KR102353979B1 - Method of preparing for liposome nanoparticles via electrospraythe - Google Patents

Abstract

The present invention relates to polymer core-liposome shell nanoparticles capable of maintaining drug effects for a long time by increasing the drug loading content and reducing the drug release rate at the same time as compared to conventional liposomes, and a method for preparing the same.
The present invention provides polymer core-liposomal shell nanoparticles in which a polymer and a drug are loaded in the core using an electrospray device.
In the manufacturing method of the present invention, since the size of the core on which the polymer and the drug are loaded is formed several times larger than the shell of the phospholipid bilayer, the drug loading amount can be increased.
In addition, the polymer core-liposome shell nanoparticles of the present invention provide a drug delivery effect for a long time because the release rate to the outside is reduced by attraction with poly(D,L-lactide-co-glycolide) filled together in the core. can do.

Description

Method of preparing polymeric liposome nanoparticles prepared by electrospray {Method of preparing for liposome nanoparticles via electrospraythe}

The present invention relates to liposome nanoparticles prepared by electrospray and a method for preparing the same, and more particularly, to increase the drug loading content compared to conventional liposomes and reduce the drug release rate at the same time to maintain the drug effect for a long time. It relates to liposome nanoparticles and a method for preparing the same.

Liposomes are one of the materials that are in the spotlight as a means of gene delivery or drug delivery today. A general lipid-cholesterol-based liposome is a safe and effective carrier composed of a thermodynamically stable lipid bilayer surrounding the inner core with a water-soluble inner core.

Korean Patent Publication No. 0792557 discloses a case in which nanoparticles are prepared in the form of liposomes and used as a drug carrier. However, since the physical stability of particles using liposomes is not secured, various studies have been attempted to secure the stability. For example, the addition of certain surfactants to liposomes induced an increase in stability, addition of electrostatically charged lipids to liposome components, or the introduction of sterols, anionic lipids or sphingo lipids. As such, studies for improving the stability of liposomes have been continuously conducted, but the above techniques have insufficient physical stability, so there is a limit to their use in the pharmaceutical field.

Korean Patent No. 10-1695836 discloses a liposome surrounded by an amino acid-based polymer grafted with an alkyl chain. However, the liposome in the registered patent has excellent formulation stability with respect to surfactant and salt, but the drug content that can be loaded inside the liposome is still limited, and as the outside of the liposome is coated with a polymer, the drug must be loaded and released inside. It is difficult to use as a drug carrier.

An object of the present invention is to provide a liposome nanoparticle capable of maintaining a drug effect for a long time by reducing the release rate of the drug while containing a large amount of drug.

One aspect of the present invention is

mixing phospholipids, cholesterol, biodegradable polymers and drugs in a solvent;

It relates to a method for producing liposome nanoparticles comprising the step of spraying the solution with an electrospray device in a state in which a voltage is applied.

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The present invention provides polymer core-liposomal shell nanoparticles in which a polymer and a drug are loaded in the core using an electrospray device.

In the manufacturing method of the present invention, since the size of the core on which the polymer and the drug are loaded is formed several times larger than the shell of the phospholipid bilayer, the drug loading amount can be increased.

In addition, the polymer core-liposome shell nanoparticles of the present invention provide a drug delivery effect for a long time because the release rate to the outside is reduced by attraction with poly(D,L-lactide-co-glycolide) filled together in the core. can do.

1 is a conceptual diagram of a polymer core-liposome shell nanoparticles.
2 is an SEM image of the liposome nanoparticles prepared in Example 1, Comparative Example 1, and Comparative Example 2.
3A is a drug release profile of Comparative Example 3 and Example 1.
Figure 4A shows the drug release mechanism of Comparative Example 2 (phospholipid, cholesterol + acetyl curcumin), Figure 4B shows the drug release mechanism of Example 1.

Hereinafter, a preferred embodiment of the present invention will be described with reference to the drawings. However, the scope of the present invention is not limited to the description or drawings of the following embodiments. That is, the terms used herein are used only to describe specific embodiments, and are not intended to limit the present invention. The singular expression includes the plural expression unless the context clearly dictates otherwise. In addition, terms such as "comprises" or "have" described in this specification are intended to designate that the features, numbers, steps, operations, components, parts, or combinations thereof described in the specification exist, but one or the It should be understood that the above does not preclude the possibility of the existence or addition of other features or numbers, steps, operations, components, parts, or combinations thereof.

The polymer core-liposomal shell nanoparticle preparation method of the present invention includes a mixing step and a spraying step.

The mixing step is a step of mixing phospholipids, cholesterol, biodegradable polymers and drugs in a solvent.

