Polypeptide-loaded hippophae rhamnoides fruit oil microcapsule powder as well as preparation method and application thereof

Abstract

The invention discloses polypeptide-loaded hippophae rhamnoides fruit oil microcapsule powder as well as a preparation method and application thereof. The hippophae rhamnoides fruit oil microcapsule powder comprises hippophae rhamnoides fruit oil and polypeptide, and the preparation method comprises the following steps: mixing the hippophae rhamnoides fruit oil and the polypeptide, and performing ball milling in a ball mill to obtain the hippophae rhamnoides fruit oil microcapsule powder According to the invention, a molecular mixture is formed through the action of mechanical force, the dissolution of the sea buckthorn fruit oil is promoted through the effects of increasing the dispersion uniformity of the medicine and the carrier, increasing the wettability and the like, and a compound is formed through the clathration of the carrier, so that the degradation of the sea buckthorn fruit oil and the loss of the antioxidant activity of the sea buckthorn fruit oil are inhibited; meanwhile, the microcapsule powder can be assembled into stable nano microemulsion in water, the taste is fresh and not greasy, the anti-oxidation and free radical scavenging capacity of the sea buckthorn fruit oil is improved, the non-alcoholic fatty liver resisting effect of the sea buckthorn fruit oil is remarkably improved, and the preparation method is simple, easy to implement, low in cost, non-toxic and harmless.

Description

Technical Field

[0001] The invention belongs to the technical field of food processing, and particularly relates to a polypeptide-loaded seabuckthorn fruit oil microcapsule powder and a preparation method and application thereof. Background Art

[0002] As people's living standards improve, life pressure increases, and people do not pay enough attention to lifestyle and eating habits, the number of people with obesity and fatty liver continues to increase. However, existing chemical drugs still cannot treat them well, and long-term medication is required, and the side effects are large. In recent years, the dietary therapy strategy has been highly praised by consumers. There is a huge market prospect for treating obesity and improving health problems such as fatty liver through dietary adjustment. However, most functional foods currently only come from natural plants, are preliminary processed products or extracts, and need to be used in large doses to exert better effects. Moreover, due to the lack of scientific and technological content, it is difficult to gain the favor of consumers. Therefore, there is an urgent need for a high-tech, efficient and safe functional food for the treatment of fatty liver.

[0003] Seabuckthorn is a perennial deciduous shrub of the genus Hippophaerhamnoides of the family Elaeagnaceae. Its Latin name is Hippophaerhamnoides L. and it is a traditional food. Seabuckthorn berry oil is the most abundant functional oil in seabuckthorn. It is rich in a large amount of Omega-9 unsaturated fatty acids and has significant therapeutic value. Scientific research has confirmed that seabuckthorn seed oil can help repair skin damage and treat skin photoaging, such as repairing skin damage caused by radiotherapy, sunlight and cosmetic lasers; it can also be used to treat some skin diseases such as eczema and burns. Oral seabuckthorn berry oil has the effect of significantly reducing alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), and triglycerides (TG) in mice with high cholesterol induced by CCl4 injection. Therefore, seabuckthorn berry oil is a functional food with special therapeutic effects on hyperlipidemia. However, the effective ingredients of seabuckthorn fruit oil are poorly water-soluble and exist in the form of oil droplets in aqueous solution. They have a greasy taste and are easily oxidized and rancid. They have a poor taste and poor efficacy when taken directly, making it difficult to gain consumer recognition, which greatly limits the further application of seabuckthorn fruit oil in food. Therefore, developing seabuckthorn fruit oil products with good water solubility, strong stability, refreshing taste and better efficacy is of practical significance to solve the above problems.

[0004] The main methods for water-dissolving seabuckthorn oil are solvent method and solvent spray drying method. These methods require the use of organic solvents, which have the problem of residual organic solvents, and the preparation process is complicated, and the loss rate of seabuckthorn oil in the process is increased. Therefore, how to avoid the above problems is the biggest obstacle faced in the development of seabuckthorn oil water-soluble products. The mechanochemical method is used to prepare microcapsule powder, and no organic solvent is needed in the process, which can avoid the pollution of organic solvents to the environment during the reaction process, and no heating is required, and the loss of active ingredients in the fruit oil is reduced. At the same time, by selecting natural sources and corresponding carriers that promote the absorption of oil-soluble drugs in the body, nano-level emulsification can be achieved, which can improve the efficacy of seabuckthorn oil. Therefore, this method is not only an innovation in the preparation process of traditional functional health edible oil water-soluble products, but also a major breakthrough in the preparation process of high-performance seabuckthorn oil. Summary of the invention

[0005] In view of the problems existing in the prior art such as poor water solubility, easy oxidation, poor taste and poor in vivo absorption of seabuckthorn fruit oil, the purpose of the present invention is to provide a polypeptide-loaded seabuckthorn fruit oil microcapsule powder and its preparation method and application, which can improve the solubility of seabuckthorn fruit oil in aqueous solution and reduce its volatility. The use of organic solvents is avoided in its preparation, meeting the requirements of green environmental protection. At the same time, the health care effect of seabuckthorn fruit oil is further improved, and a new seabuckthorn fruit oil product with good taste, stability and good effect is developed.

