KR101582191B1 - Prolamine-cation polymer complex contaning retinoid and manufacturing method thereof - Google Patents

Abstract

The present invention relates to a proline-cation polymer composite containing a retinoid and a method of preparing the same, and more particularly to a retinoid-cation polymer composite comprising a retinoid, a prolamin and a cationic polymer, And a method for producing the same.
According to the present invention, it is possible to provide a complex for capturing a retinoid, and to adjust the collection efficiency and the dissolution property by changing the molecular weight and concentration of the cationic polymer contained in the complex.

Description

TECHNICAL FIELD [0001] The present invention relates to a prolamin-cation polymer composite containing a retinoid, and a method for producing the same. BACKGROUND ART < RTI ID = 0.0 >

The present invention relates to a prodrillin-cationic polymer complex containing a retinoid and a process for its preparation.

For effective delivery, digestion, and absorption of active ingredients in the body, the captured active ingredients are retained in stomachs with high acidic conditions at low pH, and the contents slowly release in the intestines with relatively long retention times A system is required to control the system. Such systems require encapsulation, and such encapsulation techniques are techniques for trapping active ingredients in any material or tissue such that solid, liquid, or gaseous active ingredients can be released at controlled rates under specific conditions, A synthetic or natural polymer material is used as the material of the capsule.

Retinoids are a group of chemicals that are chemically related to vitamin A, and retinoids are substances that are primarily used as medicines by methods that control epithelial cell growth. Retinoids have a number of important and diverse functions in the body including vision, cell proliferation and differentiation, growth of bone tissue, immune function, and the activation role of tumor suppressor genes.

On the other hand, chitosan is a natural polymeric material obtained through deacetylation process from chitin and has very low toxicity and is excellent in biocompatibility [J. Microencapsulation, 2000, 17, 625 ~ 638], easy gelation and film formation, Lt; RTI ID = 0.0 > [J. < / RTI > Chem. Technol. Biotechnol., 1990, 49, 395-404], and also has excellent adsorptivity for jang [J. Pharm. Sci., 2003, 92, 567-576), there have been many studies on techniques for effectively delivering drugs or functional substances to the body through encapsulation using such chitosan.

Examples of the capsule manufacturing method using chitosan include emulsion cross-linking (Int. J. Pharm., 1994, 11, 217-222], emulsion droplet coalescence [Pharm. Res., 1999, 16, 1830 to 1835], reverse micellar method [J. Controlled Release, 2001, 17, 317-323], spray-drying (Int. Pharm., 1994, 217-222) or O / W / O multiple emulsion method (Korean Patent No. 10-2005-0014831).

However, in the preparation of capsules using chitosan, it is not easy to prepare capsules when only chitosan is used, and there is a lot of ionic gelling methods in which capsules are formed in a short time through rapid cross-linking reaction between polyvalent cationic chitosan and cross- It is being tried. Ionic gelation mainly forms an insoluble membrane by cross-linking reaction on the surface of the polymer, and is encapsulated through phase separation with the contents. The capsules thus prepared are mainly composed of synthetic or natural polymers and serve to control the release of the contents. At this time, the release of the contents is controlled by the chemical structure, thickness, and size of the capsule membrane, the concentration of the substance constituting the capsule, the concentration of the contents, the concentration of the crosslinking agent, and the pH (Int. J. Pharm., 2004, 274, 1-33, J. Controlled Release, 2004, 150, 5-28]. Glutaraldehyde and sulfuric acid were mainly used as crosslinking agents to be added for the ionic gelation of chitosan [J. Microencapsulation, 1998,15, 373-382; J. Microencapsulation, 2002,19, 173-180; Korean Patent 10-1998-0056584, 10-2003-0085599]. However, since these substances are toxic, their direct use in food is difficult and limited in use in pharmaceuticals.

In addition, many researchers have developed techniques for preparing chitosan capsules using low-toxicity crosslinking agents to deliver drugs to the body. In addition, various high molecular materials such as alginic acid, sodium carboxymethyl cellulose and xanthan gum are used as cross-linking agents for chitosan. In particular, attempts to manufacture chitosan capsules using tripolyphosphate (TPP) as a cross-linking agent have recently become mainstream. However, these methods have problems in encapsulating an active ingredient, such as low encapsulation yield, complicated manufacturing process, excessive release and decomposition of the above drug, and excessive release of the drug, so that the use of chitosan for encapsulation has various advantages Despite the presence of the commercialization degree and utilization is low. Therefore, it is necessary to prepare chitosan capsules which have a high yield and a high storage stability in the upper part by using a new type of cross-linking agent and are gradually released from the intestines.