As the phospholipid, a known phospholipid may be used. For example, the phospholipid is phosphatidylcholine, soybean lecithin, lysolecithin, sphingomyelin, phosphatidic acid, phosphatidylserine, phosphatidylglycerol, phosphatidyl inositol, phosphatidylethanolamine, diphosphatidylglycerol, cardiolipin (cardiolipin) and a natural phospholipid selected from the group consisting of plasmalogen; hydrogenation products of natural phospholipids; Dicetylphosphate, distearoylphosphatidylcholine, dioleoylphosphatidylethanolamine, dipalmitoylphosphatidylcholine, dipalmitoylphosphatidylethanolamine, dipalmitoylphosphatidylserine, eleostearoylphosphatidylcholine, eleostearoylphosphatidylethanolamine and ele It may be a synthetic lipid of ostearoylphosphatidylserine or a derivative of a synthetic lipid.

The biodegradable polymer may be PLGA (Poly (D,L-lactide-co-glycolide)), polycaprolactone (PCL), PLA (poly (lactic acid)), PGA (poly (glycolic acid)). .

The drug may be one in which an amino group or a hydrogen of a hydroxy group is substituted with an acetyl group. The drug may be an acetylated drug.

The drug may be a hydrophobic drug.

The drug is a known drug, and an anti-inflammatory agent, an antioxidant, and an anticancer agent in which these drugs are acetylated may be used.

The drug may be acetyl curcumin.

The solvent is dichloromethane (DCM), acetone.

The mixing step may include 20 to 30 parts by weight of cholesterol, 30 to 70 parts by weight of a biodegradable polymer, and 5 to 20 parts by weight of a drug relative to 100 parts by weight of the phospholipid.

The spraying step is a step of spraying the solution with an electrospray device while applying a voltage.

The spraying step may be sprayed at a rate of 0.5 to 1 ml/hr under 13-17 kV electric field conditions.

In the spraying step, an electric field is formed by applying a voltage, and the balance between the electrostatic force, which is a force that pulls the liquid by the electric field, and the surface tension of the liquid on the surface of the liquid is broken, and a liposome structure can be formed.

The spraying step may spray the mixed solution at a high voltage of 13 to 17 kV and a relatively fast rate of 0.5 to 1 ml/hr.

In the spraying step, the spraying distance (eg, the distance between the syringe nozzle tip and the collector plate) may be 5-20 μm.

1 is a conceptual diagram of a polymer core-liposome shell nanoparticles, and FIG. 2 c is an SEM image of the nanoparticles prepared in Example 1.

1 and 2 c, the polymer core-liposome shell nanoparticles of the present invention have a core including the biodegradable polymer and a drug, and the nanoparticle shell is a bilayer polar head portion of a liposome.

Figure 2a is a liposome prepared from phospholipids and cholesterol, Figure 2b is a liposome prepared by mixing phospholipids, cholesterol, and a drug (acetyl cumin), Figure 2c is a liposome prepared by the method of the present invention ( Electrospray of phospholipids, cholesterol, biodegradable polymers and drugs) liposomes.

For the drug, biodegradable polymer, phospholipid and cholesterol of the present invention, the above-mentioned contents can be referred to.

That is, the drug is acetylated, and preferably, the drug may be acetylated curcumin.

The biodegradable polymer may be poly(D,L-lactide-co-glycolide) (Poly(D,L-lactide-co-glycolide), PLGA).

The size of the core may be 370 ~ 420 nm, and the size of the shell may be 25 ~ 50 nm. The overall size of the liposome in the present invention can be seen from 395 to 470 nm.

The total diameter of a in FIG. 2 is about 200 to 250 nm. The total diameter of FIG. 2 b is about 300-350 nm. In the case of the present invention, the size of the core was increased up to about 2 times or more compared to the overall size of the liposome in FIG. 2 a and FIG. 2 . As a result, the polymer core-liposome shell nanoparticles of the present invention can increase the amount of drug loading compared to conventional liposomes.

The liposomes as shown in a and b of FIG. 2 have a hollow structure and have a small diameter because the outer membrane and the inner membrane are hydrophobic. The biodegradable polymer of the present invention has a high molecular weight and a large surface area. As the biodegradable polymer enters the liposome core, the diameter of the core increases that much due to the unique structure (polymer matrix) with a large surface area.

The polymer core-liposome shell nanoparticles of the present invention are filled with a drug and a biodegradable polymer, so that the release rate of the drug to the outside can be reduced by the attraction between them.

The release rate of a drug is reduced because it is in intermolecular hydrogen bonds with molecules in a polymer or liposome, or is bound by electrostatic or van der Waals forces. The drug in the liposome is bound to the liposome or polymer, and in order to release the drug out of the liposome, the above bond must be broken, thereby slowing the drug release rate.

Hereinafter, the present invention will be described in detail with reference to the accompanying embodiments and drawings. However, the accompanying examples only illustrate specific embodiments of the present invention, and are not intended to limit the scope of the present invention.