[0006] The invention discloses a polypeptide-loaded seabuckthorn fruit oil microcapsule powder, which comprises seabuckthorn fruit oil and a polypeptide carrier material.

[0007] Furthermore, sea buckthorn berry oil is extracted from sea buckthorn berries or sea buckthorn seeds, and the extraction process adopts supercritical extraction or solvent extraction.

[0008] Furthermore, the polypeptide carrier material is selected from one of sea buckthorn peptide, whey protein peptide, soybean protein peptide, wheat protein peptide, corn protein peptide, licorice peptide, astragalus peptide and millet peptide, preferably sea buckthorn peptide.

[0009] Furthermore, the mass ratio of seabuckthorn fruit oil to the polypeptide carrier material is 1:1-1:100, preferably 1:10.

[0010] A preparation method of polypeptide-loaded seabuckthorn oil microcapsule powder comprises the following steps: adding seabuckthorn oil and polypeptide carrier material into a ball mill lined with polytetrafluoroethylene according to the feed ratio and mixing them evenly; adding grinding medium; placing the ball mill in a ball mill for ball milling; and then passing through a 100-mesh sieve to obtain the seabuckthorn oil microcapsule powder.

[0011] Furthermore, the grinding medium is a stainless steel bead with a diameter of 5 to 25 mm, preferably 15 mm, and the filling rate of the grinding medium in the spherical ink tank is 8% to 84%, which is controlled by the amount of steel balls added, and the preferred filling rate is 34%.

[0012] Furthermore, the ball mill is a drum-type ball mill, and its rotation speed is 10 to 50 rpm, preferably 35 rpm.

[0013] Furthermore, the ball milling time is 0.5 to 24 hours, preferably 8 hours.

[0014] The invention discloses an application of a polypeptide-loaded seabuckthorn fruit oil microcapsule powder in the preparation of food, medicine and health care products for anti-oxidation and improvement of non-alcoholic fatty liver.

[0015] By adopting the above technology, compared with the prior art, the beneficial effects of the present invention are embodied in: 1) The seabuckthorn fruit oil microcapsule powder of the present invention can be used as the main raw material to make various solid beverages and liquid beverages, and can also be other solid and liquid food raw materials, mainly functional foods. It is easy to use, flexible, and compatible with various food raw materials and additives. It can be completely dispersed in the solution to form a nano-microemulsion, with a refreshing taste and no special smell. The storage stability and antioxidant properties are higher than those of the seabuckthorn fruit oil raw material, and it has the effects of antioxidant and improving non-alcoholic fatty liver; 2) The present invention prepares microcapsule powder by a limited preparation method, which is simple to operate. Compared with conventional preparation methods such as spray drying, it avoids the use of organic solvents, that is, it avoids the loss of sea buckthorn fruit oil that is very easy to cause during the removal of the solvent. It has the advantages of simple operation, short preparation time, safe and reliable production, low production cost, less pollution, etc., and is a solid food raw material processing and preparation method with good promotion and application. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 The solubility of seabuckthorn fruit oil and the solubility of the active ingredient β-carotene in microcapsule powder prepared with different natural protein peptide carriers; Figure 2 The effects of seabuckthorn oil content on the solubility and dissolution rate of seabuckthorn oil (A), the filling rate of the ball mill (B), the number of ball mill revolutions (C) and the ball milling time (D) on the solubility of seabuckthorn oil in the prepared microcapsule powder and the solubility of the active ingredient β-carotene; Figure 3 is the dissolution curve of seabuckthorn fruit oil / seabuckthorn peptide and seabuckthorn fruit oil in the microcapsule powder of Example 16; Figure 4 is a curve diagram of the remaining content of total carotenoids in seabuckthorn fruit oil / seabuckthorn peptide and microcapsule powder of Example 16 after being placed for different time periods; Figure 5 is a transmission electron micrograph of the solution of Example 16; Figure 6 It is the four common free radical scavenging effects of Example 16 and the physical mixture of seabuckthorn fruit oil / seabuckthorn peptide; Figure 7 This is the therapeutic effect of Example 16 and the physical mixture of seabuckthorn fruit oil / seabuckthorn peptide on non-alcoholic liver damage in mice. DETAILED DESCRIPTION

[0017] The present invention is further described below in conjunction with specific embodiments, but the protection scope of the present invention is not limited thereto: Example 1: Determination of the solubility of seabuckthorn fruit oil

[0018] Accurately weigh 0.063 g of seabuckthorn oil in a 25 ml volumetric flask and dilute with ethyl acetate. Take 0.25, 0.50, 1.00, 1.25, 1.50 and 2.00 ml of the solution in a 10 ml volumetric flask, dilute with ethyl acetate to obtain seabuckthorn oil ethyl acetate solutions with a concentration gradient of 0.063, 0.126, 0.252, 0.315, 0.378 and 0.504 mg / ml. After passing through a 0.45 μm organic filter membrane, measure the absorbance at 451 nm. The standard curve equation is: c 沙棘果油 (g / L) =1.72Abs-0.03(R 2 =0.9996). This indicates that the detection concentration range has a good linear relationship.

