KR102709458B1 - Sustained-release clonidine microsphere injection and manufacturing method thereof - Google Patents

Abstract

The present disclosure relates to a sustained-release injection containing biodegradable polymer microparticles containing clonidine as an active ingredient and a method for producing the same. The present invention can improve the clonidine content and encapsulation efficiency of the microparticles by including a specific acid in the microparticles. In addition, the formulation of the present invention not only suppresses initial drug over-release but also exhibits a zero-order release pattern, thereby continuously inducing drug release for a certain period of time.

Description

Sustained-release clonidine microsphere injection and manufacturing method thereof

The present disclosure relates to a sustained-release polymer microsphere formulation containing clonidine as an active ingredient and a method for producing the same. More specifically, the present disclosure relates to a sustained-release formulation in which clonidine is encapsulated in a biodegradable polymer microsphere as a carrier, which suppresses initial excessive release and exhibits a zero-order release pattern for a certain period of time, and a method for producing the same.

Tic disorders are caused by abnormalities in the brain's neurotransmitter system or genetic factors, and attention deficit hyperactivity disorder (ADHD) is caused by an imbalance of neurotransmitters such as dopamine and norepinephrine in the brain. Tic disorders are treated with the a-2 adrenergic receptor, antipsychotic drugs, and behavioral therapy, and the a-2 adrenergic receptor shows great efficacy in patients with tic disorders with concurrent ADHD. Clonidine is a representative a-2 adrenergic receptor that suppresses norepinephrine secretion. As a result, when the secretion of norepinephrine in the brain decreases, excessive nerve stimulation decreases, and symptoms are alleviated.

A dosage form designed to release a drug consistently and continuously in the body after subcutaneous or intramuscular injection is generally called an extended-release injection dosage form. It can reduce the burden of frequent drug administration, improve the convenience of taking medication, and maximize the therapeutic effect. Common manufacturing methods for extended-release injections include coacervation, melt injection, spray drying, and solvent evaporation. Among these methods, the solvent evaporation method, which is classified into the double emulsion solvent evaporation method (W/O/W emulsion solvent evaporation method) and the single emulsion solvent evaporation method (O/W emulsion solvent evaporation method), is the most widely used. However, microspheres manufactured by this method have a high initial release rate, and during the manufacturing process of microspheres, the drug dissolving in water or an organic solvent causes problems such as changes in the physical properties of the drug, loss of efficacy, and low encapsulation efficiency.

Literature Gaignaux, Am lie, et al. “Development and evaluation of sustained-release clonidine-loaded PLGA microparticles” International journal of pharmaceuticals 437.1-2 (2012): 20-28 and Gaignaux, Am lie, et al. "Evaluation of the degradation of clonidine-loaded PLGA microspheres." Journal of microencapsulation 30.7 (2013): 681-691 reported that sustained-release polymeric microspheres containing clonidine were prepared by a double emulsification solvent evaporation method, using a phosphate buffer solution of pH 7.0 as the inner phase and a polyvinyl alcohol solution buffered with a phosphate buffer solution of pH 8.0 as the outer phase. In this method, the use of a buffer solution was suggested, assuming that clonidine protons are released at a pH similar to the pKa of clonidine and then bond with the polymer. However, this method has the disadvantage of being difficult to produce due to the complicated manufacturing process. In addition, although the encapsulation efficiency was improved by using a buffer solution (approximately 34%), the low encapsulation efficiency is still considered to be a limitation.

Gaignaux, Amlie, et al. “Development and evaluation of sustained-release clonidine-loaded PLGA microparticles” International journal of pharmaceuticals 437.1-2 (2012): 20-28 Gaignaux, Amlie, et al. “Evaluation of the degradation of clonidine-loaded PLGA microspheres.” Journal of microencapsulation 30.7 (2013): 681-691

Therefore, the problem that the present disclosure seeks to solve is to provide a sustained-release polymer microsphere formulation containing clonidine as an active ingredient, which has good encapsulation efficiency of clonidine, can suppress initial burst release of microspheres under high loading conditions, and further exhibits good zero-order release characteristics, and a method for producing the same.

