Lauric acid-diacetone alcohol liposome and preparation method thereof

Abstract

The invention discloses a lauric acid-diacetone alcohol liposome and a preparation method thereof, and belongs to the technical field of liposome preparation. The preparation method of the lauric acid-diacetone alcohol lipidosome comprises the following steps: adding a lipidosome membrane material into an organic solvent, dissolving, and injecting into a buffer solution under stirring to obtain a lipidosome suspension; and heating the liposome suspension to react, removing the organic solvent, and mixing with lauric acid-diacetone alcohol to obtain the lauric acid-diacetone alcohol liposome. The lauric acid-diacetone alcohol liposome disclosed by the invention is a material for encapsulating the lauric acid-diacetone alcohol, can protect the lauric acid-diacetone alcohol from being influenced by external factors, and remarkably improves the solubility and the storage stability of the lauric acid-diacetone alcohol.

Description

Technical Field

[0001] The invention relates to the technical field of liposome preparation, in particular to a lauric acid-diacetone alcohol liposome and a preparation method thereof. Background Art

[0002] Lauric acid·diacetone alcohol can destroy certain functions of bacteria and viruses and inhibit the infection of bacteria and viruses. Therefore, it can be applied to the fields of food, medicine, cosmetics, agriculture and chemical industry, and lauric acid·diacetone alcohol has a good synergistic effect on cornfield herbicides, insecticides and fungicides. However, lauric acid·diacetone alcohol is sensitive to temperature, and there are problems such as being easily affected by the external environment, prone to chemical deterioration or decomposition, and poor stability, which makes lauric acid·diacetone alcohol have a short half-life and low bioavailability, limiting the performance of lauric acid·diacetone alcohol. The prior art can improve the stability of lauric acid·diacetone alcohol and control its oxidation process to a certain extent through molecular modification, addition of antioxidants and improvement of packaging materials, but there are still problems such as poor solubility and oxidative deterioration. Therefore, it is urgent to develop new dosage forms to improve the application effect of lauric acid·diacetone alcohol. Summary of the invention

[0003] The purpose of the present invention is to provide a lauric acid·diacetone alcohol liposome and a preparation method thereof to solve the problems existing in the above-mentioned prior art. The method of the present invention can encapsulate lauric acid·diacetone alcohol, protect lauric acid·diacetone alcohol from the influence of the external environment, and improve the stability of lauric acid·diacetone alcohol.

[0004] To achieve the above object, the present invention provides the following solutions:

[0005] One of the technical solutions of the present invention: a method for preparing lauric acid diacetone alcohol liposomes, comprising the following steps:

[0006] The liposome membrane material is added into an organic solvent to dissolve, and then injected into a buffer solution under stirring to obtain a liposome suspension;

[0007] The liposome suspension is heated for reaction, and then the organic solvent is removed, and then the suspension is mixed with lauric acid·diacetone alcohol to obtain the lauric acid·diacetone alcohol liposome.

[0008] Furthermore, the liposome membrane material includes soybean lecithin, cholesterol and Tween 80.

[0009] Furthermore, the mass ratio of soybean lecithin to cholesterol is (2-10):1;

[0010] The mass ratio of soybean lecithin to Tween 80 is 8:(2-6);

[0011] The mass ratio of the soybean lecithin to lauric acid diacetone alcohol is (1-5):1.

[0012] Furthermore, the dosage ratio of the soybean lecithin to the buffer solution is 60-100 mg: 5-25 mL.

[0013] Furthermore, the dissolution includes ultrasonic dissolution; the time of the ultrasonic dissolution is 5 to 30 minutes.

[0014] Furthermore, the method of removing the organic solvent includes vacuuming; the vacuuming time is 45 to 100 minutes.

[0015] Furthermore, the heating reaction is carried out at a temperature of 45 to 65° C. and for a time of 30 to 120 minutes.