[0019] An excess of the sample to be tested was added to a conical flask containing distilled water and placed in a constant temperature oscillator for 24 h (37 °C, 150 rpm) to obtain a supersaturated solution. Three parallel experiments were performed. The obtained supersaturated solution was filtered through a 0.45 μm filter membrane and appropriately diluted, and the content of sea buckthorn fruit oil in the aqueous solution was measured by the above method.

[0020] Example 2: Determination of β-carotene content in seabuckthorn fruit oil

[0021] Liquid phase conditions: chromatographic column is ODS-3 C18 column (4.6×250 mm, 5 μm); flow rate is 1.0 mL / min; injection volume is 10 μL; column temperature is 30 ℃; mobile phase is methanol: water: glacial acetic acid = 75%: 24.5%: 0.5%; detection time is 30min; detection wavelength is 440 nm. β-carotene was diluted to 40 μg / mL, 20 μg / mL, 10 μg / mL, 5 μg / mL, 3 μg / mL, and 1 μg / mL solutions according to a certain ratio. The dilutions were filtered through a 0.45 μm filter membrane and analyzed using the above conditions. C 胡萝卜素 (mg / L) = 32679*PA-10378, R 2= 0.9991, n = 5. An excess of the sample to be tested was added to a conical flask containing distilled water and placed in a constant temperature oscillator for 24 h (37 °C, 150 rpm) to obtain a supersaturated solution. Three parallel experiments were performed. The obtained supersaturated solution was filtered through a 0.45 μm filter membrane and appropriately diluted, and the β-carotene content in the aqueous solution was measured by the above method.

[0022] Embodiment 3:

[0023] Seabuckthorn peptide (6.0 g) and seabuckthorn fruit oil (extracted from seabuckthorn fruit, extracted by supercritical extraction, 0.6 g) were added to a 100 mL polytetrafluoroethylene-lined ball mill, and then 16 stainless steel ball mill beads were added as grinding media, with a ball mill diameter of 15 mm and a filling rate of 34% ((ball volume × number of balls) / total volume of the ball mill). After mixing evenly, the ball mill was placed in a drum ball mill, and the speed was set to 30 rpm for 4 hours. After the ball milling was completed, the powder was sieved through a 100-mesh sieve to obtain seabuckthorn fruit oil microcapsule powder. An appropriate amount of the powder was placed in a conical flask, 10 mL of distilled water was added, and the mixture was shaken for 24 h at 37 ° C and 150 rpm on a shaker. After the supernatant was passed through a 0.45 μm membrane, the solubility of seabuckthorn fruit oil and β-carotene were detected by ultraviolet spectrophotometry and high performance liquid chromatography.

[0024] Embodiment 4:

[0025] Add whey protein peptide (6.0 g) and sea buckthorn fruit oil (extracted from sea buckthorn fruit, the extraction process uses supercritical extraction method 0.6 g) to a 100 mL ball mill, then add 16 stainless steel ball mill beads as grinding media, the ball mill diameter is 15 mm, the filling rate is 34% ((ball volume × number of balls) / total volume of the ball mill), mix well, put the ball mill into a drum ball mill, set the speed to 30 rpm, the time is 4 hours, after the ball milling is completed, the powder is sieved through a 100 mesh sieve to obtain sea buckthorn fruit oil microcapsule powder. Take an appropriate amount of the powder and place it in a conical flask, add 10 mL of distilled water, and shake it on a shaker at 37 ° C and 150 rpm for 24 hours. After the supernatant is passed through a 0.45 μm membrane, the solubility of sea buckthorn fruit oil and β-carotene are detected by ultraviolet spectrophotometry and high performance liquid chromatography.

[0026] Embodiment 5:

[0027] Wheat protein peptide (6.0 g) and sea buckthorn fruit oil (extracted from sea buckthorn fruit, the extraction process uses supercritical extraction method, 0.6 g) were added to a 100 mL ball mill, and then 16 stainless steel ball mill beads were added as grinding media, the ball mill diameter was 15 mm, and the filling rate was 34% ((ball volume × number of balls) / total volume of the ball mill). After mixing evenly, the ball mill was placed in a drum ball mill, and the speed was set to 30 rpm for 4 hours. After the ball milling was completed, the powder was sieved through a 100 mesh sieve to obtain sea buckthorn fruit oil microcapsule powder. After the ball milling was completed, the powder was sieved through a 100 mesh sieve to obtain sea buckthorn fruit oil microcapsule powder. Take an appropriate amount of the powder and place it in a conical flask, add 10 mL of distilled water, and shake it on a shaker at 37°C and 150 rpm for 24 h. After the supernatant was passed through a 0.45 μm membrane, the solubility of sea buckthorn fruit oil and β-carotene were detected by ultraviolet spectrophotometry and high performance liquid chromatography.

