Preparation of a water-in-oil Pickering emulsion gel based on charge interaction and its application in enzyme-catalyzed sustained release

Abstract

The invention discloses the preparation of a water-in-oil Pickering emulsion gel based on charge interaction and its enzymatic sustained release application. In the method, a positively charged amino polymer is dissolved in water as a water phase, negatively charged hydrophobic nanoparticles are dispersed in oil as an oil phase, and the water phase and the oil phase are mixed to emulsify the water phase in the oil phase. The nanoparticles serve as solid surfactants to stabilize the oil-water interface, and the positively charged amino polymer in the water phase is combined with the negatively charged nanoparticles in the oil phase to form a cross-linked network, so that the droplets are closely arranged and not easy to flow and deform, thereby obtaining a stable Pickering emulsion gel. This Pickering emulsion gel has good stability, can realize the simultaneous encapsulation of hydrophilic and hydrophobic active substances, and can realize the long-term slow release of active substances in the gel in the presence of an enzyme solution, and has a wide range of oil phase selection and strong versatility.

Description

Technical Field

[0001] The invention relates to the field of emulsion gels, and in particular to the preparation of an oil-in-water Pickering emulsion gel based on charge interaction and the application of the same in enzymatic sustained release. Background Art

[0002] Emulsion gel, a soft solid material that combines the advantages of emulsion and gel, is composed of an aqueous phase and an oil phase, has superior thermodynamic stability and adjustable viscoelasticity. Due to its special composition structure and properties, it has many practical applications in controlling gastric and / or intestinal drug release, encapsulating nutrients, improving food texture, and simulating the effects of fat on food hardness and water retention capacity.

[0003] The preparation of emulsion gel generally uses natural biopolymers with good emulsifying activity, gelation and biocompatibility. Traditionally, the preparation of emulsion gel includes the production of emulsion and the conversion of emulsion into gel, using homogenization and then heating, pH or salt induction or cross-linking methods, and the preparation method is complicated and time-consuming. Using solid particles instead of surfactants as stabilizers to stabilize emulsions is one of the effective methods to improve their stability and functional properties. However, most of the emulsion gels prepared at present can only achieve the encapsulation of hydrophilic or hydrophobic substances, and have poor affinity for some active substances. The prepared emulsion gel is not stable enough and is easily affected by external conditions such as temperature and pH, which destroys the gel structure and causes the release of the encapsulated active substances. Therefore, it is of great significance to develop an emulsion gel that is not affected by conditions such as temperature and pH, has high stability, and can withstand a variety of complex external conditions to achieve effective delivery of active substances. Summary of the invention

[0004] The present invention describes the preparation of a water-in-oil Pickering emulsion gel based on charge interaction and its application of enzymatic sustained release. The method dissolves a positively charged amino polymer in water as a water phase, disperses negatively charged hydrophobic nanoparticles in oil as an oil phase, and emulsifies the water phase in the oil phase by simple vortex oscillation or high-energy emulsification. During the emulsification process, the nanoparticles act as solid surfactants to stabilize the oil-water interface, and the positively charged amino polymer in the water phase combines with the negatively charged nanoparticles in the oil phase to form a cross-linked network, and the droplets are closely arranged and not easy to flow and deform, thereby obtaining a stable Pickering emulsion gel. This Pickering emulsion gel has good stability, can realize the simultaneous encapsulation of hydrophilic and hydrophobic active substances, and can realize the long-term slow release of active substances in the gel in the presence of an enzyme solution, has a wide range of oil phase options, and is highly versatile, providing a new system for material transport and release.

[0005] The present invention first discloses a preparation method of a water-in-oil Pickering emulsion gel based on charge interaction and its application of enzymatic sustained release, comprising the following steps:

[0006] (1) dissolving the positively charged amino polymer in water as the aqueous phase and dispersing the negatively charged hydrophobic nanoparticles in oil as the oil phase;

[0007] (2) The aqueous phase and the oil phase are mixed, and the aqueous phase is emulsified in the oil phase to obtain the water-in-oil Pickering emulsion gel. During the emulsification process, the nanoparticles act as solid surfactants to stabilize the oil-water interface. The positively charged amino polymer in the aqueous phase combines with the negatively charged nanoparticles to form a cross-linked network. The droplets are closely arranged and not easily deformed by flow, thereby obtaining a stable Pickering emulsion gel.