[0016] The soybean lecithin used in the preparation of lauric acid diacetone alcohol liposomes of the present invention can improve the morphology of the liposomes, so that the morphology is uniform in size, round or elliptical without deformation or aggregation, thereby improving the structural stability of the liposomes and the transferability of the active ingredients of the drugs; cholesterol can regulate the fluidity of the liposome membrane, so that the liposome membrane maintains a certain flexibility, and enhances the ability of the liposome vesicles to resist changes in external conditions, thereby reducing drug leakage and improving the encapsulation rate of the liposomes; Tween 80 is a non-ionic surfactant that can help lipid components to be better mixed and dispersed in an organic solvent to form a stable liposome solution. In addition, Tween 80 can prevent the liposomes from aggregating or precipitating during storage or use, so that the active ingredients of the drugs are dissolved in the liposomes, thereby increasing the stability of the liposomes and improving the bioavailability and therapeutic effect of the drugs.

[0017] The second technical solution of the present invention is: a lauric acid-diacetone alcohol liposome prepared by the above preparation method.

[0018] The third technical solution of the present invention: an application of the above-mentioned lauric acid-diacetone alcohol liposome in the fields of food, medicine, cosmetics, agriculture or chemical industry.

[0019] The present invention discloses the following technical effects:

[0020] (1) The lauric acid·diacetone alcohol liposome of the present invention is a material that encapsulates lauric acid·diacetone alcohol, which can protect lauric acid·diacetone alcohol from the influence of external factors, significantly improve the solubility and storage stability of lauric acid·diacetone alcohol, solve the problem that lauric acid·diacetone alcohol is a fat-soluble drug and has a low encapsulation rate and poor storage stability when encapsulated in liposomes, and prolong the shelf life and activity maintenance time of lauric acid·diacetone alcohol.

[0021] (2) The present invention prepares lauric acid·diacetone alcohol liposomes by an ethanol injection method, which significantly improves the drug loading capacity of the liposomes and the encapsulation rate of lauric acid·diacetone alcohol.

[0022] (3) The materials used in the present invention are highly safe, inexpensive, and easily available on the market, which is conducive to the efficient use of lauric acid diacetone alcohol in the fields of food, medicine, cosmetics, agriculture, and chemical industry. In addition, the preparation method of the present invention is simple and easy to operate, and is expected to be used for the commercial production of lauric acid diacetone alcohol liposomes. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0024] Figure 1 The figure is a schematic diagram of the preparation process of the lauric acid-diacetone alcohol liposome of the present invention;

[0025] Figure 2 This is a particle size distribution diagram of lauric acid·diacetone alcohol liposomes prepared in Example 6 of the present invention;

[0026] Figure 3 This is the Zeta potential distribution diagram of the lauric acid·diacetone alcohol liposomes prepared in Example 6 of the present invention;

[0027] Figure 4 This is a Fourier infrared spectrum of the lauric acid·diacetone alcohol liposomes prepared in Example 6 of the present invention;

[0028] Figure 5 This is a TEM image of lauric acid·diacetone alcohol liposomes prepared in Example 6 of the present invention;

[0029] Figure 6 This is a TEM image of lauric acid·diacetone alcohol liposomes prepared in Example 6 of the present invention;

[0030] Figure 7 This is the in vitro cumulative release rate of the lauric acid·diacetone alcohol nanoliposomes prepared in Example 6 of the present invention. DETAILED DESCRIPTION

[0031] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0032] It should be understood that the terms described in the present invention are only for describing special embodiments and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper and lower limits of the scope is also specifically disclosed. Each smaller range between the intermediate value in any stated value or stated range and any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0033] Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

[0034] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments of the present invention description without departing from the scope or spirit of the present invention. Other embodiments derived from the present invention description will be apparent to the skilled artisan. The present invention description and examples are exemplary only.

[0035] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0036] The “parts” described in the following examples are all “parts by weight”.

[0037] The lauric acid diacetone alcohol used in the present invention is prepared by first hydrolyzing commercial coconut oil to separate the lauric acid diacetone alcohol, then subjecting the commercial coconut oil to an esterification reaction with glycerol, and finally extracting the lauric acid diacetone alcohol through separation and purification steps. The specific steps are as follows:

[0038] Step 1: Hydrolyze coconut oil

[0039] First, 100 mL of coconut oil was measured and placed in a beaker, followed by the addition of 9.2 g of 30 wt.% sodium hydroxide solution as a catalyst. The beaker was placed in a constant temperature water bath, the temperature was controlled at 100-120°C, and the heating was continued for about 2-3 hours to ensure that the coconut oil was completely hydrolyzed.