[0028] Embodiment 6:

[0029] Soy protein peptide (6.0 g) and sea buckthorn fruit oil (extracted from sea buckthorn fruit, the extraction process uses supercritical extraction method, 0.6 g) were added to a 100 mL ball mill, and then 16 stainless steel ball mill beads were added as grinding media, the ball mill diameter was 15 mm, and the filling rate was 34% ((ball volume × number of balls) / total volume of the ball mill). After mixing evenly, the ball mill was placed in a drum ball mill, and the speed was set to 30 rpm for 4 hours. After the ball milling was completed, the powder was sieved through a 100 mesh sieve to obtain sea buckthorn fruit oil microcapsule powder. After the ball milling was completed, the powder was sieved through a 100 mesh sieve to obtain sea buckthorn fruit oil microcapsule powder. Take an appropriate amount of the powder and place it in a conical flask, add 10 mL of distilled water, and shake it on a shaker at 37°C and 150 rpm for 24 h. After the supernatant was passed through a 0.45 μm membrane, the solubility of sea buckthorn fruit oil and β-carotene were detected by ultraviolet spectrophotometry and high performance liquid chromatography.

[0030] Embodiment 7:

[0031] Zein peptide (6.0 g) and sea buckthorn fruit oil (extracted from sea buckthorn fruit, the extraction process adopts supercritical extraction method, 0.6 g) were added to a 100 mL ball mill, and then 16 stainless steel ball mill beads were added as grinding media. The ball mill beads had a diameter of 15 mm and a filling rate of 34% ((ball volume × number of balls) / total volume of the ball mill). After mixing evenly, the ball mill was placed in a drum ball mill, the speed was set to 30 rpm, and the time was 4 hours. After the ball milling was completed, the powder was sieved through a 100-mesh sieve to obtain sea buckthorn fruit oil microcapsule powder.

[0032] Embodiment 8:

[0033] Licorice polypeptide (6.0 g) and sea buckthorn fruit oil (extracted from sea buckthorn fruit, the extraction process uses supercritical extraction method, 0.6 g) were added to a 100 mL ball mill, and then 16 stainless steel ball mill beads were added as grinding media, the ball mill diameter was 15 mm, and the filling rate was 34% ((ball volume × number of balls) / total volume of the ball mill). After mixing evenly, the ball mill was placed in a drum ball mill, and the speed was set to 30 rpm for 4 hours. After the ball milling was completed, the powder was sieved through a 100 mesh sieve to obtain sea buckthorn fruit oil microcapsule powder. Take an appropriate amount of the powder and place it in a conical flask, add 10 mL of distilled water, and shake it on a shaker at 37°C and 150 rpm for 24 hours. After the supernatant was passed through a 0.45 μm membrane, the solubility of sea buckthorn fruit oil and β-carotene were detected by ultraviolet spectrophotometry and high performance liquid chromatography.

[0034] Embodiment 9:

[0035] Astragalus polypeptide (6.0 g) and sea buckthorn fruit oil (extracted from sea buckthorn fruit, the extraction process uses supercritical extraction method, 0.6 g) were added to a 100 mL ball mill, and then 16 stainless steel ball mill beads were added as grinding media, the ball mill diameter was 15 mm, and the filling rate was 34% ((ball volume × number of balls) / total volume of the ball mill). After mixing evenly, the ball mill was placed in a drum ball mill, and the speed was set to 30 rpm for 4 hours. After the ball milling was completed, the powder was sieved through a 100 mesh sieve to obtain sea buckthorn fruit oil microcapsule powder. Take an appropriate amount of the powder and place it in a conical flask, add 10 mL of distilled water, and shake it on a shaker at 37°C and 150 rpm for 24 hours. After the supernatant was passed through a 0.45 μm membrane, the solubility of sea buckthorn fruit oil and β-carotene were detected by ultraviolet spectrophotometry and high performance liquid chromatography.

[0036] Embodiment 10:

[0037] Millet polypeptide (6.0 g) and sea buckthorn fruit oil (extracted from sea buckthorn fruit, the extraction process uses supercritical extraction method, 0.6 g) were added to a 100 mL ball mill, and then 16 stainless steel ball mill beads were added as grinding media, the ball mill diameter was 15 mm, and the filling rate was 34% ((ball volume × number of balls) / total volume of the ball mill). After mixing evenly, the ball mill was placed in a drum ball mill, and the speed was set to 30 rpm for 4 hours. After the ball milling was completed, the powder was sieved through a 100 mesh sieve to obtain sea buckthorn fruit oil microcapsule powder. After the ball milling was completed, the powder was sieved through a 100 mesh sieve to obtain sea buckthorn fruit oil microcapsule powder. Take an appropriate amount of the powder and place it in a conical flask, add 10 mL of distilled water, and shake it on a shaker at 37°C and 150 rpm for 24 h. After the supernatant was passed through a 0.45 μm membrane, the solubility of sea buckthorn fruit oil and β-carotene were detected by ultraviolet spectrophotometry and high performance liquid chromatography.