In order to solve the above problem, one aspect of the present invention provides a sustained-release injection containing biodegradable polymer microparticles containing clonidine, wherein the microparticles contain at least one selected from the group consisting of capric acid, mandelic acid, and hydroxy naphthoic acid (1-hydroxy-2-naphthoic acid), thereby slowly releasing clonidine from the microparticles.

Preferably, the sustained-release injection containing biodegradable polymer microparticles containing clonidine according to the present invention comprises hydroxy naphthoic acid (1-hydroxy-2-naphthoic acid) within the microparticles.

In general, when clonidine is encapsulated in biodegradable polymers such as PLGA to manufacture microspheres, not only is there a problem of an initial burst release phenomenon in which the release of the drug increases rapidly at the beginning, but there are also problems such as low encapsulation efficiency and inability to maintain a constant concentration of drug release for a desired period of time. Biodegradable polymer microspheres release the drug gradually as the polymer matrix decomposes through hydrolysis, but the initial burst release phenomenon can rapidly increase the concentration of the drug in the blood, causing toxicity. In addition, due to the slow release rate after the initial burst release, the drug falls outside the effective blood concentration range and cannot exhibit a constant efficacy.

The present inventors have discovered that by including an acid (more preferably hydroxy naphthoic acid) that is capric acid, mandelic acid, hydroxy naphthoic acid or a mixture thereof in the clonidine-containing biodegradable polymer microparticles, the acid and clonidine form a physical bond within the microparticles during and after the manufacturing process, thereby effectively solving the above-mentioned problems, thereby completing the present invention. That is, the microparticles according to the present invention not only have a high clonidine encapsulation efficiency, but also suppress initial burst release and enable sustained zero-order release. It is presumed that such an effect can be achieved through the formation of a hydrophobic bond between a specific acid and clonidine during the solidification process in the manufacturing of the microparticles, but the present invention is not limited to this mechanism.

Among the hydrophobic acids additionally included in the microspheres according to the present invention, the most preferred material is hydroxy naphthoic acid. When hydroxy naphthoic acid is used, not only can microspheres with improved encapsulation efficiency be manufactured, but also microspheres suitable for a zero-order release model showing a constant release pattern can be manufactured.

After intravenous administration of microspheres containing clonidine and hydroxynaphthoic acid, the biodegradable polymer (preferably PLGA) absorbs electrolytes in the body over time, becomes hydrated and swells, gradually releasing the active ingredient clonidine inside.

Biodegradable polymers that can be used in the microspheres according to the present invention include polylactide (PLA), polyglycolide (PGA), poly(lactide-co-glycolide) (PLGA), poly(lactide-co-glycolide) glucose, and the like.

Another aspect of the present invention also provides a method for producing a sustained-release injection containing biodegradable polymer microspheres comprising clonidine, the method comprising the steps of (S1) preparing a solution comprising a biodegradable polymer; clonidine; and an acid selected from capric acid, mandelic acid, hydroxy naphthoic acid or a mixture thereof; and (S2) adding (for example, dropping) the solution to an aqueous solution (preferably, a polyvinyl alcohol aqueous solution) while stirring to form an O/W emulsion and thereby form microspheres. In the producing method of the present invention, the microspheres may be formed when a cosolvent in the O/W emulsion diffuses and/or evaporates into the aqueous medium. That is, the present invention also provides a method for producing a biodegradable polymer microsphere-type sustained-release preparation using an O/W emulsion solvent evaporation method. This method of the present invention enables the production of a sustained-release preparation that maximizes the encapsulation amount and encapsulation efficiency of clonidine in the microspheres and induces continuous and uniform drug release.

In one aspect of the present invention, the manufacturing method according to the present invention further includes a step (S3) of solidifying microspheres through evaporation of the solvent in the O/W emulsion formed in step (S2).

In another aspect of the present invention, the manufacturing method according to the present invention may further include a step (S4) of drying, preferably freeze-drying, the microparticles produced in step (S3).

More specifically, the manufacturing method of the present invention can be manufactured through the following steps.