Zein, a protein dissolved in alcohol, is present in the endosperm tissue of maize (Zea mais) and is known as a by-product of corn processing. They are GRAS (generally recognized as safe) approved by the US Food and Drug Administration (FDA), with about a quarter of them hydrophilic and three quarters of hydrophobic amino acid residues. Due to its high hydrophobicity, it has been used to capture lipophilic substances such as gitoxin and fish oil. Jane is also widely used in adhesives, biodegradable plastics, chewing gum, food coating agents, fibers, cosmetic powders, microencapsulated pesticides, and also in the form of microspheres or jane films for the capture of insulin and heparin.

Thus, the present inventors have reached the present invention in order to produce a delivery system capable of achieving the retention rate control in the intestine with high retention efficiency of the retinoid.

It is an object of the present invention to provide a prolaminic-cationic polymer composite containing a retinoid having a proper particle size, a stable zeta potential, a constant elution and an increased collection ratio using a cationic polymer of a polymer, and a method for producing the same.

In order to accomplish the above object, in one embodiment of the present invention, there is provided a composition comprising a retinoid, a prolamine and a cation polymer, wherein the prolamin and the cation polymer are selected from the group consisting of a prolamin- Lt; / RTI >

In one embodiment of the present invention, the preparation of a retinoid-containing prolamin-cationic polymer conjugate comprising mixing a retinoid, a prolamin and a cationic polymer, wherein the prolamin and the cationic polymer coat the retinoid ≪ / RTI >

In the present invention, there is provided a process for preparing a prolamin solution, comprising the steps of: a) dissolving prolamin in an organic solvent to obtain a prolamin solution;

b) dissolving the cationic polymer in an acid to obtain a cationic polymer solution;

c) dissolving the retinoid in an organic solvent to obtain a retinoid solution;

d) adding the retinoid solution to the prolamin solution to obtain solution 1;

e) adding Solution 1 to the cationic polymer solution to obtain Solution 2;

f) adjusting the concentration by adding a surfactant to the solution 2

A cationic polymer, and a pharmaceutically acceptable carrier.

In the present invention, the organic solvent in step a) may be an alcohol including but not limited to methanol, ethanol, isopropanol, acetone, dichloromethane, chloroform, ethyl acetate and toluene. Ethanol.

In the present invention, the acid in step b) is preferably selected from the group consisting of acetic acid, hydroxyacetic acid, propanoic acid, lactic acid, pyruvic acid, oxalic acid, malonic acid, succinic acid, maleic acid, fumaric acid, malic acid, tartaric acid, citric acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, cyclamic acid, salicylic acid, p-aminosalicylic acid and pamoic acid, preferably 1% lactic acid.

In the present invention, the organic solvent in step c) may be an alcohol including but not limited to methanol, ethanol, isopropanol, acetone, dichloromethane, chloroform, ethyl acetate and toluene. .

In the present invention, the addition of the step d) may be performed under stirring. Preferably, the stirring can be performed for about 1 hour, and the stirring speed can be about 600 rpm.

In the present invention, the addition of step e) may be performed under stirring. Preferably, the stirring can be performed for about 1 hour, and the stirring speed can be about 600 rpm.

In the present invention, 'surfactant' is a substance that adsorbs at the interface in a dilute solution and reduces its surface tension, which is also referred to as a surface active agent. Surfactants are substances that not only promote water uptake but also stably disperse the active ingredient regardless of the hydrogen ion concentration and temperature.

In the present invention, the surfactant may be an alkylene oxide-based surfactant. The alkylene oxide surfactant may be at least one selected from the group consisting of C16EO2, C12EO4, C16EO10, C16EO20, C18EO10, C16EO20, C18H35EO10, C12EO23, Tween 20, Tween 40, Tween 60, Tween 80, Span 40, Triton X- Tritol TMN-10, Pluronic L121, Pluronic L64, Pluronic P65, Pluronic P85, Pluronic P103, Pluronic P123, Pluronic F68, Pluronic F127 , Pluronic F88, Pluronic 25R4, Tetronic 908, Tetronic 901, and Tetronic 90R4, preferably Tween 20.

