KR20180085301A - pH-sensitive vesicle of surface-modified ion pair of cationic polymer and fatty acid - Google Patents

Abstract

The present invention relates to a pH-sensitive vesicle of a surface-modified phospholipid vesicle which is an ion pair of a cationic polymer and a fatty acid, and a manufacturing method thereof. The pH-sensitive vesicle emits relatively few contents installed therein by being stable in a neutral pH state, and emits relatively more contents installed therein by being unstable in acidic pH and basic pH states.

Description

BACKGROUND OF THE INVENTION [0002] pH-sensitive vesicles of surface-modified ion pairs of cationic polymers and fatty acids, modified with ionic pairs of cationic polymers and fatty acids,

The present invention relates to a pH-responsive endoplasmic reticulum modified with an ion pair of a cationic polymer and a fatty acid. More specifically, the present invention relates to a pH-responsive endoplasmic reticulum which is stable at a neutral pH by modifying a phospholipid vesicle surface with an ion pair of a cationic polymer and a fatty acid. (PH-responsive vesicle), which emits relatively little of the contents to be loaded, and which is destabilized in acidic and alkaline conditions to release the loaded contents abruptly, and a method for producing the same.

Amphiphilic molecules can be self-aggregated at an aqueous phase by an entropy-driven process [Barry, E., and Dogic, Z. (2010) PNAS , 107: 10348-10353.]. The structure of the self-assembly depends on the packing parameter of the amphipathic molecule (parameter indicating the shape of the amphipathic molecule), defined as V / LA, where V is the volume of the hydrophobic tail, L is the length of the hydrophobic tail And A is the cross-sectional area of the hydrophilic head). Amphiphilic molecules are known to form bilayer vesicles in aqueous phase when packing parameters are close to 1.

When the main component of bilayer vesicles is phospholipids, bilayer vesicles are referred to as liposomes. Phosphatidylcholine (PC) is a typical phospholipid and can be self-aggregated into an endoplasmic reticulum when dispersed in a water phase. Because the packing parameter has a value close to 1, the PC can form an endoplasmic reticulum without the aid of any auxiliary molecule.

The phospholipid elastomer has a problem in that it can not concentrate the active ingredient at a specific pH value since the active ingredient is gradually released by simple diffusion at the non-specified pH values. In order to effectively exert the efficacy of the active ingredient, the phospholipid elastomer must emit a relatively large amount of the effective ingredient at the pH value of the action site. For example, when the active ingredient is to be delivered to the skin, the phospholipid elastomer emits relatively small amount of the active ingredient at the storage conditions (neutral pH value) until it is applied to the skin, and after the application to the skin, To effectively exhibit the efficacy of the active ingredient.

Accordingly, the present invention was intended to develop a pH-responsive endoplasmic reticulum which releases relatively little of the contents loaded on a stable pH at neutral pH, and releases relatively large amounts of the loaded contents in both acidic and basic conditions.

Korean Patent Registration No. 10-1319642 (Registration Date: October 31, 2013) discloses a pH sensitive drug delivery system comprising a polymer formed by introducing 2,3-dimethylmaleic acid into an amine group of glycol chitosan For the preparation of nanocomposites.

The present invention modifies the surface of a phospholipid vesicle with an ion pair of a cationic polymer and a fatty acid so that it is stable at a neutral pH and relatively low in contents to be loaded and destabilized in acidic and basic forms, (PH-responsive vesicle) which rapidly releases the pH-responsive vesicle and its production method.

In order to achieve the above object, the present invention provides a pH-responsive endoplasmic reticulum in which the surface of a phospholipid is modified by the ion pair of a cationic polymer and a fatty acid.

In addition, the present invention relates to a method for preparing an ion-pairing solution of a cationic polymer / fatty acid by preparing a solution of a cationic polymer and a fatty acid to form an ionic pair of a cationic polymer / fatty acid, preparing a phospholipid solution, To form a pH-responsive endoplasmic reticulum.

The 'ion pair' is a combination of two ions having the same properties while having opposite charges. The ion pair of the present invention is formed by an electrostatic attractive force between the carboxyl groups and the amino groups of the fatty acid molecule and the cationic polymer, respectively do. Since cationic polymers are hydrophilic and the hydrocarbon chains of fatty acids are hydrophobic, ionic pairs of cationic polymers and fatty acids exhibit amphiphilicity.