(Step S1) Step of dissolving clonidine, biodegradable polymer, and specific acid by vigorous mixing and stirring.

The non-aqueous solvent used to prepare the solution in this step is not particularly limited as long as it can dissolve the biodegradable polymer. According to a specific embodiment of the present invention, the non-aqueous solvent may be a non-aqueous solvent having a low boiling point, for example, a boiling point of 25°C to 85°C. When the boiling point of the non-aqueous solvent is within the above range, it is advantageous in terms of evaporation and drying of the microspheres after their formation. In the present invention, non-limiting examples of the non-aqueous solvent include, but are not particularly limited to, dichloromethane, dimethyl sulfoxide, dimethyl formamide, acetonitrile, ethyl acetate, chloroform, and the like.

The above biodegradable polymer is a typical polymer used in the production of microspheres, can be used as a drug delivery vehicle, has biocompatibility, and is self-degradable in the body. In the present invention, one or more biodegradable synthetic polymers such as polyglycolide (PGA), polylactide (PLA), poly(lactide-co-glycolide) (PLGA), and poly(lactide-co-glycolide) glucose can be used.

In a specific embodiment of the present invention, the biodegradable polymer may be one having a weight average molecular weight of 60,000 or less. For example, poly(lactide-co-glycolide)(50:50) having a molecular weight of about 13,000, poly(lactide-co-glycolide)(50:50) having a molecular weight of about 33,000, poly(lactide-co-glycolide)(50:50) having a molecular weight of about 52,000, poly(lactide-co-glycolide)(75:25) having a molecular weight of about 20,000, poly(lactide)(100:0) having a molecular weight of about 16,000, etc. may be used. Examples of such biodegradable polymers include Resomer RG502H, RG503H, RG504H, RG752H, and R202H from Boehringer Ingelheim.

In a specific embodiment of the present invention, the biodegradable polymer may have an intrinsic viscosity of 0.16 to 0.6 dL/g. In the present invention, if the viscosity of the biodegradable polymer is less than 0.1 dL/g, microspheres encapsulating drug powder particles may not be formed properly or the drug encapsulation rate may significantly decrease, and if it exceeds 0.6 dL/g, the size of the microspheres may be formed excessively large, and the drug release amount may be small, so that the desired level of efficacy may not be exhibited.

According to a specific embodiment of the present invention, in the step, the biodegradable polymer is dissolved in a concentration of 10 mg to 150 mg per ml of the first mixture, preferably 25 mg to 125 mg, and more preferably 50 mg to 125 mg. When the concentration of the biodegradable polymer is low, the viscosity of the polymer solution is lowered, so that the particle size tends to be smaller when dispersed by a homogenizer. However, since the proportion of the polymer in the microdroplets before the formation of microparticles is small, the particle structure may be loose when formed, and accordingly, the drug loading amount and loading efficiency also seem to be lowered. On the other hand, when the concentration exceeds 150 mg/ml, the viscosity of the polymer solution increases, so that the size of the microparticles tends to be larger.

Capric acid, mandelic acid, hydroxy naphthoic acid or a mixture thereof are used as the acid, with hydroxy naphthoic acid being particularly preferred.

(Step S2) A step of slowly mixing and stirring the above solution while adding it to the receiving medium to produce an O/W emulsion.

The aqueous medium of the present invention may contain an emulsifier. The content of the emulsifier is preferably in the range of 0.1 g to 10 g, or 3 g to 7 g per 1 L of the aqueous solvent, that is, 0.1 (w/v)% to 10 (w/v)%, or 3 (w/v)% to 7 (w/v)%. Examples of the emulsifier include, but are not limited to, polysorbate, polyethylene glycol, polyvinyl alcohol, poloxamer, and Span80. Preferably, the emulsifier in the present invention is polyvinyl alcohol.