[0008] Preferably, in step (1), the positively charged amino polymer added to the aqueous phase is polyoxyethylene diamine, polyethylene imine, polyacrylamine, etc., with a molecular weight of 1000-10000; and the hydrophobic nanoparticles are hydrophobically modified nano-scale silica.

[0009] Preferably, in step (1), the oil used in the water-in-oil Pickering emulsion gel preparation method is any oil phase that can disperse hydrophobic nanoparticles, including: camellia oil, olive oil, soybean oil, corn oil, silk oil, n-hexane, essential oil, iodized oil, polydimethylsiloxane, styrene, n-butyl acrylate, etc.

[0010] Preferably, in step (1), the concentration of the amino polymer in the aqueous phase is 2-10 mg / mL.

[0011] Preferably, in step (1), the concentration of the hydrophobic nanoparticles in the oil phase is 15-30 mg / mL.

[0012] Preferably, in step (2), the volume ratio of the oil-water phase is 4:6 to 6:4.

[0013] Preferably, in the step (2), the emulsification process can be achieved by simple vortex oscillation. In addition, high-energy emulsification methods such as high-speed homogenization, high-pressure homogenization and the like can also be used.

[0014] The present invention also discloses a drug-loaded oil-in-water Pickering emulsion gel prepared based on the above method. The preparation method of the drug-loaded oil-in-water Pickering emulsion gel comprises the following steps:

[0015] (1) dissolving a positively charged amino polymer in water as a water phase, dispersing negatively charged hydrophobic nanoparticles in oil as an oil phase; adding a pharmaceutical active ingredient to the water phase or the oil phase; wherein the hydrophilic pharmaceutical active ingredient is added to the water phase; and the hydrophobic pharmaceutical active ingredient is added to the oil phase;

[0016] (2) The aqueous phase and the oil phase are mixed, and the aqueous phase is emulsified in the oil phase to obtain the water-in-oil Pickering emulsion gel. During the emulsification process, the hydrophobic nanoparticles act as solid surfactants to stabilize the oil-water interface. The positively charged amino polymers in the aqueous phase are combined with the negatively charged nanoparticles in the oil phase to form a cross-linked network, so that the droplets are closely arranged and not easily deformed by flow, thereby obtaining a stable drug-loaded water-in-oil Pickering emulsion gel.

[0017] The present invention further provides the use of the drug-loaded oil-in-water Pickering emulsion gel in the preparation of an enzymatic sustained-release drug. The enzymatic sustained-release drug is a pharmaceutical preparation comprising the drug-loaded oil-in-water Pickering emulsion gel, and the enzymatic sustained-release drug can achieve long-term slow release of the active drug ingredient in the Pickering emulsion gel in the presence of an enzyme (such as lipase).

[0018] Compared with the prior art, the present invention has the following beneficial effects:

[0019] (1) The present invention proposes a method for preparing an oil-in-water Pickering emulsion gel based on charge interaction and its application in enzymatic sustained release. The preparation of the emulsion gel is achieved through a simple preparation method. Nanoparticles serve as solid surfactants to stabilize the oil-water interface. Amino polymers in the aqueous phase combine with the nanoparticles to form a cross-linked network. The droplets are tightly arranged and are not easily deformed by flow, which provides a new idea for the preparation of stable Pickering emulsion gels.

[0020] (2) The water-in-oil Pickering emulsion gel prepared by the present invention has strong stability, and the rheological properties of the gel are basically unchanged at 20°C to 90°C; and the gel structure does not change after centrifugation at a speed of 1200 rpm for 10 minutes.

[0021] (3) The water-in-oil Pickering emulsion gel prepared by the present invention is composed of an aqueous phase and an oil phase, and can simultaneously encapsulate hydrophilic and hydrophobic active substances, and can be widely used in the fields of material transportation.

