KR20190036767A - Salt ion-responsive lipid cubic nanostructures and methods thereof - Google Patents

Abstract

The present invention relates to a cubic lipid nanostructure having responsiveness to salt ions and a method for manufacturing the same. More specifically, the present invention relates to: the cubic lipid nanostructure having responsiveness to salt ions, capable of responding to salt ions to easily control the release of an active component contained in a lipid nanostructure by comprising an anionic polymer in a water channel and crosslinking the same with a polyvalent cation for gelation; a use of the lipid nanostructure; and the method for manufacturing the same.

Description

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a cubic lipid nanostructure having a salt-ion responsive property and a preparation method thereof.

The present invention relates to a cubic lipid nanostructure having responsiveness to salt ions and a method for producing the same. More particularly, the present invention relates to a method for preparing a lipid nanostructure capable of responding to a salt ion and facilitating the release of an active ingredient contained in the lipid nanostructure by allowing an anionic polymer to be contained in a water channel and cross- And a method for producing the same. 2. Description of the Related Art

Lipid molecules have high biocompatibility due to the characteristics derived from the living body. Due to the amphipathic nature of the molecules, nanostructures can be formed through a relatively simple manufacturing process compared to other materials. The lipid nanostructure can be formed into various types of nanostructures such as a lamellar structure, a cylinder structure, and a micelle structure based on molecular properties such as HLB (Hydrophilic Lipophilic Balance) and packing parameters.

Monoolein (MO) forms a cubic nanostructure when it reaches an equilibrium state by contacting with an appropriate amount of water. The cubic nanostructure consists of a lipid bilayer surrounding the water channels and the water channels intersecting each other [Razumas, Valdemaras., Et al. The Journal of Physical Chemistry 100.28 (1996): 11766-11774.].

The monoolein nanostructure is transparent and optically isotropic. The monoolein nanostructure comprises a water channel and a lipid matrix, and is capable of containing both a water soluble active ingredient and a fat soluble active ingredient.

The active ingredient collected in the nanostructure may be released from the water channel. In order to be used as an effective ingredient carrier, it is necessary to control the rate of release or control the release according to external environmental conditions.

On the other hand, an anionic polymer such as alginate is dissolved in an aqueous solution, and when a polyvalent cation is added thereto, the polymer chains are crosslinked to form a gel [Augst, Alexander D., et al. Macromolecular bioscience 6.8 (2006): 623-633.].

The anionic polymer may undergo reversible changes in sol gel properties due to crosslinking depending on the ion, but there is no example applied as an effective component carrier. Accordingly, there is a need for research and development of a technique capable of releasing an effective substance trapped in the structure more efficiently by taking advantage of the characteristics of the ionic polymer.

Augst, Alexander D., et al. Macromolecular bioscience 6.8 (2006): 623-633.

It is an object of the present invention to provide a lipid nanostructure capable of being sensitive to salt ions.

Another object of the present invention is to provide an effective component delivery system which contains an ionic polymer and an active ingredient in a water channel and releases the active ingredient out of the water channel in response to salt ions.

Another object of the present invention is to provide a method of releasing an active ingredient which can control the amount of active ingredient released and released.

It is another object of the present invention to provide a method for producing a lipid nanostructure capable of being sensitive to salt ions.

One aspect of the present invention is to provide a salt-ion responsive lipid nanostructure comprising an anionic polymer gel, which is a crosslinked product of an anionic polymer and a polyvalent cation, in a water channel.

In the lipid nanostructure according to an embodiment of the present invention, the lipid contained in the lipid nanostructure may include a monoolein cubic phase.

In the lipid nano-structure according to one embodiment of the present invention, the multivalent cation is ^{Mg 2 +, Ca 2 +,} Sr 2 +, Ba 2 +, Be 2+, Cr 2 +, Co 2 +, Cu 2 +, ^{Fe 2 +, Mn 2 +,} Sn 2 +, Ni 2 +, Zn 2 +, Al 3 +, Cr 3 +, Co 3 +, Cu 3 +, Ga 3 +, Au 3 +, Fe 3 +, Mn 3 ^+, Ni ^{+ 3,} Mn ^{+ 4,} it may be one containing any one or more cations selected from the group consisting of Sn ^4+, Cr ⁶⁺ and Mn ^7+.

In the lipid nanostructure according to an embodiment of the present invention, the anionic polymer may include a carboxyl group (-COOH), a sulfonic acid group (-SO ₃ H), a hydroxyl group (-OH), a phosphinic group (-HPO ₂ H ), phosphonate nikgi (-PO ₃ H _2), O sonic group (-AsO ₃ H ₂₎ and cells Reno nikgi (including an anionic polymer having one or more features selected from the group consisting of -SeO ₃ H) .