In a specific embodiment of the present invention, the emulsion in this step is prepared by mixing the solution of step S1 and the aqueous medium in a ratio of 1:5 to 1:50 based on the volume ratio. The mixing ratio is preferably 1:10 to 1:20. When the volume ratio of the solution of step S1 and the aqueous medium is less than 1:5, the gap between the fine droplets in the O/W emulsion becomes narrow, which increases the possibility of collision, and as a result, the fine droplets may clump together before the diffusion and evaporation of the cosolvent and non-aqueous solvent, making it difficult to form microspheres of uniform size. In addition, when the ratio exceeds 1:50, the amount of emulsifier added increases, which is disadvantageous in terms of cost.

(Step S3) A curing step in which microspheres are created as the co-solvent in the O/W emulsion diffuses and/or evaporates into the aqueous medium.

In the above step S2, primary polymer curing is induced as the cosolvent diffuses and/or evaporates into the aqueous medium, thereby encapsulating drug particles within the microspheres. That is, as the cosolvent within the dispersed microdroplets in the O/W emulsion rapidly diffuses and/or is extracted into the aqueous solvent phase, the biodegradable polymer is rapidly solidified to form microspheres. In one specific embodiment of the present invention, it is preferable to vigorously stir the microdroplets during this process using a magnetic stirrer so that the microspheres do not clump together. The formation time of the microspheres may be 1 to 10 minutes. When the formation time of the microspheres is less than 1 minute, the biodegradable polymer of the microspheres may be less hardened, and a clumping phenomenon may occur between the particles. On the other hand, when it exceeds 10 minutes, the encapsulation efficiency of the drug may be reduced.

After the formation of microspheres, the aqueous medium and solvent are removed from the O/W emulsion to obtain microspheres.

(Step S4) Step of (freeze) drying the generated microspheres

Through the (freeze) drying process, the structure of the microspheres becomes solid and the drug is completely encapsulated in the biodegradable polymer. According to a specific embodiment of the present invention, the drying may preferably be a freeze-drying step in order to completely remove the solvent remaining in the microspheres. The freezing step may be performed under temperature conditions of -45°C to -55°C, and the drying time may be 12 to 24 hours. According to a specific embodiment of the present invention, the freeze-drying step may be performed under vacuum conditions. If the drying time is performed under temperature conditions of less than 12 hours, the residual solvent may not be completely removed, whereas if the drying time exceeds 24 hours, the productivity may be reduced.

In the manufacturing method of the present invention, the encapsulation efficiency of clonidine in the microparticles (active ingredient encapsulated amount/added active ingredient weight × 100(%)) is 55 to 70 wt%, and preferably 60 to 70 wt%.

In the manufacturing method of the present invention, the microspheres may contain 30 to 95 wt%, preferably 45 to 75 wt%, of the biodegradable polymer relative to 100 wt% of the microspheres. If the biodegradable polymer is contained in an amount of less than 40 wt%, the microspheres tend to exhibit porous characteristics on the surface and inside, which may cause problems such as excessive initial drug release or a short release time. On the other hand, if the amount exceeds 95 wt%, the amount of microspheres administered to a patient or animal may become excessively large, making administration difficult or impossible.

The present invention provides a clonidine-containing microparticle sustained-release preparation (preferably an injection) which can exhibit a high encapsulation amount of clonidine and a zero-order drug release pattern that is sustained for a long period of time, and a method for producing the same. The manufacturing method of the present invention can produce microparticles using a minimal process and energy by producing biodegradable polymer particles encapsulating clonidine using an O/W emulsification solvent evaporation method. In particular, by including a specific acid capable of sustained-releasing clonidine in the microparticles, not only the zero-order release pattern but also the internal structure of the microparticles can be made uniform, thereby efficiently controlling drug release.

