), Y ₂ : particle size (z-average, day 1), Y ₃ : particle size (z-average, day 3), Y ₄ : Particle size (z-average, day 7), Y ₅ : Particle size (d90, initial), Y ₆ : Particle size (d90, day 1), Y ₇ : Particle size (d90, day 3), Y ₈ : Particle Size (d90, 7th day), Y ₉ : Zeta potential)

Referring to Table 8 above, the observed response values were within the 95% prediction interval. Therefore, the validity of the model generated according to the present invention was confirmed. In addition, FD was used to establish the robustness of the process variable, and as a result, as shown in FIG. 12, it stabilized over time. In all experiments shown in Table 7 above, the trend of particle size was similar.

< Experimental Example 3> Optimized Resveratrol Confirmation of the properties of the nano suspension

1. Check the elution characteristics

The in vitro drug dissolution ability of the above-described unloaded resveratrol nanosuspension formulation was compared with that of the raw material. Fig. 13 shows the dissolution profile of the resveratrol nano suspension and the resveratrol raw material.

13, the resveratrol nanosuspension showed higher dissolution rate and drug release rate than that of the resveratrol raw material. The drug release rates after 5 minutes of the resveratrol nanosuspension and raw materials were 90.2% and 3.5%, respectively, and the drug release rates after 360 minutes were 90.5% and 21.2%, respectively. From these results, it was found that the resveratrol nanosuspension according to the present invention exhibited a much higher drug release rate compared to the resveratrol powder.

2. In rats drug Dynamics Experiment

To confirm whether the nanosuspension drug delivery system can improve oral bioavailability, in vivo experiments were performed using Sprague-Dawley rats, through which the pharmacokinetic parameters of the resveratrol nanosuspension and the raw material were compared. 14 shows the plasma concentration-time profile of resveratrol after oral administration of the nanosuspension and raw materials. In addition, pharmacokinetic parameters (AUC _{0→12 h} , C _max , and T _max ) are shown in Table 9 below.

(a: It means that p<0.05 for the raw material of resveratrol, and the data are expressed as mean±standard deviation [n=4].)

Referring to FIG. 14, the plasma concentration and rapid drug absorption rate of the resveratrol nanosuspension were dramatically higher than that of the resveratrol raw material. AUC _{0 → 12 h} , C _max , and T _max values were 387.0 ± 26.0 ng·h/mL, 301.4 ± 79.6 ng/mL, and 0.44 ± 0.13 h, respectively. The oral absorption of the resveratrol nanosuspension was also higher than that of the resveratrol raw material, and as shown in Table 9, the AUC _{0 → 12 h} and C _max values showed an increase of 1.6 times and 5.7 times, respectively. For orally administered drugs, solubility is a critical rate decision step for absorption. When the nanosuspension enters the gastrointestinal tract, the nanoparticles provide a larger surface area for dissolution and the molecular diffusion of resveratrol obtained from the dissolved nanoparticles, and according to classical passive diffusion theory, this is an increased concentration at the surface of the nanocrystal. Leads to a gradient. These results indicate that the oral absorption of resveratrol was significantly increased by the nano suspension form.

3. Long-term stability check

The stability of the resveratrol nanosuspension was analyzed over a period of 6 months through confirmation of particle size, zeta potential, and drug content. As a result, as shown in FIG. 15, particles formed by the anti-solvent precipitation method were mainly in the form of a spherical shape with a smooth surface and a diameter slightly smaller than 200 nm in the initial stage of formation. After 6 months, the particle size was about 200 nm and was also spherical in shape.

(N.D: There is no data because the data cannot be measured)

In addition, as shown in Table 10 and FIG. 16, the particle size was smaller than 250 nm, and the drug content was higher than 95%. Therefore, the nanosuspension was maintained in a stable state without causing aggregation of drugs or decomposition of resveratrol for 6 months.

< Experimental Example 4> Preparation and comparison of nano suspensions of various poorly soluble drugs

Based on the experimental results obtained in the above Examples and Experimental Examples, a nano suspension with different conditions was prepared, such as using other poorly soluble drugs other than resveratrol, or using PEG400, which is known to be similar to Transcutol, or ethanol as a solvent. I tried to compare.

Thus, each was prepared according to the components shown in the following <Table 11>. That is, a drug solution was prepared by taking a certain amount of drug and putting it in a 10 mL volumetric flask, and then accurately aligning the mark using a solvent. Thereafter, in order to prepare a stabilizer (polymer and surfactant) solution, a polymer and a surfactant were taken according to a certain ratio and placed in a 100 mL volumetric flask, and the mark was adjusted using water as a solvent. Thereafter, in order to prepare nanoparticles, a certain amount of a stabilizer solution was taken and a drug solution was slowly injected while stirring at 700 rpm using a magnetic stirrer. Thereafter, after the solution injection was completed, the mixture was further stirred for 1 minute, and then the particle size of each nanoparticle was measured using a dynamic light scattering method.

As a result, as shown in <Table 11>, when PEG400 was used as a solvent for resveratrol, resveratrol aggregated to form a large lump, so that a nanosuspension containing nanoparticles could not be prepared, and in the case of ethanol, 727.9 Although nanoparticles having a size of nm were formed and the size thereof became too large, when using Transcutol HP, the solvent selected in the present invention, a nanosuspension containing nanoparticles having an appropriate particle size was prepared.

