Polyphenol compounded algal oil anti-inflammatory composition and preparation method thereof

Abstract

The invention provides an anti-inflammatory composition of polyphenol compounded algal oil and a preparation method of the anti-inflammatory composition, and belongs to the technical field of biology. The anti-inflammatory composition comprises a polyphenol compound, algae oil, modified resistant dextrin, trehalose and vitamin E succinate, wherein the polyphenol compound is prepared by compounding a clove extract and a sanguisorba officinalis extract. The polyphenol compound scavenges free radicals and protects DHA / EPA in the algal oil from oxidative degradation, the bioavailability of the polyphenol compound is improved by the fat-soluble environment of the algal oil, and the polyphenol compound and the algal oil are compounded to cover the whole inflammation process in a multi-target manner and synergistically improve the anti-inflammatory effect; the modified resistant dextrin can improve the stability and bioavailability of the polyphenol compound and the algal oil, and gastric acid protection and intestinal targeted release are realized. Animal experiment results show that the composition disclosed by the invention can play an anti-inflammatory role by inhibiting release of inflammatory factors and reducing tissue damage, and has good application potential.

Description

Technical Field

[0001] The present invention relates to the field of biotechnology, and in particular to an anti-inflammatory composition having synergistic effects between a polyphenol compound and algae oil and a preparation method thereof, and particularly to a functional anti-inflammatory preparation using a deep eutectic solvent-assisted extraction technology and a nanoemulsion delivery system. Background Art

[0002] Inflammation, as the body's defensive response to stimuli, plays a vital role in maintaining health. A moderate inflammatory response helps the body defend against invading pathogens, promptly removes damaged tissue, and ultimately restores internal stability. However, when inflammatory responses become uncontrolled and chronic, the situation becomes extremely serious. Chronic inflammation is closely linked to numerous serious health threats, including cardiovascular disease, neurodegenerative diseases, autoimmune diseases, and cancer, placing a heavy burden on global healthcare systems. Therefore, the development of safe and effective anti-inflammatory products is urgent.

[0003] Among the many substances with anti-inflammatory potential, polyphenols and algae oil stand out and have attracted much attention. Polyphenols are widely found in various parts of plants, such as roots, stems, leaves, flowers, and fruits. Their chemical structure contains multiple phenolic hydroxyl groups, which gives them unique chemical and physiological activities. Algae oil is rich in polyunsaturated fatty acids, especially DHA and EPA, and is a high-quality oil extracted from algae. Although polyphenols and algae oil have shown great potential in the anti-inflammatory field, they still face many challenges in practical application. First, polyphenols and algae oil have poor water solubility, which severely limits their absorption, distribution, and metabolism in the body, resulting in low bioavailability and difficulty in fully exerting their powerful anti-inflammatory effects. Second, polyphenols and algae oil are extremely sensitive to external factors such as light, heat, and oxygen, and are prone to oxidation and degradation reactions, resulting in reduced activity.

[0004] Although polyphenols have significant anti-inflammatory activity, their low bioavailability and poor stability limit their widespread application in practice. Therefore, developing a method to effectively improve the bioavailability and stability of polyphenols, as well as optimizing polyphenol compound anti-inflammatory compositions, has important practical significance and application value, and also points the way for research in related fields. Summary of the Invention

[0005] The present invention aims to provide an anti-inflammatory composition of polyphenols compounded with algae oil and a preparation method thereof. The anti-inflammatory composition of the present invention utilizes the synergistic anti-inflammatory mechanism of clove extract and sanguisorba officinalis extract, and is compounded with algae oil rich in DHA / EPA to achieve multi-target regulation of the entire inflammatory response. At the same time, octenylsuccinic anhydride-modified resistant dextrin is used as a carrier to construct a nano-delivery system to achieve gastric acid protection and intestinal targeted release. The anti-inflammatory composition provided by the present invention has excellent anti-inflammatory effects.

[0006] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0007] The present invention provides an anti-inflammatory composition of a polyphenol-algae oil complex, comprising the following raw materials in parts by weight: 20-30 parts of a polyphenol complex, 30-40 parts of algae oil, 15-25 parts of modified resistant dextrin, 15-25 parts of trehalose, and 1-3 parts of vitamin E succinate; the polyphenol complex is composed of a clove extract and a sanguisorba officinalis extract in a weight ratio of 1-3:1.