In the lipid nanostructure according to an embodiment of the present invention, the anionic polymer may have a weight average molecular weight of 400 to 100,000 g / mol.

The lipid nanostructure according to an embodiment of the present invention may include 2 to 9 parts by weight of an anionic polymer based on 100 parts by weight of lipid molecules.

In the lipid nanostructure according to an embodiment of the present invention, the cation may be included in an amount of 15 to 50 mol% based on the number of moles of anionic groups in the anionic polymer.

The lipid nanostructure according to an embodiment of the present invention may include an active ingredient that is released to the outside of a water channel in response to a salt ion.

Another aspect of the present invention relates to an effective ingredient delivery system comprising the above-mentioned lipid nanospheres.

Another aspect of the present invention is to adjust the amount of the active ingredient released from the water channel according to the response of the salt ion by using the anionic polymer gel contained in the water channel and the cubic lipid nanostructure containing the active ingredient And a method for releasing a salt-ion responsive active substance.

In another aspect of the present invention, there is provided a method of preparing a monoolein nanostructure comprising the steps of: preparing an aqueous solution of an anionic polymer by adding a monoolein nanostructure to a monoolein solution; and adding a polyvalent cation compound to the prepared monoolein nanostructure, And a step of forming a gel. The present invention also relates to a method for producing a salt-ion responsive cubic lipid nanostructure.

In the method for preparing a lipid nanostructure according to an embodiment of the present invention, the anionic polymer aqueous solution may further comprise an active ingredient.

The lipid nanostructure according to the present invention has an advantage that the release of the active ingredient contained in the inner water channel can be easily controlled.

In addition, the lipid nanostructure according to the present invention is capable of controlling the release and release rate of the active ingredient by being sensitive to the external environment, that is, the salt ion. Specifically, the lipid nanostructure has excellent storage stability, It has an advantage that it releases the active ingredient, increases the release rate of the active ingredient, or releases the active ingredient slowly over a long period of time.

The present invention also provides a method for manufacturing a cubic lipid nanostructure including an ionic polymer gel having a water channel having a diameter of 10 nm or less and being sensitive to salt ions by a simple method.

FIG. 1 is a schematic view of a reduction-responsive monoolein cubic nanostructure according to an embodiment of the present invention. (A) shows a case in which an ionic polymer gel is filled in a water channel, and (B) shows a case where a nanostructure is exposed to a salt solution having a salt ion, and when the ionic polymer gel filled in the water channel is crosslinked It is seen that the gel is broken down and changed into a sol.
Fig. 2 shows a differential scanning calorimetric curve of the cubic lipid nanostructure prepared in Examples and Comparative Examples.
3 is a transmission electron micrograph of the cubic lipid nanostructure (A) prepared in Comparative Example 1 and the cubic lipid nanostructure prepared in Example 2 when heated at 25 ° C to 80 ° C.
FIG. 4 is a transmission electron micrograph of the cubic lipid nanostructure prepared in Example 2 when heated at 25 ° C. to 80 ° C. FIG.
5 is a graph showing the degree of release of the dye when the cubic lipid nanostructure according to Comparative Example 1 is contacted with distilled water () and PBS (10 mM, pH 7.4) (?).
6 is a graph showing the degree of dye release when the cubic lipid nanostructure according to Example 1 is contacted with distilled water () and PBS (10 mM, pH 7.4) (○).
7 is a graph showing the degree of release of the dye when the cubic lipid nanostructure according to Example 2 is contacted with distilled water () and PBS (10 mM, pH 7.4) (?).
Figure 8 shows the relative optical density over time of alginate gelled in distilled water (●) and PBS (10 mM, pH 7.4) (○).

Hereinafter, a cubic lipid nanostructure of the present invention, a method for producing the same, and uses thereof will be described in detail with reference to the accompanying drawings. The following drawings are provided by way of example so that those skilled in the art can fully understand the spirit of the present invention. Therefore, the present invention is not limited to the following drawings, but may be embodied in other forms, and the drawings presented below may be exaggerated in order to clarify the spirit of the present invention. Hereinafter, the technical and scientific terms used herein will be understood by those skilled in the art without departing from the scope of the present invention. Descriptions of known functions and configurations that may be unnecessarily blurred are omitted.

Unless defined otherwise in the present invention, 'monoolein' means glycerol monooleate (monoolein; 1-oleoyl-rac-glycerol) with a packing parameter of slightly greater than 1.

Unless otherwise defined herein, the term " cubic " may refer to a cubic phase that is optically transparent and has an intercalated water channel in the lipid matrix. The water channels are separated by a bilayer of monoolein and may contain a water soluble compound within the water channel. The water channel may have a diameter of 10 nm or less, specifically 2 nm to 7 nm, but the diameter of the water channel is not limited to the diameter because it can vary depending on the environment of the water surface.