In addition, the 'formula' means that the ion pair is attached to the phospholipid surface or inserted between the head groups of the phospholipid bilayer.

In addition, the 'pH responsiveness' means that the release of the target substance collected in the inside can be controlled according to the pH change.

Meanwhile, in the present invention, the cationic polymer is preferably selected from among polyethyleneimine (poly (ethylene imine), chitosan, and polylysine, and most preferably polyethyleneimine (poly ethylene imine).

In the present invention, the fatty acid is preferably selected from the group consisting of octanoic acid, decanoic acid, dodecanoic acid, myristic acid, palmitic acid, stearic acid, and behenic acid, and most preferably palmitic acid.

In the present invention, the phospholipid is preferably selected from the group consisting of Dioleoylphosphatidylethanolamine (DOPE), Dilauroylphosphatidylethanolamine (DLPE), Dimyristoylphosphatidylethanolamine (DMPE) And is selected from the group consisting of dipalmitoylphosphatidylethanolamine (DPPE), distearoylphosphatidylethanolamine (DSPE), palmitoyloleoylphosphatidylethanolamine (POPE), and most preferably diolureo Dioleoylphosphatidylethanolamine (DOPE) .

On the other hand, in the present invention, when the ionic polymer / fatty acid ion pair is formed in the present invention, the molar ratio of the amine group to the carboxyl group of the fatty acid in the cationic polymer is preferably 1: 0.001 to 0.01, more preferably 1: 0.0013 to 0.006, 1: 0.002 ~ 0.004 is preferable. If the ratio is lower than the above range, the amount of the cationic polymer is too small to stabilize the membrane of the endoplasmic reticulum. If the ratio is higher than the above range, the amount of the fatty acid is too small and the cationic polymer / fatty acid ion pair does not exhibit amphiphilicity .

On the other hand, in the present invention, the weight ratio of the phospholipid: ion pair of the pH-responsive endoplasmic reticulum whose phospholipid surface is modified with ion pairs is preferably 2 to 100: 1, more preferably 3 to 80: 1, Is preferably 5 to 50: 1. At a weight ratio lower than the above range, a reverse hexagonal phase is formed instead of an endoplasmic reticulum because an amount of an ion pair is too small to stabilize an endoplasmic reticulum membrane. In a weight ratio higher than the above range, This is because a mixed micelle is formed instead of an endoplasmic reticulum.

The present invention relates to a pH-responsive vesicle in which a phospholipid vesicle surface is modified with an ion pair of a cationic polymer and a fatty acid, and a preparation method thereof. The pH-responsive endoplasmic reticulum of the present invention comprises It is stable at a neutral pH to release comparatively less of the loaded contents, and is destabilized in acidity and basicity to exert an effect of relatively releasing the loaded contents.

Figure 1 is a conceptual diagram of a pH responsive DOPE endoplasmic reticulum stabilized with a PEI / PA ion pair.
Figure 2 is a schematic diagram of a PEI / PA (1/0) solution, a PEI / PA (1/1) solution, a PEI / PA (1/2) solution, And the surface tension of the solution (DELTA).
(A) of Figure 3 is the pH value of the release medium 4.5 (━━), 6.0 (‥‥ ), 7.4 (----), 9.0 enclosed in the endoplasmic reticulum (50/1) when (- - ‥) and release profile of the knife-old, (B) is the pH value of the release medium 4.5 (━━), 6.0 (‥‥ ), 7.4 (----), 9.0 (- ‥ -) days when the ER (200/1 ). ≪ / RTI >

The present invention provides a pH-responsive endoplasmic reticulum wherein the surface of a phospholipid is modified with an ion pair of a cationic polymer and a fatty acid. The phospholipid is preferably Dioleoylphosphatidylethanolamine (DOPE).