The following drawings attached to this specification illustrate preferred embodiments of the present invention and, together with the contents of the invention described above, serve to further understand the technical idea of the present invention; therefore, the present invention should not be construed as being limited to matters described in such drawings.
Figure 1 is an electron microscope photograph of clonidine microspheres manufactured in Comparative Example 1, Comparative Example 2, and Example 1-2. The photograph on the left is a photograph observing the overall shape and surface of the microspheres, and the photograph on the right is a photograph showing the cross-section of the microspheres. The microspheres manufactured in Example 1-2 showed a smooth surface, and no breakage or deformation of the particles was observed after freeze-drying.
Figure 2 shows the experimental results for Comparative Example 2, which is microspheres composed of a biodegradable polymer and clonidine, and Examples 1-1 and 1-2, which are microspheres using hydroxy naphthoic acid with different contents, and are optical microscope photographs observing the swelling and agglomeration of particles in a PBS solution over time. It can be seen that all the microspheres swell over time, and eventually agglomeration or disintegration occurs, changing the shape of the microspheres. In particular, it was found that as the content of hydroxy naphthoic acid increased, the agglomeration and disintegration of the microspheres were delayed by up to about 42 days. This result means that the release of clonidine from the microspheres can be delayed.
Figure 3 is a graph showing the in vitro drug release behavior of Comparative Example 2, which is a microsphere composed of a biodegradable polymer and clonidine, and Examples 1-1 and 1-2, which are microspheres manufactured with different contents of hydroxy naphthoic acid, according to Experimental Example 3. In Figure 3, the sustained-release characteristics of clonidine in a PBS solution over time can be confirmed.
Figure 4 is a graph showing the in vitro drug release behavior of Example 1-2, which is a microsphere composed of a biodegradable polymer, clonidine, and hydroxynaphthoic acid, and Comparative Examples 3-3 and 3-6, which are microspheres composed of other acids, capric acid or malonic acid, according to Experimental Example 3. The sustained-release characteristics of clonidine in a PBS solution over time can be confirmed.

Hereinafter, in order to help understand the present invention, examples and the like will be described in detail. However, the examples according to the present invention may be modified in various different forms, and the scope of the present invention should not be construed as being limited to the following examples. The examples of the present invention are provided to more completely explain the present invention to a person having average knowledge in the field to which the present invention belongs.

Examples 1-3 and Comparative Example 2 : Preparation of O/W-type microspheres using biodegradable polymer PLGA, clonidine, and hydroxy naphthoic acid

750 mg of biodegradable polymer PLGA (manufacturer: EVONIK, product name: RESOMER®RG753H), 30 mg of clonidine, and 0 mg (Comparative Example 2), 12.3 mg, 24.5 mg, and 36.8 mg of hydroxy naphthoic acid (1-hydroxy-2-naphthoic acid) were added to 1.5 mL of dichloromethane and stirred vigorously to dissolve for 15 minutes. The uniformly dissolved solution was added to 250 mL of 1% (w/v) polyvinyl alcohol (manufacturer: Sigma-Aldrich, 87–90% hydrolyzed MW: 30,000–70,000) aqueous solution and dispersed at 3,000 rpm using a homogenizer (IKA, ULTRA-TURRAX T8) for 1 minute to obtain an O/W emulsion. After that, the above emulsion was refrigerated (4°C) and stirred vigorously for 4 hours under normal pressure with a magnetic stirrer (450 rpm) to form microspheres. After that, the microspheres were collected using a sieve and washed 3 to 5 times with distilled water. The washed microspheres were freeze-dried at -50°C for 24 hours to completely remove the solvent within the microspheres.

A representative example of Example 1-3 is Example 1-2.

Comparative Example 1 Comparative Example 2 Example 1-1 Example 1-2 Example 1-3 clonidine hydrochloride 30 mg - - - - Clonidine - 30 mg 30 mg 30 mg 30 mg Hydroxynaphthoic acid - - 12.3 mg 24.5 mg 36.8mg PLGA(RG753H) 750 mg 750 mg 750 mg 750 mg 750 mg

Comparative Example 1 : Preparation of W/O/W type microspheres using biodegradable polymer PLGA and clonidine hydrochloride