Unless otherwise defined in the present invention, the 'active ingredient' means a material excluding the skeleton of the lipid nanostructure and an anionic polymer contained therein and a polyvalent cation that forms a crosslinked gel with the ionic polymer, A peptide, or a protein having solubility in a cell. The active ingredient may have the same meaning as the " physiologically active ingredient ", and a water-soluble monomolecular compound, a water-soluble peptide, or a water-soluble protein may be exemplified without limitation.

In a non-limiting example, the active ingredient may include a cationic group in the molecule of the active ingredient so as to have an electrostatic interaction with the anionic group of the anionic polymer. By including a cationic group in the molecule, the release of excessive active ingredient can be suppressed initially and electrostatic interactions with the anionic group of the anionic polymer can be induced and sustained release can be induced. Also, when a nanostructure is exposed to a cation, the cation can form a salt with the anion group, so that the active ingredient can be released rapidly. The cationic group of the active ingredient may be an amine group, an ammonium group, an imidazole group, and the like, but is not limited thereto.

Unless defined otherwise in the present invention, 'gel' refers to a colloidal particle that forms a crosslinked structure due to inter-ion bonding and represents a semi-solid phase, and 'sol' refers to a crosslinked structure Is disintegrated and the colloidal particles are dispersed.

The inventor of the present invention has found that when releasing the active ingredient collected in the water channel inside the lipid nanostructure to the outside of the water channel, the active ingredient is stably trapped in the water channel without releasing the effective ingredient to the outside of the inner water channel In the study on lipid nanostructures whose release performance is controlled when they are applied to a target substance, the anionic polymer gel, which is a crosslinked mixture of anionic polymer and polyvalent cation, is contained in the water channel in the lipid nanostructure, The present inventors have found that the nanostructure efficiently releases the active ingredient in response to salt ions and can control the release amount and the release rate.

Hereinafter, the lipid nanostructure according to the present invention will be described in detail.

The cubic lipid nanostructure according to one embodiment of the present invention includes an anionic polymer gel, which is a crosslinked product of an anionic polymer and a polyvalent cation, in a water channel.

The lipid nanostructure may include a water channel therein. The water channel may have a diameter of 10 nm or less, specifically 2 nm to 7 nm, more specifically, 3 nm to 5 nm, The diameter of the water channel is not limited to the above diameter since it is variable depending on the environment of the water phase.

The lipid molecule of the lipid nanostructure may be preferably a monoolefin and is not limited to a particular molecule if it is a lipid molecule with a packing parameter of about 1.

The anionic polymer is contained in the lipid nanostructure and includes a carboxyl group (-COOH), a sulfonic acid group (-SO ₃ H), a hydroxyl group (-OH), a phosphonic group (-HPO ₂ H), a phosphonic group (-PO ₃ H _2), O sonic group (-AsO ₃ H _2), may be a polymer containing in the cell Reno nikgi (-SeO ₃ H) and repeating the anionic moiety is selected from a salt units, non-limiting (Meth) acrylic acid polymer, poly (styrene sulfonate) polymer, poly (isopropenylphosphate) polymer, and the like, and a polymer such as polyacrylic acid, May be a copolymer thereof.

The anionic polymer may have a weight average molecular weight of 400 to 100,000 g / mol, and preferably 1,000 to 50,000 g / mol, without limitation. When the above range is satisfied, it can be homogeneously mixed with lipid molecules, and it can be advantageous to realize release control performance of an active ingredient by forming a crosslinked structure with polyvalent cations, but this is only a limited example, I do not accept. In addition, the anionic polymer may have a viscosity (25 ° C) of 1 to 100 cP, specifically 2 to 80 cP. When the above range is satisfied, uniform mixing of lipid molecules as well as production of anionic polymer gel is more effective, but this is a non-limiting example and is not limited to the above-mentioned numerical range.

In the cubic lipid nanostructure according to an embodiment of the present invention, the anionic polymer may be included in an amount of 0.01 to 12 parts by weight, specifically 2 to 9 parts by weight, based on 100 parts by weight of lipid molecules. When the above range is satisfied, formation of a cubic nanostructure is smooth by uniform mixing with lipid molecules, and cross-linking with a polyvalent cation is effective in gelation and trapping and releasing of an effective component. However, Is not limited to the above numerical range.

The cubic lipid nanostructure according to an embodiment of the present invention is characterized in that the anionic polymer includes an ionic polymer gel obtained by gelation by cross-linking with polyvalent cations.