Dioleoylphosphatidylethanolamine (DOPE) is frequently used as a major constituent of phospholipid bilayer vesicles. However, DOPE is self-assembled into a reverse hexagonal phase at physiological pH values. The packing parameter of DOPE is greater than 1 (i.e., 1.41) because the acyl chain is unsaturated and the head is relatively small [Kumar, VV (1991) Proc . Natl . Acad. Sci . USA . , ≪ / RTI > 88: 444-448. Ancillary molecules are inserted between the head groups of DOPE molecules to help the DOPE molecule self-assemble into bilayer vesicles. The auxiliary molecule must be amphiphilic and the head group of the auxiliary molecule must be large enough to fill the space between the head groups of the DOPE molecule [Lee, EO, Kim, JG, and Kim, JD (1992) J. Biochem . , ≪ / RTI > 112: 671-676. Therefore, when the auxiliary molecules are dislocated or removed from the membrane of the endoplasmic reticulum, the double-layered endoplasmic reticulum is destroyed by the reversed-phase hexagonal phase and the contents of the endoplasmic reticulum are suddenly released. Thus, when DOPE is selected as a phospholipid, an amphiphilic auxiliary molecule is required, so that a rapid release effect of the content according to the pH can be expected.

In the present invention, a pH-responsive phospholipid bilayer was prepared by using ion pairs of a cationic polymer and a fatty acid as auxiliary molecules for forming DOPE endoplasmic reticulum. Fatty acid molecules and cationic polymers can form ion pairs by electrostatic attraction between their respective carboxyl and amino groups. Since cationic polymers are hydrophilic and the hydrocarbon chains of fatty acids are hydrophobic, ionic pairs of cationic polymers and fatty acids exhibit amphiphilicity. Thus, the ionic pair of cationic polymer and fatty acid is inserted between the head groups of the DOPE molecule to stabilize the bilayer to form the DOPE endoplasmic reticulum.

The cationic polymer chains are amino groups and conformational changes occur when the pH value is changed. Referring to FIG. 1, when the medium is acidified, the chain of the cationic polymer chains changes in a contracted form and an unfolded form. Due to the change in the chain shape, the cationic polymer / fatty acid ion pair loses its ability to stabilize the DOPE endoplasmic reticulum and the endoplasmic reticulum is destroyed, resulting in burst release (Fig. 1 (A)). In addition, when the medium is acidified, the electrostatic attraction between the cationic polymer and the fatty acid is reduced, so that the cationic polymer is desorbed from the fatty acid molecules and thus the endoplasmic reticulum is destroyed, resulting in burst release (Fig. 1 (B) ).

In addition, when the release medium is alkalized, the cationic polymer is deprotonated and the electrostatic attraction inside the molecule is reduced, so that the size of the molecule is reduced. The cationic polymers are shrunk and the packing parameter of the ion pair accordingly increases. In addition, alkalization reduces the ionization of the cationic polymer, reducing the electrostatic attraction of the cationic polymer and the fatty acid, thereby reducing the number of ion pairs. For this reason, when the release medium is alkalized, a sudden release from the DOPE endoplasmic reticulum occurs.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the scope of the present invention is not limited to the following embodiments, and includes modifications of equivalent technical ideas.

[ Example  One: Polyethyleneamine / Palmitic acid ( PEI / PA) mixed solution and surface tension measurement]

Polyethyleneimine (PEI), a cationic polymer, was dissolved in PBS (phosphate-buffered saline, 10 mM, pH 7.4) to a concentration of 4 mg / ml. Palmitic acid (PA), a fatty acid, was dissolved in the same buffer to give concentrations of 0.042 mg / ml, 0.084 mg / ml, and 0.2 mg / ml. 25 ml of PEI solution was mixed with the same volume of PA solution to prepare a stock solution of PEI / PA mixed solution. As a result, the concentration of PEI in the stock solution of PEI / PA mixed solution was 2 mg / ml, and the molar ratio of PEI / PA was 1/1, 1/2, 1/5. (1/1) solution, PEI / PA (1/2) solution, and PEI / PA (1/5) solution in which the molar ratio of PEI / PA is 1/1, 1/2, .

The stock solution of each mixed solution was continuously diluted by 2 times, and the surface tension of each of the diluted solutions was measured by a ring method using a surface tension machine (SEO, DST 60, South Korea). The surface tension of PEI solution (PEI / PA (1/0)) and PA solution was prepared as a control solution and the surface tension of each control solution was measured by the same method as that used for measuring the surface tension of PEI / PA mixed solution. The surface tension was measured by making the PA concentration in the control solution equal to the PA concentration in the PEI / PA (1/2) mixed solution.