[0038] Embodiment 11:

[0039] In order to compare the solubilization effect, sea buckthorn fruit oil (extracted from sea buckthorn fruit using supercritical extraction, 0.6 g) was placed in a conical flask, 10 mL of distilled water was added, and the mixture was shaken for 24 h at 37°C and 150 rpm on a shaker. After the supernatant was passed through a 0.45 μm membrane, the solubility of sea buckthorn fruit oil was 0.35 ± 0.07 g / L and the solubility of β-carotene was 0.75 ± 0.02 mg / L using UV spectrophotometry and high performance liquid chromatography.

[0040] In order to compare the efficacy of peptide carriers, the common inclusion material β-cyclodextrin was added to a 100 mL ball mill as a solubilizing carrier and sea buckthorn fruit oil (extracted from sea buckthorn fruit, the extraction process uses supercritical extraction, 0.6 g), and then 16 stainless steel ball mill beads were added as grinding media, the ball mill diameter was 15 mm, and the filling rate was 34% ((ball volume × number of balls) / total volume of the ball mill). After mixing evenly, the ball mill was placed in a drum ball mill, the speed was set to 30 rpm, and the time was 4 hours. After the ball milling was completed, the powder was sieved through a 100 mesh sieve to obtain sea buckthorn fruit oil microcapsule powder. After the ball milling was completed, the powder was sieved through a 100 mesh sieve to obtain sea buckthorn fruit oil microcapsule powder. Take an appropriate amount of the powder and place it in a conical flask, add 10 mL of distilled water, and shake it on a shaker at 37°C and 150 rpm for 24 hours. After the supernatant was passed through a 0.45 μm membrane, UV spectrophotometry and high performance liquid chromatography were used to detect the solubility of seabuckthorn oil, which was 1.82 ± 0.02 g / L, and the solubility of β-carotene, which was 5.52 ± 0.03 mg / L.

[0041] The experimental results show that sea buckthorn oil and the carotene therein are fat-soluble components and their solubility in water is extremely low.

[0042] Examples 3 to 10 use natural polypeptides with different chemical structures as carriers to prepare seabuckthorn oil-polypeptide microcapsule powders, and the solubility of seabuckthorn oil and β-carotene therein is determined, for example Figure 1 As shown in the figure, the solubilization effects of various plant protein peptides on sea buckthorn oil and β-carotene in it vary greatly, and most of the peptides have a better solubilization effect on sea buckthorn oil than sea buckthorn oil itself and sea buckthorn oil wrapped in β-cyclodextrin. Further carrier optimization results show that sea buckthorn peptide has the best effect as a carrier, and the solubility of sea buckthorn oil and β-carotene is the greatest, so the optimal peptide carrier is sea buckthorn peptide.

[0043] Example 12: Optimal amount of carrier

[0044] The process was optimized by using the formula of seabuckthorn peptide loaded with seabuckthorn fruit oil. Seabuckthorn peptide (6g, 12g, 30g, 60g, 120g) and seabuckthorn fruit oil (extracted from seabuckthorn fruit, the extraction process adopts supercritical extraction method 6.0g) were added to a 100 mL ball mill, and the ratio of seabuckthorn fruit oil: seabuckthorn peptide was controlled to be (1:1~1:20). 16 stainless steel ball mill beads were added as grinding media, the ball mill beads had a diameter of 15mm, and the filling rate was 34% ((ball volume × number of balls) / total volume of the ball mill), and after mixing evenly, the ball mill was placed in a drum ball mill, and the speed was set to 30 rpm for 4 hours. After the ball milling was completed, the powder was sieved through a 100 mesh sieve to obtain seabuckthorn fruit oil microcapsule powder. Take an appropriate amount of the powder and place it in a conical flask, add 10 mL of distilled water, and shake it on a shaker at 37°C and 150 rpm for 24 hours. After the supernatant was passed through a 0.45 μm membrane, the solubility of seabuckthorn oil was detected by UV spectrophotometry and high performance liquid chromatography, and the amount of solubilized seabuckthorn oil in the total added seabuckthorn oil (seabuckthorn oil dissolution rate) was calculated. The experimental results are shown in Figure 2 As shown in A. The test results show that, although the solubility of seabuckthorn oil increases with the increase of the amount of seabuckthorn oil, the percentage of solubilized seabuckthorn oil in the total seabuckthorn oil decreases. By balancing the solubility and the solubilization rate, the optimal content of seabuckthorn oil in the prepared seabuckthorn oil microcapsule powder is 10%.