[0022] (4) The method proposed in the present invention for preparing Pickering emulsion gel by emulsifying the water phase in the oil phase is applicable to a variety of oil-water systems and has strong versatility.

[0023] (5) Based on the preparation method of the Pickering emulsion gel of the present invention, the present invention also prepares a drug-loaded Pickering emulsion gel, wherein the active ingredient is added to the aqueous phase or the oil phase during the preparation process. The obtained drug-loaded Pickering emulsion gel can be directly used as a drug or further prepared as a pharmaceutical preparation, which can achieve long-term slow release of the active substance in the gel in the presence of an enzyme solution (such as lipase). BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 Schematic diagram of the preparation of water-in-oil Pickering emulsion gel;

[0025] Figure 2 Pickering emulsion gels prepared with different oil-water phase volume ratios;

[0026] Figure 3 The figures are actual pictures and laser confocal images of water-in-oil Pickering emulsion gels, wherein (a) is an actual picture of the preparation of water-in-oil Pickering emulsion gel, (b) is a confocal image of the water-in-oil Pickering emulsion gel prepared by dyeing the water phase, (c) is a confocal image of the water-in-oil Pickering emulsion gel prepared by dyeing the oil phase, and (d) is a confocal image of the water-in-oil Pickering emulsion gel prepared by dyeing the water phase and the oil phase together;

[0027] Figure 4 The rheological properties of water-in-oil Pickering emulsion gel;

[0028] Figure 5 is the stability of the water-in-oil Pickering emulsion gel, where (a) is the centrifugal stability and (b) is the temperature stability;

[0029] Figure 6 The stability of water-in-oil Pickering emulsion gel at different pH;

[0030] Figure 7 The release curves of water-in-oil Pickering emulsion gel in the presence or absence of enzyme;

[0031] Figure 8 The invention relates to a Pickering emulsion gel prepared from an aqueous solution of polyethyleneimine and an oily solution containing hydrophobic silica nanoparticles.

[0032] Fig. 9 A physical picture of a Pickering emulsion gel prepared from an aqueous solution containing polyoxyethylene diamine and an oily solution containing silicon dioxide;

[0033] Fig.10A Pickering emulsion prepared from an aqueous solution containing polyoxyethylene diamine and an oily solution not containing hydrophobic silica nanoparticles;

[0034] Fig.11 A Pickering emulsion prepared from an aqueous phase solution containing no polyoxyethylene diamine and an oil phase solution containing hydrophobic silica nanoparticles;

[0035] Fig.12 The Pickering emulsion is prepared by respectively using an aqueous phase solution containing polyacrylic acid and polyethylene glycol and an oil phase solution containing hydrophobic silica nanoparticles. DETAILED DESCRIPTION

[0036] The present invention is further described and illustrated below in conjunction with specific embodiments. The embodiments are merely exemplary of the present disclosure and do not limit the scope.

[0037] Example 1: Preparation of water-in-oil Pickering emulsion gel based on charge interaction

[0038] (1) dissolving polyoxyethylene diamine in water to prepare a polyoxyethylene diamine aqueous solution with a concentration of 10 mg / mL; dispersing hydrophobic silica nanoparticles in camellia oil to prepare a silica / camellia oil oil solution with a concentration of 20 mg / mL, and then vortexing and mixing evenly;

[0039] (2) According to the volume ratio of oil to water of 4:6, weigh the oil phase and water phase respectively and add them into a glass bottle, then vortex until the mixture becomes a white viscous emulsion. After standing for a few minutes, the emulsion forms a gel. If the glass bottle is turned upside down, the emulsion gel will not flow. Figure 1 and attached Figure 2 (a) shown.

[0040] (3) According to the methods in steps (1) and (2), fluorescent dye FITC is added to the aqueous phase to prepare a Pickering emulsion gel, and a laser confocal microscope is used to take a photo of the gel under an excitation light of 488 nm, as shown in the attached Figure 2 (b) as shown.