The anionic polymer gel forms a crosslinked structure of an anionic polymer and a polyvalent cation. Specifically, the anionic polymer gel crosslinks between different ionic polymer chains by polyvalent cations and they form a network structure, resulting in a semi-solid phase in which fluidity is lost. This can lower the degree of diffusion of the active ingredient in the water channel when the effective ingredient is collected inside the water channel, and thus can effectively suppress the release of the effective ingredient to the outside of the water channel.

The polyvalent cation that gels the anionic polymer has a divalent or higher ionic group. A non-limiting example of said multivalent ^{cations, Mg 2 +, Ca 2 +} , Sr 2 +, Ba 2 +, Be 2 +, Cr 2 +, Co 2 +, Cu 2 +, Fe 2 +, Mn 2 +, ^{+ 2} Sn, Ni ^2+, Zn ^{+ 2,} Al ^{+ 3,} Cr ^{+ 3,} Co ^{+ 3,} Cu ^{+ 3,} Ga ^{+ 3,} Au ^{3 +,} Fe ^{+ 3,} Mn ^{+ 3,} Ni ^{+ 3,} Mn ^{4 +} , Sn ^{4 +} , Cr ^{6 +} and Mn ^{7 +} , but is not limited thereto.

The cation may be contained in an amount of 15 to 50 mol% based on the number of moles of anionic groups in the anionic polymer. When the above range is satisfied, it is advantageous in that the formation of the ionic polymer gel is smooth and the diffusion rate of the active ingredient contained therein is increased when the salt is soled by the salt ion. However, It is not limited by the numerical range.

In the present invention, the polyvalent cation is a monovalent ion as a contradictory concept. When monovalent ions are introduced into the water channel, they cause an ion exchange reaction with the polyvalent ions and convert an anionic polymer gel into a sol. Specifically, when the monovalent ions are introduced into the water channel, the anionic polymer chains are destroyed by the ion exchange with the monovalent ions due to the polyvalent cations. As a result, the ionic polymer gel is changed into the sol Thereby increasing the diffusivity of the active ingredient in the channel. The cubic lipid nanostructure according to the present invention having a sol-gel phase-change property according to the above-mentioned ion contains an active ingredient in the inner water channel, thereby stably collecting the active ingredient or releasing the active ingredient And the emission rate can be controlled.

In the cubic lipid nanostructure according to an embodiment of the present invention, the lipid molecule contained in the lipid nanostructure may be a cubic lipid nanostructure including a monoolein cubic phase. Monoolein is a lipid molecule having a packing parameter of about 1 and can self-assemble upon melt-mixing with water to form a stable cubic phase.

The cubic lipid nanostructure according to an embodiment of the present invention includes an active ingredient that is released to the outside of the water channel by being sensitive to salt ions. Here, the salt ion means a monovalent cationic group. Specifically, the monovalent cation (M ⁺ ) may be any one or more selected from the group consisting of an alkali metal ion and an ammonium ion, but is not limited thereto. For example, sodium ion (Na ⁺ ), potassium ion (K ⁺ ), lithium ion (Li ⁺ ) and ammonium ion (NH ₄ ⁺ ) may be used.

When the lipid nanostructure is exposed to a substance having a salt ion, the cross-linking structure may be destroyed by ion exchange of a polyvalent ion and a monovalent ion as described above. The lipid nanostructure is a salt-anion-responsive lipid nanostructure. When the concentration of the salt ion is high, the lipid nanostructure has a property of releasing a large amount of effective components collected in the water channel.

In another aspect, the present invention includes an effective ingredient delivery system comprising the above-mentioned lipid nanostructure. The method of releasing an active ingredient according to the present invention is a method of releasing an active ingredient released from a water channel in response to salt ions by using an anionic polymer gel contained in a water channel and a cubic lipid nanostructure containing an active ingredient Thereby adjusting the amount.

The active ingredient delivery system according to an embodiment of the present invention may contain an active ingredient in the inner aqueous channel of the lipid nanostructure, and the active ingredient may be a physiologically active ingredient. The active ingredient may include, but is not limited to, various physiologically active ingredients such as anticancer agents, antibiotics, antifungal agents, antibacterial agents, antioxidants, whitening agents, antiinflammatory agents and hypoglycemic agents.

The lipid nanostructure may be used as an active ingredient delivery material as it contains an active ingredient in the inner water channel. The active ingredient carrier includes an anionic polymer gel in an inner water channel inside the lipid nanostructure, and the anionic polymer gel is a semi-solid state of the crosslinked structure, thereby lowering the effective component in the water channel, thereby releasing the active ingredient It is possible to stabilize it without providing it. On the other hand, when exposed to salt ions, for example, when applied to the skin, the skin contains a body fluid having a high salt concentration, so that the anionic gel filling the water channel breaks down the crosslinked structure between the chains of the anionic polymer, The active ingredient can be released to the outside of the water channel as the degree of diffusion of the effective ingredient in the water channel is increased and the release rate and the release amount of the active ingredient can be controlled according to the response of the salt ion.