[0040] Step 2: Separation of lauric acid and diacetone alcohol

[0041] After hydrolysis, the reaction mixture was allowed to cool naturally to room temperature, and then a separatory funnel was used to separate the upper layer of oil from the lower layer of glycerol and water. The oil phase was then extracted with ethanol, which dissolved lauric acid and diacetone alcohol and obtained a crude extract.

[0042] Step 3: Esterification

[0043] The extracted lauric acid and diacetone alcohol were mixed with glycerol in a molar ratio of 1:3. 2 mL of sulfuric acid was added to the mixture as a catalyst for the esterification reaction. The mixture was then heated at 90-110°C for 6 hours.

[0044] Step 4: Purification of the product

[0045] First, the acidic substances in the reaction mixture are neutralized with sodium carbonate solution, and then the product is washed with water to remove unreacted raw materials and possible by-products. Finally, the washed product is dried to obtain pure lauric acid diacetone alcohol.

[0046] Example 1

[0047] A method for preparing lauric acid diacetone alcohol liposomes (ethanol injection method):

[0048] (1) Weigh 80 mg of soybean lecithin, cholesterol (the amount of cholesterol is 40 mg, 20 mg, 13.3 mg, 10 mg or 8 mg respectively), and 40 mg of Tween 80 and dissolve them in 30 mL of anhydrous ethanol. Place the mixture in a beaker and perform ultrasonic cleaning for 20 min in an ultrasonic cleaning apparatus to completely dissolve the soybean lecithin, cholesterol, and Tween 80 and mix them thoroughly to obtain a liposome membrane material solution.

[0049] (2) Use a sterile syringe to draw up the liposome membrane material solution and slowly inject it into 15 mL of phosphate buffer solution under constant stirring to obtain a blank liposome suspension.

[0050] (3) The blank liposome suspension was heated at normal pressure in a constant temperature water bath at 55°C for 120 min. After the reaction, the anhydrous ethanol was removed by vacuum for 60 min. The suspension was then mixed with 20 mg of lauric acid·diacetone alcohol (manually stirred and mixed), and filtered through a 0.22 μm microporous organic filter membrane. The filtrate was lauric acid·diacetone alcohol liposomes. The schematic diagram of the preparation process is shown in Figure 1 .

[0051] The lauric acid·diacetone alcohol encapsulation efficiency of the lauric acid·diacetone alcohol liposomes prepared in this example was determined by the following method:

[0052] The content of lauric acid·diacetone alcohol in liposomes was measured by ultraviolet spectrophotometry. 10 mL of lauric acid·diacetone alcohol solution of a certain concentration was prepared with anhydrous ethanol, and the absorption wavelength of lauric acid·diacetone alcohol was scanned in the range of 200-400 nm using an ultraviolet spectrophotometer. 3 mg of lauric acid·diacetone alcohol was accurately weighed and diluted with anhydrous ethanol into a series of standard solutions with concentration gradients of 25, 50, 75, 100 and 125 μg / mL, respectively, with anhydrous ethanol as the blank control. It was found that 246 nm was the maximum absorption wavelength of lauric acid·diacetone alcohol. The absorbance values ​​of lauric acid·diacetone alcohol-anhydrous ethanol solutions at different concentrations were measured at 246 nm by an ultraviolet spectrophotometer. A linear regression was performed with the mass concentration of lauric acid·diacetone alcohol as the abscissa (X-axis) and the absorbance value as the ordinate (Y-axis). The regression equation of the standard curve was obtained: Y=9.8554X+0.0115, R 2 =0.9992, indicating that the lauric acid·diacetone alcohol solution has a good linear relationship in the mass concentration range of 25-125 μg / mL.