Clonidine hydrochloride (30 mg) was added to 0.35 ml of a phosphate buffer solution (inner phase) having a pH of 7.0, and biodegradable polymer PLGA (1000 mg) (manufacturer: EVONIK, product name: RESOMER®RG753H) was added to 5 ml of dichloromethane (organic phase). These were then stirred vigorously for 15 minutes to dissolve them. The homogeneously dissolved inner phase was added to the organic phase, and a W/O emulsion was obtained using an ultrasonicator (Sonic Dismembrator-Model 500, Fisher Scientific, Inc. NH, USA) at 20% power for 1 minute. After that, the W/O emulsion was added to 200 ml of 1% (w/v) polyvinyl alcohol (manufacturer: Sigma-Aldrich, 87-90% hydrolyzed, MW: 30,000-70,000) aqueous solution buffered with a pH 8.0 phosphate buffer, and dispersed using a homogenizer (IKA, ULTRA-TURRAX T8) at 2,000 rpm for 1 minute to obtain a W/O/W emulsion. After that, the emulsion was refrigerated (4 °C) and stirred vigorously with a magnetic stirrer (450 rpm) for 3 hours under normal pressure to form microspheres. Thereafter, the microspheres were collected using a sieve and washed 3-5 times with distilled water. The washed microspheres were freeze-dried at -50 °C for 24 hours to completely remove the solvent inside the microspheres.

Comparative Example 2 : Manufacturing of O/W type microspheres using biodegradable polymer PLGA and clonidine

750 mg of biodegradable polymer PLGA (manufacturer: EVONIK, product name: RESOMER®RG753H) and 30 mg of clonidine were added to 1.5 ml of dichloromethane and stirred vigorously to dissolve for 15 minutes. Thereafter, microspheres were manufactured in the same manner as in Example 1-1.

Comparative Example 3 : Preparation of microspheres using biodegradable polymer PLGA, clonidine, and pharmaceutically acceptable acid

750 mg of biodegradable polymer PLGA (manufacturer: EVONIK, product name: RESOMER®RG753H), 30 mg of clonidine, and 0.13 mol of each pharmaceutically acceptable acid as a hydrophobic solidifying agent were added to 1.5 mL of dichloromethane and dissolved for 15 minutes with vigorous stirring. Thereafter, microspheres were manufactured in the same manner as in Example 1. The pharmaceutically acceptable acids used were as shown in Table 2 below.

Example 1-2 Comparative Example 3-1 Comparative Example 3-2 Comparative Example 3-3 Comparative Example 3-4 Comparative Example 3-5 Comparative Example 3-6 Clonidine 30 mg 30 mg 30 mg 30 mg 30 mg 30 mg 30 mg Hydrophobic
Solidifying agent

(0.13 mol) Hydroxy
Naphthoic acid

24.5 mg Benzoic acid


15.9 mg Camphorsulfonic acid


30.3 mg Capric acid


22.5 mg Caproic acid


15.1 mg Glycolic acid


9.9 mg Mandelic acid


13.6 mg PLGA
(RG753H) 750 mg 750 mg 750 mg 750 mg 750 mg 750 mg 750 mg

Experimental Example 1 : Measurement of the shape of microspheres

The microsphere morphology obtained in Comparative Examples 1, 2, and Examples 1-2 was confirmed using an electron microscope and an optical microscope. For observing the cross-section of the microsphere with an electron microscope, a sample of 10 mg of the microsphere was cut after treating it with liquid nitrogen. About 10 mg of the manufactured microsphere was coated with platinum for about 2 minutes using a coater (Quorum A 150 TES, 10 mA). Thereafter, the cross-section and surface of the microsphere were observed using a scanning electron microscope (Tescan Mira 3, LMU FEG-SEM), respectively.

The measurement results are shown in Figures 1 and 2. According to the measurement results, microparticles of about 38 to 125 μm in size were confirmed in Examples 1-1, 1-2, Comparative Examples 1 and 2.

Figure 1 is an electron microscope photograph of clonidine-containing microspheres manufactured in Comparative Example 1, Comparative Example 2, and Example 1-2. The microspheres manufactured in Comparative Example 1 had a microporous structure on the surface and in the cross-section. The microspheres manufactured in Comparative Example 2 had a smooth surface and a dense matrix in the cross-section. Example 1-2 was a clonidine microsphere containing hydroxy naphthoic acid, an aromatic acid, and showed a smooth surface. In Example 1-2, a few holes were observed on the surface and inside the microspheres, but there did not seem to be a major problem in using them as injections. In none of the microspheres manufactured in Comparative Example 1, Comparative Example 2, and Example 1-2 were particles observed to be broken or deformed after freeze-drying.