The method of producing a cubic lipid nanostructure according to the present invention comprises the steps of preparing a monoolein nanostructure by adding an anionic polymer aqueous solution to a monoolein solution and then adding a polyvalent cation compound to the monoolein nanostructure To form an anionic polymer gel therein. The step of preparing the monoolein nanostructure is a step of adding the anionic polymer aqueous solution to the monoolein solution, mixing and hydrating the monoolein solution. That is, it forms a cubic monoolein nanostructure, and an anionic polymer can be incorporated in the resulting internal water channel.

Specifically, a method for preparing a cubic lipid nanostructure according to an embodiment of the present invention includes the steps of preparing a monoolein nanostructure by adding an anionic polymer aqueous solution to a monoolein solution.

At this time, the anionic polymer aqueous solution may have an anionic polymer concentration of 0.01 to 30% by weight, specifically 0.1 to 20% by weight, more specifically 1 to 10% by weight. If it is less than the above range, it is difficult to control the release of the active ingredient through the water channel, irrespective of whether it is sensitive to salt ions, because the crosslink density is too low. When the concentration exceeds the above range, The water-soluble polymer is difficult to dissolve in the aqueous phase, and even if dissolved, the viscosity is too high, so that the monoolefin solution may be difficult to hydrate. On the other hand, when the above range is satisfied, it is easy to form a continuous phase gel by polyvalent cations, and when the formed gel is soled by salt ions, the viscosity is too high, It is effective in terms of performance, but it is a non-limiting example and is not limited to the above numerical range.

The monoolein solution may contain monoolein heated at 30 to 70 ° C, preferably melted by heating at 50 to 60 ° C, optionally further containing a solvent such as water can do.

The anionic polymer aqueous solution may contain 20 to 50% by weight, specifically 23 to 40% by weight, more specifically 25 to 37% by weight based on the total weight of the monoolein and the anionic polymer aqueous solution. When the above range is satisfied, the formation of the cubic monoolefin nanostructure is smooth and effective in terms of ease of absorption of the polyvalent ionic compound to be added for crosslinking the anionic polymer. However, It is not limited by the numerical range.

Next, a polyvalent cation compound is added to the prepared monoolein nanostructure to form an anionic polymer gel therein.

The multivalent cation compound may have a multivalent cation concentration of 0.05 to 20% by weight, specifically 0.1 to 10% by weight, more specifically 0.5 to 5% by weight, in a non-limiting manner. If the concentration of the polyvalent cation is less than the above range, it may be difficult to form a crosslinked structure between the anionic polymer chains. If the concentration exceeds the above range, an anionic polymer gel formed due to excessive polyvalent cation is not destroyed by the salt ion And may be undesirable.

The aqueous anionic polymer solution may be mixed with the monoolein solution after warming to a temperature similar to the temperature of the heated monoolein solution before mixing with the monoolein solution. In this case, the anionic polymer aqueous solution may be added to the monoolein solution and mixed therein by slowly adding the anionic polymer aqueous solution to the heated monoolein solution and inducing mixing in the stationary state. From the above method, it is possible to obtain a lipid nanostructure in which an inner water channel is uniformly formed. Specifically, it is preferable that the anionic polymer aqueous solution layer is formed on the upper layer of the monoolefin solution, and the monoolefin solution and the anionic polymer aqueous solution can be mixed from the interface. Further, such mixing is carried out in a closed atmosphere to ensure a stable cubic phase and is effective in terms of adsorption efficiency. The anionic polymer aqueous solution and the monoolein solution may be further cooled after being mixed and allowed to cool to room temperature until the transparent gel phase is obtained.

In the method of manufacturing a lipid nanostructure according to an embodiment of the present invention, the anionic polymer aqueous solution may be a manufacturing method further comprising an active ingredient.

The active ingredient may be a water-soluble substance which flows into the water channel. The active ingredient is a substance in which the lipid nanostructure of the present invention is released in response to a salt ion, and the active ingredient may be a water-soluble physiologically active ingredient.

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 examples.

(Example 1)

A cubic lipid nanostructure containing alginate gelled with a polyvalent ion

The monoolein cubic phase was prepared by melt hydration. 2 g of monoolein (MO, monoolein, BASF Monomuls 90-018) was placed in a 10 ml vial and melted in a water bath at 50 ° C to prepare a monoolein solution.