[0053] 1 mL of the lauric acid·diacetone alcohol liposome prepared in this example was accurately measured, 2 mL of petroleum ether was added, and the mixture was centrifuged at 3000 rpm / min for 5 min to extract free lauric acid·diacetone alcohol, and the petroleum ether and liposome were separated. The same operation was repeated 5 times, and the petroleum ether extract was diluted to 10 mL with anhydrous ethanol. The absorbance was measured at the optimum wavelength of 246 nm, and the absorbance was substituted into the standard curve to determine the free coconut oil content (W1). The encapsulation efficiency was calculated according to the following formula:

[0054]

[0055] Where:

[0056] EE is the encapsulation efficiency of lauric acid·diacetone alcohol, %;

[0057] W1 is the amount of free lauric acid diacetone alcohol in the product, mg;

[0058] W0 is the feed amount of lauric acid diacetone alcohol when preparing the product, mg.

[0059] The results of the determination of the encapsulation efficiency of lauric acid·diacetone alcohol are shown in Table 1.

[0060] Table 1 Effect of the mass ratio of soybean lecithin to cholesterol on the encapsulation efficiency of lauric acid·diacetone alcohol

[0061] Mass ratio of soybean lecithin and cholesterol Encapsulation efficiency %(EE) 2∶1 88.92 4∶1 91.41 6∶1 91.89 8∶1 92.24 10∶1 88.28

[0062] Example 2

[0063] A method for preparing lauric acid diacetone alcohol liposomes:

[0064] (1) Weigh 80 mg of soybean lecithin, 10 mg of cholesterol, and Tween 80 (the amount of Tween 80 is 20 mg, 30 mg, 40 mg, 50 mg, or 60 mg, respectively) and dissolve them in 30 mL of anhydrous ethanol. Place the mixture in a beaker and perform ultrasonic cleaning for 20 min in an ultrasonic cleaning apparatus to completely dissolve the soybean lecithin, cholesterol, and Tween 80 and mix them thoroughly to obtain a liposome membrane material solution.

[0065] (2) Use a sterile syringe to draw up the liposome membrane material solution and slowly inject it into 15 mL of phosphate buffer solution under constant stirring to obtain a blank liposome suspension.

[0066] (3) The blank liposome suspension was heated at normal pressure in a constant temperature water bath at 55°C for 120 min. After the reaction, the ethanol was removed by vacuum for 60 min. The suspension was then mixed with 20 mg of lauric acid / diacetone alcohol (manually stirred and mixed), and filtered through a 0.22 μm microporous organic filter membrane. The filtrate was the lauric acid / diacetone alcohol liposome.

[0067] The results of the determination of the encapsulation efficiency of lauric acid·diacetone alcohol are shown in Table 2.

[0068] Table 2 Effect of the mass ratio of soybean lecithin to Tween 80 on the encapsulation efficiency of lauric acid·diacetone alcohol

[0069] Mass ratio of soybean lecithin and Tween 80 Encapsulation efficiency %(EE) 4∶1 82.45 8∶3 91.71 2∶1 92.77 8∶5 92.15 4∶3 89.26

[0070] As can be seen from Table 2, with the increase in the proportion of Tween 80 input, the liposome encapsulation efficiency showed a trend of first increasing and then decreasing. When the mass ratio of soybean lecithin to Tween 80 was 2:1, the encapsulation efficiency (EE=92.77%) was the highest.

[0071] Example 3

[0072] A method for preparing lauric acid diacetone alcohol liposomes:

[0073] (1) Weigh 80 mg of soybean lecithin, 10 mg of cholesterol, and 40 mg of Tween 80, dissolve them in 30 mL of anhydrous ethanol, place them in a beaker, and perform ultrasonic cleaning in an ultrasonic cleaning instrument for 20 min to completely dissolve the soybean lecithin, cholesterol, and Tween 80 and fully mix them to obtain a liposome membrane material solution.

[0074] (2) The liposome membrane material solution was drawn up with a sterile syringe and slowly injected into a phosphate buffer solution (the amount of phosphate buffer solution was 5 mL, 10 mL, 15 mL, 20 mL or 25 mL) under continuous stirring to obtain a blank liposome suspension.