Experimental Example 2 : Measurement of clonidine content and encapsulation efficiency in microspheres

About 10 mg of the microspheres manufactured in the above Examples 1, Comparative Examples 2 and 3 were each placed in 2 ml of dichloromethane (Honeywell) and completely dissolved, which was then used as a test solution. The absorbance value measured using an HPLC meter was converted to calculate the content of clonidine encapsulated in the microspheres. The encapsulation rate was calculated as the encapsulation amount relative to the administered amount of the drug.

About 10 mg of the microspheres manufactured in the above Comparative Example 1 were each added to 2 ml of dichloromethane (Honeywell) and completely dissolved, and then the mobile phase was added and used as a test solution. Thereafter, the same procedure as in Example 1, Comparative Example 2, and Comparative Example 3 was followed.

HPLC analysis conditions were as follows. The mobile phase was prepared with Acetonitrile:Buffer = 8:2 ratio. The UV detector was 242 nm, the separation column was 4.6 mm × 25 cm, packing L10, the injection size was 10 μL, the flow rate was 0.8 mL/min, the analysis time was 8 min, and the analysis range was measured in the concentration range of 1000 μg/ml to 5.0576 μg/ml.

Table 3 below shows the results of measuring the clonidine content and encapsulation efficiency of microparticles.

Clonidine content (%)
(Drug loading amount/microparticle weight) × 100(%) Theoretical clonidine content
(Drug dosage/microparticle weight) × 100(%) Clonidine encapsulation efficiency (%)
(Drug loading amount/Drug administration amount) × 100% Example 1-1 2.28 ± 0.05 3.79 60.16 ± 1.40 Example 1-2 2.49 ± 0.06 3.73 66.66 ± 1.73 Example 1-3 2.08 ± 0.13 3.67 56.61 ± 3.44 Comparative Example 1 1.01 ± 0.10 2.91 34.63 ± 1.82 Comparative Example 2 2.14 ± 0.10 3.85 55.71 ± 2.60

The encapsulation efficiency of clonidine in Examples 1-1, 1-2, and 1-3 was approximately 57-67%. In Example 1-3, since the content of hydroxy naphthoic acid was high, microsphere formation was not good and the encapsulation efficiency was less than 60%. In addition, in Comparative Example 2, which did not contain hydroxy naphthoic acid, a hydrophobic release delaying agent, the drug leaked into the external water phase during the manufacturing process, resulting in a high drug loss rate and relatively low encapsulation efficiency. Comparative Example 1 had the lowest encapsulation efficiency due to drug leakage caused by two solvent diffusion processes.

Meanwhile, Table 4 below shows the results of measuring the content of clonidine and the encapsulation efficiency in the microparticles manufactured in Comparative Example 3.

Clonidine content (%)
(Drug loading amount/microparticle weight) × 100 (%) Theoretical clonidine content
(Drug dosage/microparticle weight) × 100 (%) Clonidine encapsulation efficiency (%)
(Drug loading amount/Drug administration amount) × 100% Example 1-2
Hydroxy naphthoic acid 2.49 ± 0.06 3.73 66.66 ± 1.73 Comparative Example 3-1
Benzoic acid 1.34 ± 0.02 3.77 36.03 ± 0.43 Comparative Example 3-2 Camphorsulfonic acid 1.37 ± 0.04 3.70 36.46 ± 1.11 Comparative Example 3-3 Capric Acid 2.22 ± 0.05 3.74 60.09 ± 1.44 Comparative Example 3-4 Caproic Acid 1.49 ± 0.05 3.77 39.91 ± 1.44 Comparative Example 3-5 Glycolic Acid 1.84 ± 0.02 3.80 48.30 ± 0.62 Comparative Example 3-6 Mandelic Acid 2.54 ± 0.03 3.78 67.18 ± 0.70

Most of the added acids form salts with clonidine during the manufacturing process of the microspheres, which is expected to cause clonidine to escape into the external phase and be lost when the O/W type microspheres are solidified during the manufacturing process. That is, it is presumed that when the added acids form salts with clonidine, they are ionized upon contact with the water phase and quickly move to the external phase. For this reason, it is thought that the clonidine content and encapsulation rate in the microspheres decrease. The microspheres containing capric acid of Comparative Example 3-3 and mandelic acid of Comparative Example 3-6 had a structure with 8 or more carbon atoms, which increased hydrophobic bonds with clonidine and effectively prevented clonidine from leaking into the external phase during the solidification process, thereby showing high clonidine content and encapsulation efficiency in the microspheres.