Further, the alginate solution was dissolved in distilled water so that the alginate concentration was 1.22 w / v%. Sodium alginate (Sigma Alrich) was used as the alginate. 0.7 ml of the aqueous alginate solution was preheated in a water bath at 50 ° C and slowly added to the prepared monoolein solution. The alginate aqueous solution was completely absorbed into the monoolein solution until it became a transparent semi-solid, Lt; 0 > C).

0.158 ml of calcium chloride (Ca ^{2 +} ) solution (542 mM) was placed on the cubic lipid nanostructure to gel the alginate contained in the water channel of the cubic lipid nanostructure, and the cap was sealed tightly. Then, (MOCP_1) was prepared by storing in a thermostat (27 ° C) until it was completely absorbed by the liposome (cubic lipid nanostructure containing alginate). The final concentration of alginate in the aqueous solution contained in the cubic lipid nanostructure was 1 w / v%, and the final concentration of calcium chloride was 100 mM.

(Example 2)

A monoolefin cubic lipid nanostructure (MOCP_2) was prepared in the same manner as in Example 1, except that the alginate aqueous solution had an alginate concentration of 2.44 w / v%. The final concentration of alginate in the aqueous solution contained in the cubic lipid nanostructure was 2 w / v%, and the final concentration of calcium chloride was 100 mM.

(Comparative Example 1)

The procedure of Example 1 was repeated except that the alginate-free, monoolefin solution was hydrated with distilled water to prepare a cubic lipid nanostructure, thereby obtaining a monoolein cubic lipid nanostructure (MOCP_3). At this time, the cubic lipid nanostructure had a water content equal to that of the cubic lipid nanostructure according to Example 1.

(Comparative Example 2)

The procedure of Example 1 was repeated except that the alginate-free, monoolefin solution was hydrated with a calcium chloride-free solution of alginate to prepare a cubic lipid nanostructure, thereby obtaining a monoolein cubic Lipid nanostructure (MOCP_4) was prepared. At this time, the cubic lipid nanostructure had a water content equal to that of the cubic lipid nanostructure according to Example 1.

(Example 3)

Phase Transition Temperature of Cubic Lipid Nanostructure by Differential Scanning Calorimetry

The phase transition temperatures of the cubic lipid nanostructures prepared in Examples and Comparative Examples were measured by differential scanning calorimetry. A predetermined amount of the cubic lipid nanostructure prepared in Comparative Examples 1 and 2 and Examples 1 and 2 was placed in an aluminum pan (TA Instruments, Aluminum Pans Hermetic) and sealed with a press (TA Instruments, Tzero Sample Press Kit). Then, the calories were analyzed at a heating rate of 2 ° C / min in a temperature range of 30 ° C to 80 ° C using a differential scanning calorimeter (TA Instruments, DSC Q2000). FIG. 2 shows a differential scanning calorimetric curve of the cubic lipid nanostructure according to Comparative Examples 1 and 2 and Examples 1 and 2, and the phase transition occurs when the nanostructure is heated. In the cubic lipid nanostructure according to Comparative Example 1, an endothermic peak was found at about 60.5 DEG C in the differential scanning calorimetry curve, which is caused by the phase transition of the cubic lipid nanostructure. The cubic lipid nanostructure of Comparative Example 2 had a phase transition temperature of about 46.8 DEG C in a differential scanning calorimetry curve. This was lower than that of Comparative Example 1, and the glycerol residue of the monoolein was dehydrated by the CaCl ₂ ion, indicating a lower phase transition temperature. The lipid nanostructures according to Examples 1 and 2 were found to have phase transition temperatures of 44.2 ° C and 45.3 ° C, respectively. They have similar phase transition temperatures, confirming that alginate does not affect the structure of the monoolein cubic lipid nanostructure.

(Example 4)

Observation of structure of cubic lipid nanostructure using transmission electron microscope (TEM)

TEM images were taken with a transmission electron microscope (LEO 912AB OMEGA, Germany). 5 ml of an aqueous solution of Pluronic F-127 (0.6% (w / v)) was added to each cubic lipid nanostructure contained in a 10 ml vial and the cubic lipid nanostructure was pulverized with a spatula into small pieces, To < RTI ID = 0.0 > micronize < / RTI > 1 ml (2% (w / v)) of aqueous solution of tungstophosphoric acid was added to 1 ml of the atomized cubic lipid nanostructure suspension, and the mixture was stained at room temperature (25 ° C) for 4 hours in a dark condition. A grid coated with Formvar / copper was immersed in a suspension of the dyed cubic lipid nanostructure and taken out after a few seconds, and then the grid was dried at room temperature in a dark, dust-free state.