The measurement results are shown in Fig. The surface tension was about 71.5 dyne / cm when the PEI concentration was 0, and the surface tension of the PEI / PA (1/0) solution did not show any significant change until the PEI concentration increased to 1 mg / ml. This is considered to be because PEI is a water-soluble polymer and thus it is difficult to place it at the air / water interface.

The surface tension of the PEI / PA (1/1) solution rapidly decreased to 42 dyne / cm when the PEI concentration was increased to 0.25 mg / ml and then increased to 40 dyne / cm. This is because the PA is attached to the PEI by the electrostatic attraction between the carboxyl group and the amino group. Since PEI is hydrophilic and the hydrocarbon chain of PA is hydrophobic, the PEI / PA ion pair exhibits surface-activity when PA is attached to PEI to form a PEI / PA ion pair. For this reason, the surface tension of the PEI / PA (1/1) solution decreases sharply with increasing PEI concentration.

The surface tension of the PEI / PA (1/2) solution decreased as the PEI concentration increased, and the surface tension profile was similar to that of the PEI / PA (1/1) solution. However, the surface tension of the PEI / PA (1/2) solution decreased faster than the surface tension of the PEI / PA (1/1) solution, and the plateau value in the area where the surface tension no longer changed Was lower than that of PEI / PA (1/1) solution. This means that the PEI / PA (1/2) ion pair has a higher surface activity than the PEI / PA (1/1) mixture.

HLB (Hydrophilic-lipophilic balance) is one of the major factors that indicate the surface activity of surfactants [Lee, EO, Kim, JG, and Kim, J. Biochem. , ≪ / RTI > 112: 671-676. When the molar ratio of PEI / PA is 1/2, the HLB value is lower than that of 1/1, so that PEI / PA (1/2) ion pairs are arranged more at the interface and result in high surface activity.

The surface tension profile of the PEI / PA (1/5) solution was almost the same as the surface tension profile of the PEI / PA (1/2) solution because the surface activity of the PEI / PA (1/5) PA < / RTI > (1/2) ion pair.

Figure 2 is a schematic diagram of a PEI / PA (1/0) solution, a PEI / PA (1/1) solution, a PEI / PA (1/2) solution, And the surface tension of the solution (DELTA).

[ Example  2: PEI / PA < / RTI > Diol leoyl Phosphatidyl  Preparation and Characterization of Dioleoylphosphatidylethanolamine (DOPE)

(One) PEI / PA < / RTI > Diol leoyl Phosphatidyl  Manufacture of Dioleoylphosphatidylethanolamine (DOPE)

2 ml of DOPE solution (solution dissolved in chloroform at a concentration of 10 mg / ml) was placed in a 50 ml round bottom flask and the organic solvent was evaporated under reduced pressure in a rotary evaporator to form a dry phospholipid film on the flask wall. At the same time, 0.1 mg to 1 mg of PEI / PA (1/2) ion pair and 1.8 mg of deoxycholate were dissolved in 2 ml of calcein solution (50 mM in PBS (10 mM, pH 7.4)). The mixed solution was put into a round bottom flask having a dried phospholipid membrane, and the flask was whirled by hand until the phospholipid film was completely removed from the flask wall and a suspension was formed. The weight ratio of phospholipid / (PEI / PA) ion pairs in the mixed suspension was 200/1, 100/1, 50/1, 20/1.

Each mixture suspension was sonicated in a bath-type sonicator (Ultrasonic processor, VCX 500, SONIX, USA) for 20 minutes and stopped for 20 seconds. To anneal the vesicle membrane, the vesicle suspension was left at room temperature for 4-5 hours. The vesicle suspension was passed through a Sephadex G-75 column (Sephadex G-75 column, 1.6 cm x 40 cm) and gel-chromatographed to remove the deoxylcholate and the non-encapsulated calcein. To the isolated vesicle suspension, PBS (10 mM, pH 7.4) buffer solution was added to give a final lipid concentration of 2 mg / ml. Thereafter, the endoplasmic reticulum prepared from the mixed suspension of phospholipid / (PEI / PA) ion pair weight ratio of 20/1, 50/1, 100/1, 200/1 was administered to the endoplasmic reticulum (20/1), the endoplasmic reticulum ), An endoplasmic reticulum (100/1), and an endoplasmic reticulum (200/1).