[0045] Example 13: Optimization of ball mill filling rate

[0046] The process was optimized with a formula of seabuckthorn peptide loaded with seabuckthorn fruit oil in a ratio of 10:1. Seabuckthorn peptide (6.0 g) and seabuckthorn fruit oil (extracted from seabuckthorn fruit, the extraction process adopts supercritical extraction method, 0.6 g) were added to a 100 mL ball mill. Different numbers of ball milling beads were added as grinding media. The diameter of the ball milling beads was 15 mm. The filling rates were controlled to be 8%, 16%, 25%, 34%, 58%, 67%, 75%, and 84%, respectively. After mixing evenly, the ball mill was placed in a drum ball mill. The speed was set to 30 rpm for 4 hours. After the ball milling was completed, the powder was sieved through a 100-mesh sieve to obtain seabuckthorn fruit oil microcapsule powder. The solubility detection step was the same as in Example 3. The solubility of seabuckthorn fruit oil and β-carotene in the prepared seabuckthorn fruit oil microcapsule powder was as follows: Figure 2 As shown in B in the figure. The test results show that if the filling rate is too low, the grinding is insufficient and the solubilization effect is poor; while if the filling rate is too high, the running space of the grinding media is reduced and the solubilization effect is poor, so the preferred filling rate is 34%.

[0047] Example 14: Optimization of ball mill speed

[0048] The process was optimized with a formula of seabuckthorn peptide loaded with seabuckthorn fruit oil in a ratio of 10:1. Seabuckthorn peptide (6.0 g) and seabuckthorn fruit oil (extracted from seabuckthorn fruit or seabuckthorn seeds, the extraction process uses supercritical extraction or solvent extraction 0.6 g) were added to a 100 mL ball mill, and then 16 stainless steel ball mill beads were added as grinding media. The ball mill beads had a diameter of 15 mm and a filling rate of 34%. After mixing evenly, the ball mill was placed in a drum ball mill. The speed of the ball mill was controlled to be 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, and 60 rpm, respectively, for 4 hours. After the ball milling was completed, the powder was sieved through a 100-mesh sieve to obtain seabuckthorn fruit oil microcapsule powder. The solubility detection step was the same as in Example 3. The solubility of seabuckthorn fruit oil and β-carotene in the prepared seabuckthorn fruit oil microcapsule powder was as follows: Figure 2 As shown in C. The test results show that if the rotation speed is too low, the grinding and dispersion are not sufficient, resulting in poor solubilization effect; while if the grinding rate is too high, the system exotherm increases, causing oxidation and polymerization of the essential oil, and the solubility decreases significantly, so the preferred rotation speed is 35 rpm.

[0049] Example 15: Optimization of ball milling time

[0050] Seabuckthorn peptide (6.0 g) and seabuckthorn fruit oil (0.6 g) were added to a 100 mL ball mill, and then 16 stainless steel ball mill beads were added as grinding media. The ball mill beads had a diameter of 15 mm and a filling rate of 34%. After mixing evenly, the ball mill was placed in a drum ball mill. The speed of the ball mill was controlled to 30 rpm, and ball milling was started. Samples were taken at 0.5, 1, 2, 4, 8, 16, and 24 hours, respectively. The solubility detection steps were the same as in Example 3. The solubility of seabuckthorn fruit oil and β-carotene in the prepared seabuckthorn fruit oil microcapsule powder was as follows: Figure 2 As shown in D in the figure. The test results show that when the grinding time is short, the sea buckthorn fruit oil fails to fully contact with the carrier, resulting in poor solubilization effect; and as the grinding time increases, the solubilization effect gradually decreases and the solubility tends to be flat, so considering energy saving comprehensively, the preferred grinding time is 8h.

[0051] Example 16: Optimal Process and Conditions for Seabuckthorn Oil Microcapsule Powder

[0052] After the formulation and process optimization, the preparation method of seabuckthorn fruit oil microcapsule powder prepared with seabuckthorn peptide as carrier is as follows: seabuckthorn peptide (6.0 g) and seabuckthorn fruit oil (extracted from seabuckthorn fruit, the extraction process adopts supercritical extraction method, 0.6 g) are added to a 100 mL ball mill, and then 16 stainless steel ball mill beads are added as grinding media, the ball mill beads have a diameter of 15 mm, and the filling rate is 34% ((ball volume × number of balls) / total volume of the ball mill), and after mixing evenly, the ball mill is placed in a drum ball mill, the speed is set to 35 rpm, and the time is 8 hours. After the ball milling is completed, the powder is sieved through a 100 mesh sieve to obtain seabuckthorn fruit oil microcapsule powder. The solubility of seabuckthorn fruit oil in the microcapsule powder is 9.21±0.11 g / L, and the solubility of β-carotene is 72.71±4.72 mg / L.