FIGS. 3 and 4 are transmission electron micrographs of the cubic lipid nanostructure prepared in Comparative Example 1 and the cubic lipid nanostructure prepared in Example 2, respectively, when heated at 25 ° C. to 80 ° C. FIG. Black and white streaks were observed and no significant structural differences were found between the nanostructures. Tungstophosphoric acid is easily solubilized in water and is present in the inner channel of the nanostructure, but is almost absent in the lipid bilayer. The water channel corresponds to the white line in the photograph and the lipid bilayer corresponds to the black line. The diameter of the dead channel was about 3-4 nm in electron micrographs and no significant structural differences were found between cubic lipid nanostructures. The phase transition temperature of the cubic lipid nanostructure was significantly reduced by CaCl ₂ , but the phase transition temperature was still high (FIG. 2). Thus, the cubic lipid nanostructure was able to maintain its structure in the preparation of samples for transmission electron microscopy (performed at room temperature (25 ° C)). The alginate is soluble in water and may be present in the water channel without interaction with the hydrocarbon chain of the monoolein. In addition, since the molecule does not cause intermolecular interactions (for example, electrostatic interaction and hydrogen bonding) with the glycerol residue of the monoolein, the alginate hardly affects the packing state of the monoolein, and the structure of the cubic lipid nanostructure It can be understood that there is no influence on

(Example 5)

Salt-ion responsive emission observation

In order to conduct the salt ion responsive emission experiment, a dye - loaded cubic lipid nanostructure was prepared. At this time, amaranth (dye) was dissolved in the aqueous solution so that the final concentration of the dye in the aqueous solution contained in the lipid nanostructure was 0.2% (w / v).

The cubic lipid nanostructures prepared according to Examples and Comparative Examples were placed in 10 ml vials and 5 ml of distilled water or phosphate buffer (PBS) (10 mM, pH 7.4) was added to each vial, And shaken by hand using a shaking incubator (JSR, JSSI-100C Compact shaking incubator) at room temperature (25 캜) in a dark place. To determine the amount of dye (amaranth) released from the cubic lipid nanostructure, 0.2 ml of the release medium is taken from the vial at the specified time for 24 hours and the same amount of fresh release medium is added to the vial to constantly maintain the volume of the release medium Respectively. The release medium (0.2 ml) taken from the vial was diluted to 1 ml with the new release medium and the absorbance of the amaranth was measured at 520 nm with a UV spectrophotometer (JENWAY, 6505 UV / VIS spectrometer). The amount of released amaranth was determined using a calibration curve obtained by plotting Amaranth 0 concentration for absorbance at 550 nm. The release rate was defined as the percentage of the amount of dye released over a specific time period relative to the amount of dye loaded on the nanostructure.

5 is a graph showing the degree of release of the dye when the cubic lipid nanostructure according to Comparative Example 1 was contacted with distilled water (占) and PBS (10 mM, pH 7.4) (?), Was initially released at a very high rate of release and was released for the most 2 hours. This occurred regardless of the release medium of distilled water or PBS (10 mM, pH 7.4). The release rates of distilled water and PBS at 24 hours were 23.7% and 25.9%, respectively.

6 shows the degree of dye release of the cubic lipid nanostructure according to Example 1. The release rate of the dye was steadily increased. The nanostructure according to Example 1 contained an alginate chain crosslinked with a polyvalent ion (Ca ^{2 +} ), and the diffusion rate of the dye through the cubic lipid nanostructure was slowed down. That is, it showed sustained sustained release after the initial rapid release. Also, the release rate was higher when PBS was used as the release medium. Specifically, the maximum release rate when PBS was used as the release medium was 42.6%, and the maximum release rate when using distilled water as the release medium was 31.8%. This shows that the nanostructure has a response characteristic depending on the salt ion, and it is confirmed that the release characteristic is effectively controlled. That is, as the polyvalent cationic crosslinking agent (i.e., Ca ²⁺ ) is exchanged with a monovalent cation (i.e., Na ⁺ ) contained in the buffer solution, the release of gelled alginate in the water channel dissolves or changes into a sol form, The density can be reduced and the dye can be continuously and easily released through the water channel.

FIG. 7 shows the degree of dye release of the cubic lipid nanostructure according to Example 2. Compared with FIG. 6, the initial rapid release behavior was lower than that of FIG. 6, and the sustained release characteristic was exhibited. This is due to the gelated alginate contained in the water channel. As the difference in alginate concentration between the alginate and the alginate is different from that in Example 1, there is also a difference in the cross-link density of the gelled alginate. As a result, And the release rate was also low. Specifically, the maximum release rate in PBS was 37.2%, which was 27% higher than the maximum release rate in distilled water. This is due to the fact that the cubic lipid nanostructure containing gelled alginate due to ion exchange between Ca ^{2 +} and Na ⁺ promotes release in response to salt ions.