In the complexes and methods of manufacture of the present invention, retinoids include, but are not limited to, retinol, tretinoin, isotretinoin, alitretinoin, etretinate, acitretin, And at least one selected from the group consisting of adapalene, bexarotene, and tazarotene. Preferably, the retinoid may be retinol.

In the present invention, 'retinol' is a type of vitamin A and may be referred to as vitamin A in a narrow sense. Egg yolk, liver, milk is a lot, and the carotene in the plant is changed to vitamin A in the body. The epidermal cells of the skin play an important role in maintaining the original function.

In the present invention, 'prolamin' includes, but is not limited to, selected from the group consisting of zein, gliadin, hordein, secalin and kafirin And may be characterized by at least any one of them. Preferably, the prolamin can be a zein.

In the present invention, 'Jane' is a protein extracted from corn and is a kind of propolamine which is dissolved in alcohol.

In the present invention, the 'cationic polymer' may be chitosan. The chitosan may be poly [ß- (1-4) -2-amino-2-deoxy-D-glucopyranose] Chitosan, trimethylated chitosan, carboxymethyl chitosan and N- (2-hydroxypropyl-3-trimethyl ammonium chitosan chloride).

In the present invention, 'chitosan' is a cationic polymer having an amino group, which is a cation, and has a characteristic of strongly catching and discharging a harmful substance having an anion.

In the present invention, the cationic polymer may have a low molecular weight or a high molecular weight. The low molecular weight cationic polymer has a molecular weight of about 20 cp to about 300 cp, and preferably a high molecular weight cationic polymer, and can have a molecular weight of about 800 cp to about 2000 cp.

In the present invention, the prolamin to cation polymer may have a volume ratio of about 1:10 to about 1:40. Preferably, the propylamine to cationic polymer may have a volumetric ratio in the range of about 1: 5 to about 1:15 or about 1:15 to about 1:25.

In the present invention, the retinoid may be present in a concentration of about 0.1 ppm to about 500 ppm. Preferably from about 50 ppm to about 150 ppm, from about 150 ppm to about 250 ppm, and from about 250 ppm to about 350 ppm.

In the present invention, the propylamine-cationic polymer complex may have a particle size of from about 0.1 nm to about 2000 nm. Preferably, the particle size of the propylamine-cationic polymer complex may be from about 500 nm to about 1500 nm.

When the complex of the present invention is orally administered, the dissolution profile is shown by passing through the stomach (pH 1.2) and intestine (pH 7.4). Particularly, in the case of chitosan, the elution at pH 1.2 does not take place because it is not decomposed at the top.

The present invention provides a prodrillin-cationic polymer composite for coating a retinoid, and it is possible to control the stability of the complex, the collection efficiency and the dissolution property of the complex by changing the concentration of the retinoid, the molecular weight and the concentration of the cationic polymer, have.