In addition, when aprepitant was used as a drug and Transcutol P was used as a solvent, when the composition of the stabilizer solution was a combination of PVP VA64/PVPK12, a nanosuspension was prepared with an average particle size of 291.4 nm.

In the case of using cyclosporine and transcutol P, it was confirmed that a preferred suspension was prepared at 193.5 nm when the combination of PVP VA64/PVP K12/SLS was used.

In the case of using celecoxib and Transcutol P, when the composition of the stabilizer solution is a combination of PVP VA64/PVPK12, the average particle size is 234.1 nm, when the combination of PVP VA64/PVP K12, the average particle size is 235.2 nm, When the combination of PVP VA64/PVP K12/SLS was 274.3 nm, the overall nanosuspension was well prepared.

< Experimental Example 5> MHY498((Z)-5- (2,4- Dihydroxybenzylidene ) Thiazolidine -2,4- D On) nano suspension preparation and comparison

A nanosuspension using transcutol for MHY498, which is known as a poorly soluble compound, was prepared under the conditions as shown in Table 12 below. Piperidine was added to a solution of 2,4-dihydroxybenzaldehyde and thiazolidine-2,4-dione substituted in ethanol. . After the reaction was subjected to several steps, MHY498 was obtained as a solid yellow powder. A detailed synthesis method of MHY498 is described in Korean Patent No. 10-2013-0045887 (published on May 6, 2013).

That is, a drug solution was prepared by taking a drug of a certain concentration and putting it in a 10 mL volumetric flask, and then accurately aligning the mark using Transcutol HP as a solvent. Thereafter, in order to prepare a stabilizer solution, PVP K30 was taken according to a certain ratio and put into a 100 mL volumetric flask, and the mark was adjusted using water as a solvent. Thereafter, in order to prepare nanoparticles, a certain amount of a stabilizer solution was taken and a drug solution was slowly injected while stirring at 700 rpm using a magnetic stirrer. Thereafter, after the solution injection was completed, the mixture was further stirred for 1 minute, and then the particle size of each nanoparticle was measured using a dynamic light scattering method.

As a result, as shown in Table 12, in the configuration using Transcutol HP as a solvent and PVP K30 as a stabilizer, the average particle size is 35.3 nm to 142.9 nm regardless of the concentration of the MHY498 solution or the ratio of the solvent. It was confirmed that a nano suspension was prepared.

Since the specific parts of the present invention have been described in detail above, for those skilled in the art, it is obvious that this specific technique is only a preferred embodiment, and the scope of the present invention is not limited thereby. Do. That is, the substantial scope of the present invention is defined by the appended claims and their equivalents.

Claims (8)

Dissolving a poorly soluble drug in a solvent, Transcutol;
Dissolving polyvinylpyrrolidone, a water-soluble polymer, in water to prepare an aqueous polyvinylpyrrolidone polymer solution; And
A method of preparing a nanosuspension comprising injecting a solution in which the poorly soluble drug is dissolved in the polyvinylpyrrolidone polymer aqueous solution. The method of claim 1, wherein the poorly soluble drug is resveratrol, MHY498((Z)-5-(2,4-dihydroxybenzylidene)thiazolidine-2,4-dione), aprepitant, cyclosporine, sele Coxib, coenzyme Q10, valsartan, dabigatran etexilate, sorafenib and silymarin method for producing a nano-suspension, characterized in that selected from the group consisting of. delete The method of claim 1, wherein the polyvinylpyrrolidone is polyvinylpyrrolidone K12, polyvinylpyrrolidone K17, polyvinylpyrrolidone K25, polyvinylpyrrolidone K30, and polyvinylpyrrolidone vinyl acetate VA64. Method for producing a nano-suspension, characterized in that selected from the group consisting of. Dissolving a poorly soluble drug in a solvent, Transcutol;
Dissolving a water-soluble polymer polyvinylpyrrolidone and a surfactant in water to prepare a polyvinylpyrrolidone polymer and an aqueous surfactant solution; And
A method for preparing a nanosuspension comprising injecting a solution in which the poorly soluble drug is dissolved in the polyvinylpyrrolidone polymer and the aqueous surfactant solution. The method of claim 5, wherein the surfactant is sodium lauryl sulfate (SLS), sucrose fatty acid ester, polyoxyethylene alkyl ether, polyoxyethylene glycolated natural or hydrogenated castor oil, polyoxyethylene sorbitan fatty acid ester [Tweens )], polyethylene glycol, polyoxyethylene stearate, polyoxyethylene-polyoxypropylene copolymer (poloxamer or poloxamine), polyethylene glycolated phospholipid and alpha-tocopherol polyethylene glycol succinate selected from the group consisting of Method for producing a nano-suspension, characterized in that. The method of any one of claims 1, 2, 4 to 6, wherein the nanosuspension comprises poorly soluble drug nanoparticles having an average diameter of 1 nm to 300 nm. Manufacturing method. A nanosuspension prepared by the manufacturing method according to any one of claims 1, 2, and 4 to 6.

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