[0075] (3) The blank liposome suspension was heated at normal pressure in a constant temperature water bath at 55°C for 120 min. After the reaction, the ethanol was removed by vacuum for 60 min. The suspension was then mixed with 20 mg of lauric acid / diacetone alcohol (manually stirred and mixed), and filtered through a 0.22 μm microporous organic filter membrane. The filtrate was the lauric acid / diacetone alcohol liposome.

[0076] The results of the determination of the encapsulation efficiency of lauric acid·diacetone alcohol are shown in Table 3.

[0077] Table 3 Effect of the amount of phosphate buffer on the encapsulation efficiency of lauric acid·diacetone alcohol

[0078] Amount of phosphate buffer (mL) Encapsulation efficiency %(EE) 5 89.26 10 92.33 15 93.04 20 90.97 25 82.31

[0079] As can be seen from Table 3, as the volume of phosphate buffer increases, the encapsulation efficiency of liposomes increases from 89.26% to 93.04% and then decreases to 82.31%. This shows that when the volume of phosphate buffer is small, the solvent concentration is too high, and lauric acid·diacetone alcohol cannot be completely dissolved in the liposomes, resulting in a decrease in the encapsulation efficiency; and when the volume of phosphate buffer is too large, the concentration of lauric acid·diacetone alcohol in the solution is too low, which also affects the encapsulation efficiency. The appropriate volume of phosphate buffer can promote the effective arrangement of the phospholipid bilayer of the liposome, increase the contact area between the liposome and lauric acid·diacetone alcohol, and thus improve the encapsulation efficiency. When the volume of phosphate buffer is 15mL, the encapsulation efficiency % (EE=93.04%) reaches the highest.

[0080] Example 4

[0081] A method for preparing lauric acid diacetone alcohol liposomes:

[0082] (1) Weigh 80 mg of soybean lecithin, 10 mg of cholesterol, and 40 mg of Tween 80, dissolve them in 30 mL of anhydrous ethanol, place them in a beaker, and perform ultrasonic cleaning in an ultrasonic cleaning instrument for 20 min to completely dissolve the soybean lecithin, cholesterol, and Tween 80 and fully mix them to obtain a liposome membrane material solution.

[0083] (2) Use a sterile syringe to draw up the liposome membrane material solution and slowly inject it into 15 mL of phosphate buffer solution under constant stirring to obtain a blank liposome suspension.

[0084] (3) The blank liposome suspension was heated at normal pressure in a constant temperature water bath at 55°C for 120 min. After the reaction, the ethanol was removed by vacuum for 60 min. The suspension was then mixed with lauric acid·diacetone alcohol (the amount of lauric acid·diacetone alcohol was 80 mg, 40 mg, 26.67 mg, 20 mg or 16 mg, respectively) (manually stirred and mixed), and filtered through a 0.22 μm microporous organic filter membrane. The filtrate was the lauric acid·diacetone alcohol liposome. The results of the determination of the lauric acid·diacetone alcohol encapsulation efficiency are shown in Table 4.

[0085] Table 4 Effect of the mass ratio of soybean lecithin to lauric acid·diacetone alcohol on the encapsulation efficiency of lauric acid·diacetone alcohol

[0086] Mass ratio of soybean lecithin and lauric acid diacetone alcohol Encapsulation efficiency %(EE) 1∶1 83.5 2∶1 86.5 3∶1 90.6 4∶1 92.50 5∶1 88.35

[0087] As can be seen from Table 4, as the mass ratio of soybean lecithin to lauric acid·diacetone alcohol increases, the liposome encapsulation efficiency increases from 83.5% to 92.50% and then decreases to 88.35%. When the mass ratio of soybean lecithin to lauric acid·diacetone alcohol is 4:1, the encapsulation efficiency % (EE=92.50%) reaches the highest.

[0088] Example 5

[0089] A method for preparing lauric acid diacetone alcohol liposomes:

[0090] (1) Weigh 80 mg of soybean lecithin, 10 mg of cholesterol, and 40 mg of Tween 80, dissolve them in 30 mL of anhydrous ethanol, place them in a beaker, and perform ultrasonic cleaning in an ultrasonic cleaning instrument for 20 min to completely dissolve the soybean lecithin, cholesterol, and Tween 80 and fully mix them to obtain a liposome membrane material solution.