Example 1: Synthesis of NGCDs

Acetyl Curcumin Synthesis

0.542 mmol (200 mg) of (1E,6E)-1,7-bis(4-hydroxy-3-methoxyphenyl) hepta-1,6-diene-3,5-dione (curcumin) was mixed with 5ml of acetaldehyde and 5ml of It was placed in a flask containing ethanol. The reaction mixture was refluxed for 20 minutes in the presence of an acidic medium (100 μL H2SO4 0.001 N). The color of curcumin changed from yellow to dark brown. After cooling, it was filtered with distilled water to obtain a dark brown solid acetylated curcumin.

liposome synthesis

i) 100 mg of L-α-phosphatidylcholine (phospholipid) and 25 mg of cholesterol were mixed in a bottle containing 1 mL DCM (dichloromethane) (Comparative Example 1, pure liposome-LNPs preparation)

ii) 100 mg of L-α-phosphatidylcholine (phospholipid) and 25 mg of cholesterol were mixed in a bottle containing 1 mL DCM (dichloromethane). 50 mg of PLGA was added to this solution (phospholipid, cholesterol + PLGA, Example 1 - PCSLNPs preparation)

8.1 mg of the drug (curcumin or AC) was put into the two solutions prepared above and mixed.

The prepared three solutions (Comparative Example 1, Comparative Example 2 (drug mixture in Comparative Example 1), Example 1) were put into Becton Dickinson (BD) Luer-Lok™ syringes, and a 26G needle was used. A 10 μm gap was placed between the syringe and the collector.

Under a 15 KV electric field, each solution was sprayed onto the glass by pumping at a rate of 0,6 mL/h. After spraying, the solvent was removed, and 20ml PBS was added to obtain the liposomes prepared in Comparative Example 1, Comparative P and Example 1.

Experiment: Drug Release Profile

5 mL of drug-loaded LNPs were put into a dialysis membrane (50 mL PBS buffer, pH 5.2, 7.4), and then the membrane was inserted into a beaker. The drug release profile was investigated by measuring the absorption rate of the released drug using a UV-vis spectrophotometer.

Figure 2a is the liposome prepared in Comparative Example 1, Figure 2b is the liposome prepared in Comparative Example 2, Figure 2c is Example 1 (phospholipid, cholesterol, biodegradable polymer and drug electrospray ) is an SEM image of the prepared liposome. In comparison, the liposome sizes of 1 and 2 are 200-250 nm and 300-350 nm, respectively, and the size of Example 1 is 395-470 nm. The liposome size of Example 1 was increased.

3A is a drug release profile of Comparative Example 3 (the same structure as in Example 1, but the drug is unacetylated curcumin), and FIG. 3B is a drug release profile of Example 1. Referring to FIG. 3 , it can be seen that the liposome of Example 1 releases the drug more slowly than Comparative Example 3.

Figure 4A is Comparative Example 2 and shows the drug release (curcumin/AC (acetylcurcumin)) mechanism of LNPs, and Figure 4B is Example 1 and shows the drug release (curcumin/AC (acetylcurcumin)) mechanism of PCSLNPs.

Referring to FIG. 4 , in Example 1 (containing PLGA), AC (acetylated curcumin) is difficult to release to the outside of the liposome by attraction with the polymer (PLGA).

In the above, although preferred embodiments of the present invention have been described in detail, these are only for the purpose of explanation, and the protection scope of the present invention is not limited thereto.

Claims (11)

mixing phospholipids, cholesterol, biodegradable polymers and drugs in a solvent;
As a method for preparing liposome nanoparticles comprising the step of spraying the mixed solution with an electrospray device in a state in which a voltage is applied,
The biodegradable polymer is PLGA (Poly (D,L-lactide-co-glycolide)), polycaprolactone (PCL), PLA (poly (lactic acid)) or PGA (poly (glycolic acid)),
The drug is acetylated, hydrophobic,
The method comprises mixing 20-30 parts by weight of cholesterol, 30-70 parts by weight of a biodegradable polymer, and 5-20 parts by weight of a drug with respect to 100 parts by weight of the phospholipid in the solvent,
The solvent is dichloromethane (DCM) or acetone,
The spraying step is spraying at a rate of 0.5 to 1 ml/hr under 13-17 kV electric field conditions,
In the spraying step, when the solution is sprayed under voltage applied, the balance between the electrostatic force, which is a force that pulls the liquid by an electric field from the liquid surface of the sprayed solution, and the surface tension of the liquid is broken, and the core-shell lipid bilayer structure of liposome nanoparticles is formed,
The core includes the biodegradable polymer and a drug,
The size of the core is 370 ~ 420nm, and the size of the shell is 25 ~ 50nm liposome nanoparticle manufacturing method, characterized in that. delete delete The method according to claim 1, wherein the drug is acetylated curcumin.
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