Likewise, the hydroxy naphthoic acid added in Example 1-2 increases the hydrophobic bond between clonidine and hydroxy naphthoic acid after forming a salt with clonidine. For this reason, it is thought that it plays a role in preventing clonidine from being lost to the water phase during the microsphere manufacturing process, and thus it is judged that the high content is shown.

Experimental Example 3: Measurement of in vitro drug release behavior of microspheres

In order to measure the release behavior of clonidine drug, 150.0 mg of each microsphere manufactured in Examples 1-1, 1-2, Comparative Examples 2, 3-3, and 3-6 was weighed. Then, each was placed in 50 ml of PBS (phosphate buffer saline, pH 7.4) and stored in an isotherm at 37°C. Thereafter, 1 ml was extracted for each date, and the amount of released drug was analyzed using an HPLC meter. The measured absorbance values were converted to obtain the concentration of the drug released for each date, and this was expressed by calculating the cumulative percentage. A clonidine solution was used as a control group in this experiment. The measurement results are shown in Figs. 3 and 4.

FIG. 3 is a graph showing the results of in vitro drug release behavior measurements obtained by conducting an experiment according to Experimental Example 3 on the microspheres manufactured in Examples 1-1, 1-2, and Comparative Example 2 according to the present invention. Compared to Comparative Example 2, Example 1 containing hydroxy naphthoic acid showed a higher drug release delay effect over time as well as an initial release inhibition effect. In addition, as the content of hydroxy naphthoic acid increased, not only the release delay effect improved but also a further improved zero-order release pattern was shown. It is thought that the addition of hydroxy naphthoic acid delays the release of the drug by strengthening the bond between clonidine and the biodegradable polymer inside the microspheres through the van der Waals force, thereby preventing the drug from rapidly leaking into the external water phase.

FIG. 4 is a graph showing the results of in vitro drug release behavior measurement obtained by conducting an experiment according to Experimental Example 3 on microspheres manufactured in Examples 1-2, Comparative Examples 3-3, and 3-6 according to the present invention. In the case of Example 1-2 containing hydroxy naphthoic acid, a hydrophobic solidifying agent, not only was the initial release noticeably delayed compared to Comparative Examples 3-3 and 3-6, but the drug release delay effect was also high. In addition, Example 1-2 showed a better zero-order release pattern. This is thought to be because the van der Waals force acts stronger than capric acid and mandelic acid when hydroxy naphthoic acid is added as a hydrophobic solidifying agent, but the present invention is not limited to this mechanism. As a result, it is judged that the bond between clonidine and the biodegradable polymer inside the microspheres becomes stronger, delaying the release of the drug.

Claims (6)

A sustained-release injection containing biodegradable polymer microparticles containing clonidine, wherein the microparticles contain hydroxy naphthoic acid, thereby slowly releasing clonidine from the microparticles. delete A sustained-release injection in claim 1, wherein the biodegradable polymer is at least one selected from the group consisting of polylactide (PLA), polyglycolide (PGA), poly(lactide-co-glycolide) (PLGA), and poly(lactide-co-glycolide) glucose. A step of preparing a solution comprising a biodegradable polymer; clonidine; and hydroxy naphthoic acid; and
A step of forming microspheres by adding the above solution to an aqueous solution and stirring to form an O/W emulsion,
A method for producing a sustained-release injection containing biodegradable polymer microparticles containing clonidine. delete In claim 4, the manufacturing method is characterized in that the encapsulation efficiency of clonidine in the microparticles (active ingredient encapsulated amount/added active ingredient weight × 100(%)) is 55 to 70 wt%.