[0091] (2) Use a sterile syringe to draw up the liposome membrane material solution and slowly inject it into 15 mL of phosphate buffer solution under constant stirring to obtain a blank liposome suspension.

[0092] (3) The blank liposome suspension was heated at normal pressure for 120 min in a constant temperature water bath (the temperature of the constant temperature water bath was 45°C, 50°C, 55°C, 60°C or 65°C, respectively). After the reaction was completed, the anhydrous ethanol was removed by vacuum for 60 min. The suspension was then mixed with 20 mg of lauric acid / diacetone alcohol (manually stirred and mixed), and filtered through a 0.22 μm microporous organic filter membrane. The filtrate was the lauric acid / diacetone alcohol liposome.

[0093] The results of the determination of the encapsulation efficiency of lauric acid·diacetone alcohol are shown in Table 5.

[0094] Table 5 Effect of constant temperature water bath temperature on the encapsulation efficiency of lauric acid diacetone alcohol

[0095] Temperature of constant temperature water bath (℃) Encapsulation efficiency %(EE) 45 86.23 50 91.17 55 92.14 60 90.56 65 87.09

[0096] As can be seen from Table 5, with the increase of preparation temperature, the encapsulation efficiency of liposomes increased from 86.23% to 92.14% and then decreased to 87.09%. This shows that within the appropriate temperature range, with the increase of temperature, the release rate of lauric acid·diacetone alcohol in liposomes is accelerated, thereby improving the stability and uniformity of liposomes; when the preparation temperature is low, the phase change of lecithin cannot be effectively triggered, resulting in poor permeability of liposome membrane, thus affecting its encapsulation efficiency; and when the preparation temperature is too high, the liposome synthesis may be too fast, which increases the fluidity of liposome membrane, leading to structural changes and possibly causing leakage of lauric acid·diacetone alcohol, thereby reducing the encapsulation efficiency. When the preparation temperature is 55°C, the encapsulation efficiency % (EE=92.14%) reaches the highest.

[0097] Example 6

[0098] A method for preparing lauric acid diacetone alcohol liposomes:

[0099] (1) Weigh 80 mg of soybean lecithin, 10 mg of cholesterol, and 40 mg of Tween 80, dissolve them in 30 mL of anhydrous ethanol, place them in a beaker, and perform ultrasonic cleaning in an ultrasonic cleaning instrument for 20 min to completely dissolve the soybean lecithin, cholesterol, and Tween 80 and fully mix them to obtain a liposome membrane material solution.

[0100] (2) Use a sterile syringe to draw up the liposome membrane material solution and slowly inject it into 15 mL of phosphate buffer solution under constant stirring to obtain a blank liposome suspension.

[0101] (3) The blank liposome suspension was heated at normal pressure in a constant temperature water bath at 55°C for 120 min. After the reaction, the ethanol was removed by vacuum for 60 min. The suspension was then mixed with 20 mg of lauric acid-diacetone alcohol (manually stirred and mixed), and filtered through a 0.22 μm microporous organic filter membrane. The filtrate was lauric acid-diacetone alcohol liposomes.

[0102] Effect Example 1

[0103] The particle size, Zeta potential and polydispersity coefficient of the lauric acid·diacetone alcohol liposomes prepared in Example 6 were measured. The specific measurement method is as follows:

[0104] Take an appropriate amount of lauric acid·diacetone alcohol liposomes, dilute them with deionized water, and use a nano-laser particle size analyzer to measure the particle size and polydispersity coefficient. Use a potential analyzer to measure the Zeta potential. The results are shown in Figure 2 and Figure 3 .

[0105] from Figure 2It can be seen that the average particle size of the lauric acid·diacetone alcohol liposomes prepared in Example 6 is 182.7±0.29 nm, and the polydispersity index PDI is 0.386±0.023, indicating that the particle size of the lauric acid·diacetone alcohol liposomes is uniform and evenly distributed.