(2) PEI / PA < / RTI > Diol leoyl Phosphatidyl  Characterization of Dioleoylphosphatidylethanolamine (DOPE)

< Measurement of fluorescence intensities of calcans embedded in the endoplasmic reticulum>

The degree of quenching of the fluorescence intensity of the fluorescent substance encapsulated in the endoplasmic reticulum is a measure of the degree of vesicle formation (vesicle formation efficiency) [Hong, Y. J., and Kim, J. C. (2010) J. Nanosci. Nanotechnol., 10: 8380-8386.].

 The degree of quenching of the fluorescence intensity of the calcein (fluorescent substance) encapsulated in the DOPE endoplasmic reticulum containing the PEI / PA ion pair can be determined using Equation 1 below [Wang, M., and Kim , JC (2013) Colloid. Polym. Sci., 291: 2319-2327.]. The concentration of DOPE was 0.01% (w / v) in the vesicle suspension to measure the fluorescence intensity. Fluorescence intensity was measured by excitation with 484-nm light from a fluorescence spectrometer (F-2500, HITACHI, Japan) to measure the intensity of light at a wavelength of 524 nm.

[Equation 1]

% Quenching = (1 - F _i / F _f ) x 100

F _i = fluorescence intensity of the vesicle suspension measured after removal of the uncleaved calcein using chromatography

F _t : fluorescence intensity measured after completely dissolving the endoplasmic reticulum with Triton X-100

As a result, the degree of fluorescence of the calcein encapsulated in the DOPE ER containing PEI / PA ion pair decreased as the content of PEI / PA ion pair increased. Specifically, the fluorescence intensities of the calcein encapsulated in the endoplasmic reticulum (200/1), the endoplasmic reticulum (100/1), the endoplasmic reticulum (50/1) and the endoplasmic reticulum (20/1) were 70.7%, 68.7% 35.3% and 14%, respectively.

When the weight ratio of phospholipid / (PEI / PA) ion pair was 200/1 and 100/1, the degree of fluorescence was relatively high, indicating that the DOPE endoplasmic reticulum was efficiently formed at 200/1 and 100/1.

On the other hand, PEI / PA ion pair can form mixed micelles with DOPE when PEI / PA ion pairs are present in excess. If the PEI / PA ion pair is excessively inserted between the head groups of DOPE, the surface curvature of the DOPE aggregate will increase and become micelles. For this reason, the degree of fluorescence is relatively low when the ratio of DOPE / (PEI / PA) ion pair is high.

&Lt; Measurement of hydration diameter of the endoplasmic reticulum &

The hydration diameter of DOPE endoplasmic reticulum containing PEI / PA ion pairs was measured by light scattering. (20/1, 50/1, 100/1, and 200/1) were diluted with PBS (10 mM, pH 7.4) The hydration diameter was measured using a light scattering apparatus (ZetaPlus 90, Brookhaven Instrument Co., USA).

The average diameter of the ER was increased with increasing PEI / PA ion content. Specifically, the average diameters of the vesicle (200/1), the vesicle (100/1), the vesicle (50/1) and the vesicle (20/1) were 289 nm, 339 nm, 687 nm and 702 nm, respectively. When DOPE molecules are assembled into bilayer vesicles with the help of PEI / PA ion pairs, the hydrocarbon chains of PA are inserted into the phospholipid bilayer and the bulky PEI chains are directed to the water phase. Because of the PEI chain, when the PEI / PA ion pair is inserted into the endoplasmic reticulum membrane, the size of the endoplasmic reticulum becomes larger.

[ Example  3: Observation of accelerated emission according to change of pH value]

To observe the accelerated release according to the change of the pH value of the dioleoylphosphatidylethanolamine (DOPE) endoplasmic reticulum (200/1, 50/1) containing the PEI / PA ion pair prepared in Example 2 above.

1.9 ml of the buffer solution was placed in a cuvette for ultraviolet visible light spectrometer as a release medium and fixed in a cuvette holder. (Citrate buffer, 10 mM, pH 4.0, pH 5.0, pH 6.0), phosphoric acid buffer solution (PBS buffer, 10 mM, pH 7.4) and glycine buffer Medium. 0.1 ml of the vesicle suspension was injected into a 1.9 ml buffer solution contained in the cuvette and the fluorescence intensity was measured. The excitation wavelength was 494 nm and the emission wavelength was 524 nm. The degree of emission was calculated using the following equation (Wang, M., and Kim, J. C. (2013) Colloid. Polym. Sci., 291: 2319-2327.].