[0053] Example 17: Dissolution Test

[0054] The dissolution rate of seabuckthorn fruit oil in the microcapsule powder was determined by taking seabuckthorn fruit oil and its main component β-carotene as indicators. The method is as follows: Example 16 (0.55 g, equivalent to about 50 mg of seabuckthorn fruit oil) and a physical mixture of seabuckthorn fruit oil (50 mg) and seabuckthorn peptide (0.5 g) (referred to as seabuckthorn fruit oil / seabuckthorn peptide) were accurately weighed for dissolution experiment. The paddle method was used: 900 mL of purified water was used as the dissolution medium at a temperature of 37 o C, rotation speed 100 rpm, dissolution time 120 min; 5 ml of each solution was taken at 5 min, 10 min, 20 min, 30 min, 60 min, 75 min, 90 min and 120 min, and 37 o 5 ml of purified water was added, and the sample solution was filtered through a 0.45 μm microporous filter membrane as a test sample. The concentration of seabuckthorn oil was determined by ultraviolet spectrophotometry, and the dissolution curve of seabuckthorn oil with solubility as the measurement index was obtained. Figure 3 From the solubilization effect on seabuckthorn fruit oil, it can be seen that the microcapsule powder shown in Example 16 not only significantly improves the solubility of seabuckthorn fruit oil and its key components, but also effectively promotes the rapid dissolution of seabuckthorn fruit oil, and is significantly better than the physical mixture, indicating that seabuckthorn peptides have good surface activity and promote the water dispersion of fat-soluble seabuckthorn fruit oil.

[0055] Example 18: Accelerated aging assessment of seabuckthorn fruit oil microcapsule powder

[0056] Take the microcapsule powder of Example 16 (5.5 g, of which the content of seabuckthorn fruit oil is about 0.5 g) and the physical mixture of seabuckthorn fruit oil and seabuckthorn peptide in a ratio of 1:10 (5.5 g, of which the content of seabuckthorn fruit oil is about 0.5 g), spread the sample flat to make the thickness uniform, seal it with an aluminum foil bag, and place it in an assessment box (set temperature 45 oC±2 humidity RH 65%±2) for 6 months, then sample 0.55g at 0.5, 1, 2, 3, 4, 5, and 6 months, dissolve in 95% ethanol, make up to 100mL, dilute to a certain concentration, record the dilution multiple, and use HPLC to determine the mass m of β-carotene in the sample. t , β-carotene remaining = (m t / m0)×100%; the result is Figure 4 As shown, after accelerated aging test, the residual amount of β-carotene in Example 16 can still reach more than 95%, indicating that all the examples have good stability and can be stored for a long time at room temperature.

[0057] Example 19: Observation of solution microstructure

[0058] To further observe the structure of nanocapsules formed in the aqueous solution of Example 16 ( Figure 5 ), a JEM 2100EX transmission electron microscope was used to observe the morphology of nanocapsules at an accelerating voltage of 100 kV. First, the microcapsule powder of Example 16 was diluted in ultrapure water, ultrasonicated for 5 min, and then placed on a copper mesh, and then stained with uranyl acetate (2% w / w). The results showed that there were a large number of microcapsules in the solution under the microscopic level. The microcapsules were uniformly dispersed and regular spherical structures, and the color inside the sphere was obviously darker, which was clearly distinguished from the peripheral light-colored part, indicating that the seabuckthorn fruit oil was successfully encapsulated by the seabuckthorn peptide, and micelle-like self-assembly behavior occurred in the aqueous solution.

[0059] The particle size and zeta potential of Example 16 were further measured by using a Malvern Nano ZS90 dynamic light scattering nanoparticle size analyzer. Example 16 was dissolved in 25°C distilled water (0.1 mg / ml), and then an appropriate amount of the solution was taken and placed in the detector for measurement. The measurement was repeated 20 times. The experimental results showed that the particle size of the microcapsule powder solution of Example 16 was 131.30 nm, and the zeta value was (-25.2 ± 0.3). The microcapsule powder of Example 16 formed stable nano-scale microcapsules in the solution.

[0060] Example 20: Study on the free radical scavenging ability of seabuckthorn oil microcapsule powder

[0061] Through experiments, the antioxidant capacity of the seabuckthorn fruit oil / peptide mixture of Example 16 at different concentrations was determined, with vitamin C as a control. Figure 6 The results showed that the conjugated structure of carotene and unsaturated fatty acids in seabuckthorn fruit oil determined its excellent antioxidant properties. Figure 6The results of the scavenging rates of Example 16, seabuckthorn oil / peptide mixture and VC on DPPH free radicals, superoxide free radicals, hydroxyl free radicals and ABTS free radicals are shown respectively. The results show that when the concentration of the positive control vitamin C is 0.8 mg / mL, the scavenging rates of DPPH, superoxide anion, hydroxyl and ABTS free radicals are 85.2%, 73.2%, 65.32% and 97.10%, respectively. Compared with the seabuckthorn oil / peptide mixture, the ability of Example 16 to scavenge the above four types of free radicals is significantly improved, and the scavenging rate increases by 5%-10%. The scavenging effect on hydroxyl free radicals is even better than that of VC, which fully demonstrates that Example 16 has a more excellent free radical scavenging effect and better antioxidant capacity.