1 is a graph showing the particle size and zeta potential of a complex according to the concentrations of zein and chitosan when low molecular weight chitosan and 100 ppm retinol are used.
FIG. 2 is a graph showing the particle size and zeta potential of a complex according to the concentrations of zein and chitosan when low molecular weight chitosan and 200 ppm retinol are used.
FIG. 3 is a graph showing the particle size and zeta potential of a complex according to the concentrations of zein and chitosan when low molecular weight chitosan and 300 ppm retinol are used.
FIG. 4 is a graph showing the particle size and zeta potential of a complex according to the concentration of zein and chitosan when high molecular weight chitosan and 100 ppm retinol are used.
FIG. 5 is a graph showing the particle size and zeta potential of a complex according to the concentrations of zein and chitosan when high molecular weight chitosan and 200 ppm retinol are used.
FIG. 6 is a graph showing the particle size and zeta potential of a complex according to the concentration of zein and chitosan when high molecular weight chitosan and 300 ppm retinol are used.
1: 10, 1: 10, Jane: 0.5% chitosan = 1: 20, Jane: 1% chitosan = 1: 10, 1: 20, Jane: 1.5% Chitosan = 1: 10, Jane: 1.5% Chitosan = 1: 20)
7 is a graph showing the particle size and zeta potential of nanoparticles according to the concentration of retinol (Fig. 7: ●: 100 ppm, ○: 200 ppm, ▼: 300 ppm)
8 is a graph showing elution of a complex according to the concentrations of zein and chitosan when low molecular weight chitosan and 100 ppm retinol are used.
9 is a graph showing elution of a complex according to the concentration of zein and chitosan when using low molecular weight chitosan and 200 ppm retinol.
FIG. 10 is a graph showing elution of a complex according to the concentration of zein and chitosan when using low molecular weight chitosan and 300 ppm retinol.
11 is a graph showing elution of a complex according to the concentrations of zein and chitosan when using high molecular weight chitosan and 100 ppm retinol.
12 is a graph showing elution of a complex according to the concentration of zein and chitosan when using high molecular weight chitosan and 200 ppm retinol.
FIG. 13 is a graph showing elution of a complex according to the concentration of zein and chitosan when using high molecular weight chitosan and 300 ppm retinol. FIG.
14 is a graph showing the elution of nanoparticles according to the concentration of retinol (in Fig. 14: ●: 100 ppm, ○: 200 ppm, ▼: 300 ppm)
15 is a graph showing the collection efficiency of the complex according to the molecular weight of chitosan, the concentration of zein and chitosan when 100 ppm of retinol is used.
16 is a graph showing the collection efficiency of the complex according to the molecular weight of chitosan, the concentration of zein and chitosan when using 200 ppm of retinol.
17 is a graph showing the collection efficiency of the complex according to the molecular weight of chitosan, the concentration of zein and chitosan when 300 ppm of retinol is used.
18 is a graph showing the collection efficiency of nanoparticles according to the concentration of retinol.
19 is an electron transmission microscope image of the surfaces of retinol-containing zein nanoparticles (A), zein-low molecular weight chitosan complex (B) and zein-high molecular weight chitosan complex (C).

Hereinafter, the present invention will be described in detail with reference to the following examples. However, the following examples are illustrative of the present invention, and the contents of the present invention are not limited by the following examples.

Example

Example  1-1. Preparation of Retinol-Containing Jane-Chitosan Complex

Each solution was prepared by dissolving jane in 75% ethanol, dissolving chitosan in 1% lactic acid, and dissolving retinol in methanol (purchased from Sigma, Jane, Chitosan and Retinol).

At the room temperature, the retinol solution obtained in the above manner was dropped into the Jane solution and stirred at 600 rpm for 1 hour to coat retinol with Jane (coating material). The chitosan solution obtained by the above method was dropped at a ratio of 1:20 (volume of the chitosan solution: coating material) while stirring at a speed of 600 rpm for 1 hour to encapsulate the zeta-chitosan complex containing retinol .

The mixture was stirred at a speed of 600 rpm while dropping 1% of Tween 20 to the complex, and the concentration of Tween 20 was adjusted to 0.44%. Ethanol was removed by using a concentrator to measure particle size, zeta potential and the like.

Example  1-2. Preparation of Retinol-Containing Jane Nanoparticles

For comparison with the complex of Example 1-1, a jane and retinol solution were prepared in the same manner as in Example 1-1, except for chitosan.

At the room temperature, the retinol solution obtained in the above manner was dropped into the Jane solution and stirred at 600 rpm for 1 hour to coat retinol with Jane (coating material).

The mixture was stirred at a speed of 600 rpm while dropping 1% of Tween 20 to the complex, and the concentration of Tween 20 was adjusted to 0.44%. Ethanol was removed by using a concentrator to measure particle size, zeta potential and the like.

Example  2. Identification of Retinol in Jane-Chitosan Complex and Jane Nanoparticles

The following conditions were used to analyze retinol. The complexes and nanoparticles of Example 1 were analyzed by HPLC (HITACHI, High-Technologies Corporation, Tokyo, Japan). At this time, C18 reverse phase column (3.9 × 300 mm, Waters, USA) was used for the column, and the column oven temperature was analyzed at 40 ° C. The mobile phase was measured with 75% methanol, fluorescence detector (Ex: 340 nm, Em: 460 nm). The instrumental analysis conditions of the HPLC used in the instrumental analysis are as follows.