[0106] from Figure 3 It can be seen that the average Zeta potential of the lauric acid·diacetone alcohol liposomes prepared in Example 6 is -37.5±0.48 mV, indicating that the lauric acid·diacetone alcohol liposomes have good stability.

[0107] Effect Example 2

[0108] The infrared spectrum of the lauric acid·diacetone alcohol liposomes prepared in Example 6 was measured by the following method:

[0109] The lauric acid diacetone alcohol liposomes were mixed with dried potassium bromide powder in a mass ratio of 1:100, dried, ground and pressed into tablets, and heated at 4000-500 cm -1 The results are shown in Figure 4 .

[0110] from Figure 4 It can be seen that the lauric acid diacetone alcohol liposomes have a wavelength of 3200-3500 cm -1 、2700~3050cm -1 There is an absorption peak at 1400~1500cm -1 The absorption peak red-shifted to 1700-1800 cm -1 , confirming that lauric acid·diacetone alcohol was successfully loaded into liposomes.

[0111] Effect Example 3

[0112] The morphological characteristics of the lauric acid·diacetone alcohol liposomes prepared in Example 6 were determined by the following method:

[0113] The lauric acid·diacetone alcohol liposomes prepared in Example 6 were taken and diluted 100 times with deionized water. A trace amount of the diluted solution was taken and dropped onto a copper mesh for transmission electron microscopy. After 10 minutes, the aqueous solution at the bottom of the copper mesh was removed with filter paper and allowed to stand until naturally dried. The mesh was then restained with a 2% phosphotungstic acid solution, and then the phosphotungstic acid staining solution was removed with filter paper. The copper mesh was placed on the filter paper and dried. The morphology of the liposomes on the copper mesh was observed using a transmission electron microscope and photographed. The results are shown in FIG. Figures 5-6 .

[0114] from Figure 5 and Figure 6 It can be seen that the lauric acid·diacetone alcohol liposomes prepared in Example 6 are round or oval in shape, uniform in size and smooth in surface.

[0115] Effect Example 4

[0116] Determine the lauric acid diacetone alcohol liposomes (i.e. Figure 7 The in vitro cumulative release rate of lauric acid diacetone alcohol nanoliposomes) was determined by the following method:

[0117] Appropriate amounts of lauric acid·diacetone alcohol liposomes prepared in Example 6 and lauric acid·diacetone alcohol (as a blank control) were taken respectively, placed in a certain amount of PBS buffer, and subjected to in vitro release determination by dialysis. After a certain period of time, the release solution was removed, and the absorbance of the liposome samples and the blank control at different times was measured. The in vitro cumulative release rate was calculated by the standard curve regression equation (in Example 1). The results are shown in Table 2. Figure 7 .

[0118] from Figure 7 It can be seen that after 8 hours of in vitro release, the cumulative release rate of lauric acid·diacetone alcohol liposomes was over 50%, and the cumulative release rate of lauric acid·diacetone alcohol solution exceeded 75%; after 24 hours of in vitro release, the cumulative release rate of lauric acid·diacetone alcohol solution reached over 90%, and the cumulative release rate of lauric acid·diacetone alcohol liposomes was 76.48%, indicating that lauric acid·diacetone alcohol liposomes have stronger sustained-release properties than lauric acid·diacetone alcohol solution.

[0119] Effect Example 5

[0120] The storage stability of the lauric acid·diacetone alcohol liposomes prepared in Example 6 was determined by the following method:

[0121] The lauric acid·diacetone alcohol liposomes were placed at 4°C and 25°C, and the encapsulation efficiency of the lauric acid·diacetone alcohol liposomes was measured after storage for 0 days, 15 days and 30 days to evaluate the storage stability. The results are shown in Table 6.

[0122] Table 6 Storage stability test results at 4℃ and 25℃

[0123]

[0124] As can be seen from Table 6, during the 30-day storage time, the encapsulation efficiency of the lauric acid·diacetone alcohol liposomes prepared in Example 6 decreased to a certain extent, but the overall stability was good. The liposome encapsulation efficiency at 25°C was lower than that at 4°C, indicating that the lauric acid·diacetone alcohol liposomes were more stable at 4°C than at room temperature. In addition, the lauric acid·diacetone alcohol liposomes prepared in Example 6 of the present invention can be stably stored at a temperature of 0 to 40°C (the suitable storage temperature of ordinary lauric acid·diacetone alcohol is only 2 to 8°C).