(Example 6)

Stability of gelated alginate in salt water

An alginate aqueous solution was prepared by adding alginate to the distilled water to a concentration of 2% (w / v). CaCl ₂ was added to distilled water to prepare a calcium solution having a concentration of 200 mM. 2 ml of each solution was placed in a 10 ml vial and vortexed for several seconds with a vortex mixer to prepare a gelled alginate suspension. The same volume (4 ml) of distilled water or PBS (10 mM, pH 7.4) was added to the gelled alginate suspension and vortexed for 10 seconds to prepare a mixture. The optical density of the prepared mixture at 600 nm was measured at a given time for 24 hours. The relative optical density was calculated as a percentage of the optical density at a given time for the initial optical density.

Figure 8 shows the relative optical density over time of distilled water and gelled alginate in PBS. When the gelled alginate was in distilled water, the optical density hardly changed for 24 hours, indicating that the gelled alginate was stable in distilled water . When the gelated alginate was in an aqueous salt solution (PBS (10 mM, pH 7.4)), the optical density decreased significantly with the lapse of time due to the dissolution of the gelled alginate. This is because Na ⁺ , a monovalent cation of PBS, can replace Ca ^{2 +} bound to the alginate chain, thereby decreasing the cross-linking density or increasing the solubility of the gelated alginate. 6 and 7, respectively.

As a result, the anionic polymer and the anionic polymer gel formed by cross-linking by the polyvalent cation to the anionic polymer in the inner water channel had a large number of cubic lipid nanostructures in the presence of salt ions It can be expected to expand the application range of the active ingredient as a carrier since it has the property of releasing the active ingredient in positive or exhibiting the property of sustained release and stably collecting the active ingredient in the absence of salt ion .

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the invention is not limited to the disclosed exemplary embodiments.

Claims (12)

A salt-ion responsive cubic lipid nanostructure comprising an anionic polymer gel that is a cross-link of an anionic polymer and a polyvalent cation within a water channel. The method according to claim 1,
Wherein the cubic lipid nanostructure comprises a monoolein cubic phase. The method according to claim 1,
The multivalent cations are ^{Mg 2 +, Ca 2 +,} Sr 2 +, Ba 2 +, Be 2 +, Cr 2 +, Co 2 +, Cu 2 +, Fe 2 +, Mn 2 +, Sn 2 +, Ni 2 ^+, Zn ^{+ 2,} Al ^{+ 3,} Cr ^3+, Co ^{+ 3,} Cu ^{+ 3,} Ga ^{+ 3,} Au ^{3 +,} Fe ^{+ 3,} Mn ^{+ 3,} Ni ^{+ 3,} Mn ^{4 +,} Sn ^{4 +,} Cr & lt ^{; 6 +} & gt ^; and Mn < ^{7 +} & gt ^; , wherein the cation-responsive cubic lipid nanostructure comprises at least one cation selected from the group consisting of Cr & lt ^{; 6 +} > The method according to claim 1,
The anionic polymer includes a carboxyl group (-COOH), a sulfonic acid group (-SO ₃ H), a hydroxyl group (-OH), a phosphonic group (-HPO ₂ H), a phosphonic group (-PO ₃ H ₂ ) Sonic group (-AsO ₃ H ₂₎ and cells Reno nikgi (-SeO ₃ H) salt containing one or more functional groups which is selected from the group consisting of ionic responsive cubic lipid nanostructure. The method according to claim 1,
Wherein the anionic polymer has a weight average molecular weight of 400 to 100,000 g / mol. The method according to claim 1,
Wherein the lipid nanostructure comprises an anionic polymer in an amount of 2 to 9 parts by weight based on 100 parts by weight of the lipid molecules, and the salt-ion responsive cubic lipid nanostructure. The method according to claim 1,
The cation-responsive cubic lipid nanostructure comprises 15 to 50 mol% of the cation based on the molar number of anionic groups in the anionic polymer. The method according to claim 1,
Wherein the cubic lipid nanostructure comprises an active ingredient that is released to the outside of the water channel in response to salt ions. 9. An effective ingredient delivery system comprising any one of the cubic lipid nanostructures selected from the group consisting of the above-mentioned materials. Characterized in that the amount of the active ingredient released from the water channel is controlled by the response of the salt ion by using the anionic polymer gel contained in the water channel and the cubic lipid nanostructure containing the active ingredient, A method of releasing an effective substance. Adding an anionic polymer aqueous solution to a monoolein solution to prepare a monoolein nanostructure; and
Adding a polyvalent cation compound to the prepared monoolein nanostructure to form an anionic polymer gel therein
/ RTI > The method of claim 1, 12. The method of claim 11,
Wherein the anionic polymer aqueous solution further comprises an active ingredient.

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