Retinol HPLC  Analysis condition

Mobile phase methanol: distilled water (75:25, v / v)

Flow rate 0.8 mL / min

Detector HITACHI Fluorescence Detector L-2485

Detection wavelength (nm) Ex: 340 nm, Em: 460 nm

Column Waters μBondapak ™ C18 (10 μm, 3.9 × 300 mm)

Injection volume 10 μl

Example  3. Particle sizes of zein-chitosan complex and zein nanoparticles Zeta potential  Measure

In the production method of Example 1, composites were prepared by varying the conditions, and the particle size and the gentle potential were measured.

(1) Particle size measurement

The average particle size of the emulsion droplets was measured using a particle size analyzer (Microtrac nanotrac 250, Microtrac Inc., USA).

(2) Zeta potential  Measure

Since zeta potential is one of the important factors for stability, the zeta potential was measured for use in confirming stability. The higher the absolute value of the zeta potential, the higher the stability. The zeta potential was measured using a zeta potential analyzer (Zetaplus, Brookhaven Instruments corporation, USA).

3-1. The particle size of the zein-chitosan complex according to the addition amount of chitosan and Zeta potential  Measure

Chitosan of low molecular weight (20-300 cp) and high molecular weight (800-2000 cp) was added at 0.1, 0.5 and 1% of the total weight of the complex, and the particle size and zeta potential according to the concentration of each zein were measured.

As a result, the average particle size of the composite was about 1 탆 to about 2 탆, and the particle size also increased with an increase in the amount of chitosan. This tendency was conspicuous when high molecular weight chitosan was used. It was found that when the high molecular weight chitosan 1% was used (Jane: chitosan = 1:20), the average particle size decreased to 200 nm.

According to the results of the molecular weight of chitosan, it was found that when the high molecular weight chitosan was used, the particle size was lower than that of the low molecular weight chitosan. The zeta potential was measured at a level of about 20 mV to about 25 mV. On the other hand, as the content of retinol increased, the average particle size decreased and the zeta potential tended to decrease slightly (Figs. 1 to 6).

3-2. The particle size of Jane nanoparticles and Zeta potential  Measure

The particle size and zeta potential of the nanoparticles were measured by the method of Example 1 using only zein without adding chitosan, and the average particle size was about 3 nm to about 4 nm. The zeta potential was -30 mV (Fig. 7).

Example  4. Elution rate of Jane-chitosan complex and Jane nanoparticles

A predetermined amount of retinol-trapped zein-chitosan complex and zein nanoparticles prepared in Example 1 was placed in an elution solvent, and samples were collected over time using a dissolution testing apparatus (vision elite 8, Hanson, USA) , Tokyo, Japan) was used to measure retinol content. HCl solution (pH 1.2) and PBS buffer (pH 7.4) were used as the elution solvent, and the temperature was maintained at 37 ° C and the stirring speed was maintained at 100 rpm.

(Fig. 8 to 13), the dissolution of the complex containing chitosan was completed within about 6 hours to about 7 hours, compared with the dissolution test of the zein nanoparticles within about 3 hours.

In the case of the Jane nanoparticles, the elution was completed within a short time as the retinol content decreased (FIG. 14). At pH 1.2, no elution was observed.

Example  5. Jane-chitosan complex and zein nanoparticles Capture  efficiency

The encapsulation efficiency of retinol was calculated according to the following equation.

Collection efficiency (%) = (collected retinol content / total retinol content) x 100

As a result, it was confirmed that as the retinol content increases, the collection efficiency decreases in the case of the zein - chitosan complex. On the other hand, as a result of the measurement of the collecting efficiency according to the molecular weight of chitosan, the collecting efficiency increased when using a high molecular weight chitosan, compared with a low molecular weight, and the tendency was shown to be more than 80% (Figs. 15 to 17).

The retention efficiency of Jane nanoparticles was about 65% when the retinol content was 100 ppm. As with the Jane-chitosan complex, the retention efficiency tended to decrease as the retinol content increased (FIG. 18).