[0041] (4) According to the methods in steps (1) and (2), the fluorescent dye Nile red is added to the oil phase to prepare a Pickering emulsion gel, and a laser confocal microscope is used to take a photo of the gel under an excitation light of 543 nm, as shown in the attached figure. Figure 2 (c) as shown.

[0042] (5) According to the method in steps (1) and (2), FITC was added to the aqueous phase and Nile red was added to the oil phase to prepare a Pickering emulsion gel, and the gel was photographed under the excitation light of 488 nm and 543 nm using a laser confocal microscope, as shown in the attached figure. Figure 2 (d) as shown.

[0043] (6) According to the method in step (1), an oil-water phase solution is prepared. By adjusting the volume ratio of the oil-water phase, the results show that the preparation of the water-in-oil Pickering emulsion gel can be achieved when the volume ratio of the oil-water phase is 4:6 to 6:4, such as Figure 3 shown.

[0044] Example 2: Determination of properties of water-in-oil Pickering emulsion gel based on charge interaction

[0045] (1) Rheological properties: According to the method in Example 1, a water-in-oil Pickering emulsion gel was prepared with an oil-water phase volume ratio of 4:6, and the rheological properties of the obtained gel were measured using a flat plate system of a rotational rheometer. The strain scan results are shown in the attached figure. Figure 4 As shown in (a), the storage modulus of the gel decreases exponentially when the strain is 10%, and the loss modulus intersects with the storage modulus curve when the strain is 100%, indicating that the obtained oil-in-water Pickering emulsion gel is in a gel state. The frequency scanning results are shown in the attached Figure 4 As shown in (b), when the frequency scanning range is 0.1-50rad / s, the storage modulus of the gel is always greater than the loss modulus, which further proves that the obtained oil-in-water Pickering emulsion gel is in a gel state, and within this frequency scanning range, the storage modulus and loss modulus do not change much with the increase of frequency. The time scanning results are shown in the attached Figure 4 As shown in (c), within the scanning time of 1800s, the storage modulus and loss modulus of the gel did not change much. At 25℃ and 37℃, the storage modulus and loss modulus of the gel measured were not much different, indicating that the effect of temperature on the rheological properties of the gel was small. The effect of temperature on the gel was further determined, and the temperature scanning results are shown in the attached figure. Figure 4 As shown in (d), the storage modulus and loss modulus of the gel remain basically unchanged with increasing temperature, indicating that the obtained oil-in-water Pickering emulsion gel has good stability at 20℃-90℃.

[0046] (2) Centrifugal stability: According to the method in Example 1, a water-in-oil Pickering emulsion gel was prepared with an oil-water volume ratio of 4:6. The prepared gel was poured into a centrifuge tube and centrifuged at 500 rpm, 800 rpm, 1000 rpm, 1200 rpm, 1500 rpm, 1800 rpm, 2000 rpm, 2500 rpm and 3000 rpm for 10 min, and then the emulsion gel was taken out to observe its stability. When the centrifugal speed was less than or equal to 1200 rpm, the water phase and the oil phase in the gel did not separate, and the stability was good. Figure 5 (a) shown.

[0047] (3) Temperature stability: According to the method in Example 1, a water-in-oil Pickering emulsion gel was prepared with an oil-water volume ratio of 4:6. The prepared gel was placed at -20°C, 4°C and 50°C for 12 h, and the stability of the gel was observed after it was taken out. Figure 5 (b) When the temperature is between 4°C and 50°C, the emulsion gel is stable; when the temperature is below freezing, the emulsion gel is destroyed due to the crystallization of water, however, a stable Pickering emulsion gel can be obtained again by shaking.

[0048] (4) pH stability: According to the method in Example 1, a water-in-oil Pickering emulsion gel was prepared with an oil-water volume ratio of 4:6. The prepared gel was placed in solutions with pH = 2, 7 and 12 for 24 h, and the state of the gel and its optical microscope were observed. Figure 6 As shown, the gel has good stability under acidic and neutral conditions, and the gel structure is destroyed under highly alkaline conditions.