[0125] Comparative Example 1

[0126] Preparation method of lauric acid diacetone alcohol liposomes:

[0127] (1) Weigh 80 mg of soybean lecithin, 10 mg of cholesterol, and 40 mg of Tween 80 and dissolve them in 30 mL of anhydrous ethanol. Place the mixture in a test tube and heat it in a constant temperature water bath at 50°C for 2 h to completely dissolve the soybean lecithin, cholesterol, and Tween 80 and mix them thoroughly. Then transfer the thoroughly mixed solution to a 200 mL round-bottom flask and remove the anhydrous ethanol by rotary evaporation under reduced pressure (the rotary evaporation speed is 15 rpm) in a constant temperature water bath at 50°C to form a uniform transparent film on the inner wall of the flask. Transfer the round-bottom flask with the film to a vacuum pump and evacuate for 1 h to remove the residual anhydrous ethanol to obtain a lipid film.

[0128] (2) Add 15 mL of phosphate buffer to the round-bottom flask and continue to rotary evaporate and hydrate the membrane for 30 min to completely elute and dissolve the membrane material to obtain a suspension.

[0129] (3) The round-bottom flask was placed in an ultrasonic cleaner for 10 min to allow the membrane material to be fully mixed and uniformly obtained to obtain a blank liposome suspension.

[0130] (4) The blank liposome suspension was mixed with 20 mg of lauric acid-diacetone alcohol, heated in a constant temperature water bath at 50° C. for 15 min, and filtered through a 0.22 μm microporous organic filter to obtain lauric acid-diacetone alcohol liposomes.

[0131] The encapsulation efficiency of the lauric acid·diacetone alcohol liposome prepared in this comparative example was 88.31%.

[0132] Comparative Example 2

[0133] Same as Example 6, except that cholesterol is replaced by an equal amount of stigmasterol.

[0134] The encapsulation efficiency of the lauric acid·diacetone alcohol liposome prepared in this comparative example was 86.38%.

[0135] The embodiments described above are only descriptions of the preferred modes of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary technicians in this field should all fall within the protection scope determined by the claims of the present invention.

Claims

1. A method for preparing lauric acid-diacetone alcohol liposomes, characterized in that: The following steps are involved: The liposome membrane material is added into an organic solvent to dissolve, and then injected into a buffer solution under stirring to obtain a liposome suspension; The liposome suspension is heated for reaction, and then the organic solvent is removed, and then the suspension is mixed with lauric acid·diacetone alcohol to obtain the lauric acid·diacetone alcohol liposome.

2. The preparation method according to claim 1, characterized in that: The liposome membrane material comprises soybean lecithin, cholesterol and Tween 80.

3. The preparation method according to claim 2, characterized in that: The mass ratio of soybean lecithin to cholesterol is (2-10):1; The mass ratio of soybean lecithin to Tween 80 is 8:(2-6); The mass ratio of the soybean lecithin to lauric acid diacetone alcohol is (1-5):

1.

4. The preparation method according to claim 2, characterized in that: The dosage ratio of the soybean lecithin to the buffer solution is 60-100 mg: 5-25 mL.

5. The preparation method according to claim 1, characterized in that: The dissolution includes ultrasonic dissolution; the time of the ultrasonic dissolution is 5 to 30 minutes.

6. The preparation method according to claim 1, characterized in that: The method of removing the organic solvent includes vacuuming; the vacuuming time is 45 to 100 minutes.

7. The preparation method according to claim 1, characterized in that: The temperature of the heating reaction is 45-65° C. and the time is 30-120 min.

8. Lauric acid-diacetone alcohol liposome prepared by the preparation method according to any one of claims 1 to 7.

9. Use of the lauric acid-diacetone alcohol liposome according to claim 8 in the fields of food, medicine, cosmetics, agriculture or chemical industry.

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