Example  6. Surface observation of Jane-chitosan complex and Jane nanoparticles

The surfaces of the zein-chitosan complexes and zein nanoparticles prepared using an electron transmission microscope (LIBRA 120 EFTEM, Carl Zeiss, Germany) were observed.

As a result, it was confirmed that the particle size distribution of the zein nanoparticles was smaller than that of the zein-chitosan complex (FIG. 19).

While the present invention has been described with reference to exemplary embodiments, it will be understood by those skilled in the art that various changes may be made and equivalents may be substituted for elements thereof without departing from the scope of the invention. You will know. In addition, many modifications may be made to adapt a particular situation and material to the teachings of the invention without departing from the essential scope thereof. Accordingly, it is intended that the invention not be limited to the particular embodiment disclosed as the best mode contemplated for carrying out this invention, but that the invention be construed as including all embodiments falling within the scope of the appended claims.

Claims (21)

Wherein said prolamin and chitosan comprise retinoids and retinoids are present in an amount of up to 200 ppm, wherein said prolamin and chitosan are selected from the group consisting of 1 < RTI ID = 0.0 > : ≪ / RTI > 5 to 1: 15. The method according to claim 1,
Wherein the retinoid is at least one selected from the group consisting of retinol, tretinoin, isotretinoin, alitretinoin, etretinate, acitretin, adapalene, bexarotene, and tazarotene. 3. The method of claim 2,
Wherein the retinoid is retinol. The method according to claim 1,
Wherein the prolamine is selected from the group consisting of pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine and pyrrolidine. 5. The method of claim 4,
Lt; RTI ID = 0.0 > furanamine < / RTI > delete The method according to claim 1,
(1-4) -2-amino-2-deoxy-D-glucopyranose], thiolated chitosan, trimethylated chitosan, carboxymethyl chitosan and N- (2- -Trimethylammonium chitosan chloride; and a prolamin-chitosan complex. The method according to claim 1,
Wherein the chitosan has a molecular weight of 800 cp to 2000 cp. The method according to claim 1,
Wherein the prolamin to chitosan has a volume ratio of 1:10. The method according to claim 1,
Wherein the retinoid is present in a concentration from 0.1 ppm to 200 ppm. A method for producing a prolamin-chitosan complex comprising mixing a retinoid, a prolamin and a chitosan, wherein the prolamin and the chitosan are coated with a retinoid and the retinoid content is 200 ppm or less, Wherein the large chitosan has a volume ratio of 1: 5 to 1: 15. 12. The method of claim 11,
a) dissolving the prolamin in an organic solvent to obtain a prolamin solution;
b) dissolving chitosan in an acid to obtain a chitosan solution;
c) dissolving the retinoid in an organic solvent to obtain a retinoid solution;
d) adding the retinoid solution to the prolamin solution to obtain solution 1;
e) adding Solution 1 to the chitosan solution to obtain Solution 2;
f) adjusting the concentration by adding a surfactant to the solution 2
≪ / RTI > or a pharmaceutically acceptable salt thereof. 12. The method of claim 11,
Wherein the retinoid is at least one selected from the group consisting of retinol, tretinoin, isotretinoin, alitretinoin, etretinate, acitretin, adapalene, bexarotene, and tazarotene. 14. The method of claim 13,
Wherein the retinoid is retinol. ≪ RTI ID = 0.0 > 11. < / RTI > 12. The method of claim 11,
Wherein the at least one compound is at least one selected from the group consisting of pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine, pyrrolidine and pyrrolidine. 16. The method of claim 15,
Lt; RTI ID = 0.0 > a < / RTI > prolaminemizer. delete 12. The method of claim 11,
(1-4) -2-amino-2-deoxy-D-glucopyranose], thiolated chitosan, trimethylated chitosan, carboxymethyl chitosan and N- (2- -Trimethylammonium chitosan chloride, and a method for producing the prodrillin-chitosan complex. 12. The method of claim 11,
Wherein the chitosan has a molecular weight of 800 cp to 2000 cp. 12. The method of claim 11,
Lt; RTI ID = 0.0 > 1: 10. ≪ / RTI > 12. The method of claim 11,
Wherein the retinoid is present in a concentration of from 0.1 ppm to 200 ppm.

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