[0049] Example 3: Enzymatic sustained release properties of water-in-oil Pickering emulsion gel

[0050] In vitro degradation experiments of water-in-oil Pickering emulsion gels were conducted at 37°C. 0.5 mL of emulsion gel was added to the bottom of a 3 mL glass bottle, and 1 mL of PBS buffer containing or not containing 5 mg / mL lipase was added to the top of the gel. Hydrophilic FITC was encapsulated in the aqueous phase, and then the top buffer was collected every other day to detect the dye concentration, and fresh buffer was replaced after each measurement. The concentration of released FITC was measured using a UV-Vis spectrometer at a peak wavelength of 495 nm. In the presence of the enzyme, the gel was completely released in nearly 40 days, and the resulting release curve is shown in Figure 2. Figure 7 shown.

[0051] Example 4: Pickering emulsion gel prepared based on polyethyleneimine

[0052] (1) dissolving polyethyleneimine in water to prepare a polyethyleneimine aqueous solution with a concentration of 10 mg / mL; dispersing hydrophobic silica nanoparticles in camellia oil to prepare an oil phase solution with a concentration of 20 mg / mL silica / camellia oil, and then vortexing and mixing evenly;

[0053] (2) According to the volume ratio of oil to water of 4:6, weigh the oil phase and water phase respectively and add them into a glass bottle, then vortex until the mixture becomes a white viscous emulsion. After standing for a few minutes, the emulsion forms a gel. If the glass bottle is turned upside down, the emulsion gel will not flow. Figure 8 shown.

[0054] Comparative Example 1: Preparation of water-in-oil Pickering emulsion gel

[0055] (1) dissolving polyoxyethylene diamine in water to prepare a polyoxyethylene diamine aqueous solution with a concentration of 10 mg / mL; dispersing hydrophobic silica nanoparticles in camellia oil to prepare a silica / camellia oil oil solution with a concentration of 20 mg / mL;

[0056] (2) According to the volume ratio of oil to water phase of 4:6, the aqueous phase solution containing polyoxyethylene diamine and the oil phase solution containing hydrophobic silica nanoparticles are mixed, and then vortexed to mix them evenly to obtain a stable Pickering emulsion gel, such as Fig. 9 As shown;

[0057] (3) According to the volume ratio of oil to water phase of 4:6, the aqueous phase solution containing polyoxyethylene diamine was mixed with the oil phase solution without hydrophobic silica nanoparticles, and then vortexed to mix evenly. However, a stable Pickering emulsion gel could not be obtained, but a Pickering emulsion was obtained, as shown in 10;

[0058] (4) According to the volume ratio of oil to water phase of 4:6, the aqueous phase solution without polyoxyethylene diamine was mixed with the oil phase solution containing hydrophobic silica nanoparticles, and then vortexed to mix evenly. However, a stable Pickering emulsion gel could not be obtained, but a Pickering emulsion, such as Fig.11 shown.

[0059] Comparative Example 2: Preparation of water-in-oil Pickering emulsion

[0060] (1) dissolving polyacrylic acid and polyethylene glycol in water respectively to prepare an aqueous phase solution of polyacrylic acid and polyethylene glycol with a concentration of 10 mg / mL; dispersing hydrophobic silica nanoparticles in camellia oil to prepare an oil phase solution of silica / camellia oil with a concentration of 20 mg / mL;

[0061] (2) According to the volume ratio of oil to water of 4:6, the aqueous solutions of polyacrylic acid and polyethylene glycol were mixed with the oil phase solution respectively, and then vortexed to mix evenly. However, no stable Pickering emulsion gel could be obtained. Fig.12 shown.

[0062] By comparing Example 1 with Comparative Example 1, it is found that when the aqueous phase solution of the oil-water system contains only an amino polymer (polyoxyethylene diamine) or the oil phase solution contains only silica, a Pickering emulsion gel cannot be formed. Only when the oil-water system contains both silica and an amino polymer (polyoxyethylene diamine) can a stable Pickering emulsion gel be formed.

[0063] By comparing Example 1 with Comparative Example 2, it is found that when the oil phase of the oil-water system contains silicon dioxide and the water phase contains a polymer solution without amino groups (polyacrylic acid and polyethylene glycol), a Pickering emulsion gel cannot be formed.