&Quot; (2) &quot;

% release = (F _t - F _i ) / (F _f - F _i ) x 100

F _t : fluorescence intensity at a given pH value

F _i : the value of the initial fluorescence intensity at pH 7.4

F _f : value of fluorescence intensity after completely dissolving the endoplasmic reticulum with Triton X-100

Since the endoplasmic reticulum was prepared at pH 7.4 and the release was observed at different pH values, the fluorescence intensities (F _t and F _f ) measured at different pH values were converted to values of fluorescence intensity at pH 7.4. Calibration curves (curves showing changes in fluorescence intensity with changes in calcein concentration) were made at the respective pH values of the release experiments.

The measurement results are shown in Fig. Figure 3 (A) is the release profile of the calcein encapsulated in the endoplasmic reticulum (50/1) when the pH of the release medium is 4.5, 6.0, 7.4, 9.0. At all pH values tested, the release rate increased rapidly and the release was almost complete in 10 minutes. The burst release from the DOPE endoplasmic reticulum is not due to simple diffusion through the endoplasmic reticulum membrane but because the endoplasmic reticulum membrane is disassembled.

Among the tested pH values, the release level was lowest at pH 7.4. Specifically, the release rates at pH 4.5, pH 6.0, pH 7.4 and pH 9.0 for 120 seconds were 85%, 24.1%, 10.1% and 62.0%, respectively. Since the pH value of the release medium used when making the ER was pH 7.4, the ER does not undergo a change in pH value when the ER is added to the release medium with a pH value of pH 7.4. Thus, the PEI / PA ion pair does not undergo a shape change of the molecule. For this reason, the endoplasmic reticulum is relatively stable in release at pH 7.4.

If the release medium is acidified, PEI becomes protonated and becomes bulky due to electrostatic repulsion within the molecule. According to the equation ^{(((1+ (10 (pK} -pH)) -1) -1 × 100) derived using the Henderson Hasselbalch equation for the basic compound, if the pK value of said amino group in the PEI 9 pH 7.4 , pH 6.0 and pH 4.5, the ionization degree of the amino group was calculated to be 97.5%, 99.9% and 100%, respectively. Since the pH value of the release medium decreases from pH 7.4 to pH 4.5, The shape of the PEI chain does not change much during the reduction from pH 7.4 to pH 4.5, but the shape of the PEI chain does not change significantly, but when the pH change is greater, for example, 4.5, the shape of the PEI chain is changed by the acidification, thereby promoting the release of the internal substance. In the present invention, since the range of pH change is narrow, it is not possible to observe the release of the internal substance by the above principle. To pH 4.5 During the decay, the PA molecules lose their electrostatic attraction with the PEI chains and fall out of the PEI chains, resulting in the release of internal material, which is the release of internal material by the loss of electrostatic attraction, A detailed explanation is as follows.

(1 / (1 + 10 ^(pK-pH) ) derived from the Henderson Hasselbalch equation for acidic compounds × 100), when the pK value of the carboxyl group of PA was 4.44, the ionization degree of carboxyl groups of PA was calculated as 99.9%, 97.3% and 53.4% at pH 7.4, pH 6.0 and pH 4.5, respectively. Thus, as the pH value of the release medium decreases from pH 7.4 to pH 4.5, the ionization degree decreases by almost 50%, so almost half of the PA molecules lose their electrostatic attraction with the PEI chains and are desorbed from the PEI chains. That is, half of the PEI / PA ion pair will disintegrate and lose the ability to stabilize the DOPE endoplasmic reticulum membrane. For this reason, the release medium is promoted when the release medium is acidified.