[0062] Example 21: Study on the therapeutic effect of non-alcoholic fatty liver disease

[0063] 40 healthy male ICR mice were randomly divided into a normal group, a model group, an SJY group, and a SJY-SJT group. The mice were fed a high-fat diet and intraperitoneally injected with tetracycline saline solution at a dose of 15 mg / 100 g at 10 am every day for 5 consecutive days to establish a non-alcoholic liver injury model. In the SJY group, seabuckthorn fruit oil and seabuckthorn peptide were suspended in pure water at a ratio of 1:10 and administered by gavage at a dose of 30 mg / 100 g; in the SJY-SJT group, Example 16 was dissolved in pure water and administered by gavage at a dose of 30 mg / 100 g; the normal control group and the model group were given pure water by gavage according to body weight (the normal group was normal animals, and the model group was non-alcoholic liver injury model animals without treatment); the drug was administered continuously for 15 days. After the end, the weight of the mice in each group was weighed, the liver was dissected and removed, and the stained sections were fixed ( Figure 7 ), use the corresponding kits to measure serum and liver homogenate total cholesterol, triglycerides, high-density lipoprotein, low-density lipoprotein, aspartate transaminase, alanine transaminase and other biochemical indexes, see Figure 7 .

[0064] The results of blood biochemical indexes of mice showed that compared with the normal control group, the total TG, TC, HDL-C, LDL-C, AST and ALT of the model group mice serum were significantly increased (*P<0.01), indicating the formation of liver damage; while the SJY-SJT group and the SJY group could effectively reduce the related factors, among which the serum TC, HDL-C, ALT and LDL-C of the SJY-SJT group were significantly better than those of the SJY group (P<0.01). Further liver-related factor detection and liver section showed that the TC and LDL-C of the liver in the SJY-SJT group were reduced. Liver section observation showed that the hepatic cords in the SJY-SJT group were neatly arranged, and the hepatic sinusoids were not significantly expanded or squeezed; more hepatocytes were slightly loose in the tissue (black arrow); there were few lymphocytes and neutrophils infiltration, which was close to the normal group, better than the SJY group, and significantly better than the model group. The above experimental results show that seabuckthorn fruit oil can relieve fatty liver to a certain extent, and after being prepared into seabuckthorn fruit oil microcapsule powder-Example 16, its ability to relieve fatty liver is significantly increased, and it is more suitable for development as a functional food.

Claims

1. A polypeptide-loaded seabuckthorn fruit oil microcapsule powder, characterized in that: Its ingredients include sea buckthorn fruit oil and polypeptide carrier material.

2. The polypeptide-loaded seabuckthorn fruit oil microcapsule powder according to claim 1, characterized in that: Sea buckthorn berry oil is extracted from sea buckthorn berries or sea buckthorn seeds, and the extraction process uses supercritical extraction or solvent extraction.

3. The polypeptide-loaded seabuckthorn fruit oil microcapsule powder according to claim 1, characterized in that: The polypeptide carrier material is selected from one of seabuckthorn peptide, whey protein peptide, soybean protein peptide, wheat protein peptide, corn protein peptide, licorice polypeptide, astragalus polypeptide and millet polypeptide.

4. The polypeptide-loaded seabuckthorn fruit oil microcapsule powder according to claim 1, characterized in that: The mass ratio of seabuckthorn fruit oil to polypeptide carrier material is 1:1-1:

100.

5. A method for preparing the polypeptide-loaded seabuckthorn fruit oil microcapsule powder according to any one of claims 1 to 4, characterized in that: According to the feed ratio, sea buckthorn fruit oil and polypeptide carrier material are added to a ball mill lined with polytetrafluoroethylene and mixed evenly, grinding media is added, and the ball mill is placed in a ball mill for ball milling, and then passed through a 100-mesh sieve to obtain sea buckthorn fruit oil microcapsule powder.

6. The method for preparing a polypeptide-loaded seabuckthorn fruit oil microcapsule powder according to claim 5, characterized in that: The grinding media are stainless steel beads with a diameter of 5 to 25 mm. The filling rate of the grinding media in the spherical ink tank is 8% to 84%, which is controlled by the amount of steel balls added.

7. The method for preparing a polypeptide-loaded seabuckthorn fruit oil microcapsule powder according to claim 5, characterized in that: The ball mill is a drum-type ball mill, and its rotation speed is 10 to 50 rpm.

8. The method for preparing a polypeptide-loaded seabuckthorn fruit oil microcapsule powder according to claim 5, characterized in that: The ball milling time is 0.5~24h.

9. Use of the polypeptide-loaded seabuckthorn fruit oil microcapsule powder according to any one of claims 1 to 4 in the preparation of food, medicine and health care products, characterized in that: Used for preparing foods, medicines and health products for anti-oxidation and improving non-alcoholic fatty liver.

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