[0064] The above examples are only specific embodiments of the present invention. Obviously, the present invention is not limited to the above examples, and many variations are possible. All variations that can be directly derived or associated with the contents disclosed by a person skilled in the art should be considered as the protection scope of the present invention.

Claims

1. A method for preparing a water-in-oil Pickering emulsion gel based on charge interaction, characterized in that: The following steps are involved: (1) dissolving a positively charged amino polymer in water as an aqueous phase, and dispersing negatively charged hydrophobic nanoparticles in oil as an oil phase; the concentration of the positively charged amino polymer in the aqueous phase is 2-10 mg / mL, and the concentration of the negatively charged hydrophobic nanoparticles in the oil phase is 15-30 mg / mL; (2) The aqueous phase and the oil phase are mixed, and the volume ratio of the oil phase to the aqueous phase is 4:6 to 6:4; the aqueous phase is emulsified in the oil phase to obtain the water-in-oil Pickering emulsion gel. During the emulsification process, the hydrophobic nanoparticles act as solid surfactants to stabilize the oil-water interface, and the positively charged amino polymers in the aqueous phase are combined with the negatively charged nanoparticles in the oil phase to form a cross-linked network, so that the droplets are closely arranged and not easy to flow and deform, thereby obtaining a stable Pickering emulsion gel.

2. The method for preparing the water-in-oil Pickering emulsion gel according to claim 1, characterized in that: The process of emulsifying the water phase in the oil phase is achieved by vortex oscillation or high-energy emulsification.

3. The method for preparing the water-in-oil Pickering emulsion gel according to claim 1, characterized in that: In the step (1), the positively charged amino polymer added to the aqueous phase is polyoxyethylene diamine, polyethylene imine or polyacrylamine, and has a molecular weight of 1000-10000; and the hydrophobic nanoparticles are hydrophobically modified nano-scale silica particles.

4. The method for preparing the water-in-oil Pickering emulsion gel according to claim 1, characterized in that: In the step (1), the oil is an oil that can disperse hydrophobic nanoparticles.

5. The method for preparing the water-in-oil Pickering emulsion gel according to claim 4, characterized in that: The oil is a mixture of one or more of camellia oil, olive oil, soybean oil, corn oil, silk oil, n-hexane, essential oil, iodized oil, polydimethylsiloxane, styrene or n-butyl acrylate.

6. A drug-loaded water-in-oil Pickering emulsion gel prepared by the method of claim 1, characterized in that: The preparation method of the drug-loaded water-in-oil Pickering emulsion gel comprises the following steps: (1) dissolving a positively charged amino polymer in water as an aqueous phase, dispersing negatively charged hydrophobic nanoparticles in oil as an oil phase; adding a pharmaceutical active ingredient to the aqueous phase or the oil phase; wherein the hydrophilic pharmaceutical active ingredient is added to the aqueous phase; and the hydrophobic pharmaceutical active ingredient is added to the oil phase; the concentration of the positively charged amino polymer in the aqueous phase is 2-10 mg / mL, and the concentration of the negatively charged hydrophobic nanoparticles in the oil phase is 15-30 mg / mL; (2) The aqueous phase and the oil phase are mixed, and the volume ratio of the oil phase to the aqueous phase is 4:6 to 6:4; the aqueous phase is emulsified in the oil phase to obtain the water-in-oil Pickering emulsion gel. During the emulsification process, the hydrophobic nanoparticles act as solid surfactants to stabilize the oil-water interface, and the positively charged amino polymers in the aqueous phase are combined with the negatively charged nanoparticles in the oil phase to form a cross-linked network, so that the droplets are closely arranged and not easy to flow and deform, thereby obtaining a stable drug-loaded water-in-oil Pickering emulsion gel.

7. Use of the drug-loaded water-in-oil Pickering emulsion gel according to claim 6 in the preparation of enzymatic sustained-release drugs.

8. The use according to claim 7, characterized in that: The enzyme is lipase.

Images