If the release medium is alkalized, the PEI molecule is depolarized and the electrostatic attraction within the molecule is reduced, thus reducing the size of the molecule. In fact, the ionization degree of the amino group of PEI at pH 7.4 and pH 9.0 was calculated to be 97.5% and 50%, respectively. Thus, when alkalized, the PEI molecules are shrunk and thus the packing parameter of the PEI / PA ion pair increases. In addition, alkalization reduces the ionization of PEI, which reduces the electrostatic attraction of PEI and PA, reducing the number of PEI / PA ion pairs. For this reason, when the release medium is alkalized, the release from the DOPE endoplasmic reticulum is promoted. On the other hand, at pH 7.4 and pH 9.0, the ionization degree of PA carboxyl groups was calculated as 99.9% and 100%, respectively. Since almost all of the carboxyl groups are ionized at pH 7.4 and pH 9.0, changes in PA ionization are not included in the reason that the release is promoted by alkalization.

FIG. 3 (B) is the release profile of the calcein encapsulated in the endoplasmic reticulum (200/1) when the pH of the release medium is 4.5, 6.0, 7.4, 9.0. The release profile was similar to the release profile of the calcein encapsulated in the vesicle (50/1). As with release from the endoplasmic reticulum (50/1), release was promoted when the release medium was acidified (decreased from pH 7.4 to pH 4.5) and basified (increased from pH 7.4 to pH 9.0). Specifically, the release rates for 120 seconds at pH 4.5, pH 6.0, pH 7.4 and pH 9.0 were 16.8%, 1.1%, 0.9% and 11.5%, respectively. At each pH value the degree of release was much lower than the degree of release from the endoplasmic reticulum (50/1). The PEI / PA ion pair is a pH-sensitive stabilizer, and therefore the DOPE endoplasmic reticulum containing a low content of PEI / PA ion pairs is less sensitive to changes in the pH value of the release medium. For this reason, the vesicle (200/1) is less sensitive to changes in the pH value in terms of release than the vesicle (50/1).

(A) of Figure 3 is the pH value of the release medium 4.5 (━━), 6.0 (‥‥ ), 7.4 (----), 9.0 enclosed in the endoplasmic reticulum (50/1) when (- - ‥) and release profile of the knife-old, (B) is the pH value of the release medium 4.5 (━━), 6.0 (‥‥ ), 7.4 (----), 9.0 (- ‥ -) days when the ER (200/1 ). &Lt; / RTI &gt;

Claims (8)

A pH - responsive endoplasmic reticulum with phospholipids modified by ion pairs of cationic polymers and fatty acids.
The method according to claim 1,
The cationic polymer may be,
Wherein the pH-responsive endoplasmic reticulum is any one selected from the group consisting of poly (ethylene imine), chitosan, and polylysine.
The method according to claim 1,
The fatty acid may be,
But are not limited to, octanoic acid, decanoic acid, dodecanoic acid, myristic acid, palmitic acid, stearic acid, behenic acid, And the pH-responsive endoplasmic reticulum.
The method according to claim 1,
The phospholipid,
(DOPE), dilauroylphosphatidylethanolamine (DLPE), dimyristoylphosphatidylethanolamine (DMPE), dipalmitoylphosphatidylethanolamine (DPPE), and the like. Wherein the pH-responsive endoplasmic reticulum is any one selected from the group consisting of distearoylphosphatidylethanolamine (DSPE), palmitoyloleoylphosphatidylethanolamine (POPE), and the like.
A cationic polymer / fatty acid mixture solution was prepared to form cationic polymer / fatty acid ion pairs,
After preparation into a phospholipid solution,
Wherein the ion-pair is modified on the surface of the phospholipid by adding a mixed solution of the phospholipid solution and the cationic polymer.
6. The method of claim 5,
The cationic polymer may be,
Characterized in that the pH-responsive endoplasmic reticulum is any one selected from the group consisting of poly (ethylene imine), chitosan, and polylysine.
6. The method of claim 5,
The fatty acid may be,
But are not limited to, octanoic acid, decanoic acid, dodecanoic acid, myristic acid, palmitic acid, stearic acid, behenic acid, Wherein the pH-responsive endoplasmic reticulum is selected from the group consisting of:
6. The method of claim 5,
The phospholipid,
(DOPE), dilauroylphosphatidylethanolamine (DLPE), dimyristoylphosphatidylethanolamine (DMPE), dipalmitoylphosphatidylethanolamine (DPPE), and the like. Wherein the surfactant is any one selected from the group consisting of distearoylphosphatidylethanolamine (DSPE), palmitoyloleoylphosphatidylethanolamine (POPE), and the like.

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