[0008] Preferably, the preparation method of the clove extract comprises: mixing clove buds, a low eutectic solvent and water, extracting the mixture through a high-voltage pulse electric field, centrifuging, and purifying the supernatant through a macroporous resin to obtain the clove extract.

[0009] More preferably, the deep eutectic solvent is composed of choline chloride and urea in a molar ratio of 1:1.5-2.5, and the weight ratio of the clove buds, the deep eutectic solvent and water is 1:(18-23):(2-4).

[0010] Preferably, the preparation method of the Sanguisorba officinalis extract comprises: mixing the Sanguisorba officinalis root with a 30%-40% ethanol solution by volume, ultrasonically treating for 40-50 minutes, centrifuging, mixing the precipitate, a low eutectic solvent and water, ultrasonically extracting, centrifuging, and purifying the supernatant with a macroporous resin to obtain the Sanguisorba officinalis extract.

[0011] More preferably, the eutectic solvent is composed of choline chloride and lactic acid in a molar ratio of 1:1.5-2.5, and the weight ratio of the precipitate, the deep eutectic solvent and water is 1:8-12:1-2.

[0012] Preferably, the preparation method of the modified resistant dextrin preferably comprises: mixing resistant dextrin with water, adjusting the pH value to 8.5-9, adding octenyl succinic anhydride for reaction, centrifuging, and precipitating the modified resistant dextrin.

[0013] More preferably, the reaction temperature is 58-62° C., the rotation speed is 250-350 rpm, and the reaction time is 6-7 h.

[0014] The present invention also provides a preparation method of the above-mentioned anti-inflammatory composition, comprising: mixing a polyphenol complex, algae oil and vitamin E succinate to obtain an oil phase; mixing modified resistant dextrin, trehalose and water to obtain an aqueous phase; adding the oil phase to the aqueous phase, homogenizing to obtain a nanoemulsion, and freeze-drying to obtain the anti-inflammatory composition.

[0015] Preferably, the homogenization comprises: first shearing at 8000-12000 rpm for 4-6 minutes, and then high-pressure homogenization at 45-55 MPa for 8-12 minutes.

[0016] The present invention also provides a use of the anti-inflammatory composition in preparing anti-inflammatory drugs.

[0017] Compared with the prior art, the present invention has the following beneficial effects:

[0018] The present invention provides an anti-inflammatory composition comprising a polyphenol compound and algae oil, comprising: a polyphenol compound, algae oil, modified resistant dextrin, trehalose, and vitamin E succinate. The polyphenol compound is compounded from clove extract and sanguisorba officinalis extract. The polyphenol compound scavenges free radicals, protecting DHA / EPA in the algae oil from oxidative degradation. The fat-soluble environment of the algae oil improves the bioavailability of the polyphenol compound. The combination of the two can provide multi-target coverage throughout the entire inflammation process, synergistically enhancing the anti-inflammatory effect. The modified resistant dextrin can improve the stability and bioavailability of the polyphenol compound and algae oil, achieving gastric acid protection and intestinal targeted release.

[0019] This invention, for the first time, uses a dual eutectic solvent system of choline chloride-urea and choline chloride-lactic acid to perform customized extractions targeting the polyphenolic properties of cloves and sanguisorba officinalis, respectively, thereby enriching polyphenols with anti-inflammatory properties and improving the anti-inflammatory effect. The invention innovatively combines polyphenol complexes with algae oil to achieve dual synergistic antioxidant and anti-inflammatory effects, significantly improving the anti-inflammatory effect. The invention uses a nanoemulsion system constructed by modifying resistant dextrin with octenylsuccinic anhydride, significantly improving oral bioavailability.

[0020] The results of animal experiments show that the composition of the present invention can exert anti-inflammatory effects by inhibiting the release of inflammatory factors and reducing tissue damage, and has good application potential. DETAILED DESCRIPTION

[0021] The present invention provides an anti-inflammatory composition of a polyphenol-algae oil complex, comprising: 20-30 parts of a polyphenol complex, 30-40 parts of algae oil, 15-25 parts of modified resistant dextrin, 15-25 parts of trehalose and 1-3 parts of vitamin E succinate; the polyphenol complex is composed of a clove extract and a sanguisorba officinalis extract in a weight ratio of 1-3:1.

[0022] The preparation method of the clove extract of the present invention preferably comprises: grinding clove buds through an 80-120 mesh sieve to obtain clove bud powder, mixing the powder with a deep eutectic solvent (composed of choline chloride and urea in a molar ratio of 1:1.5-2.5) and water in a weight ratio of 1:(18-23):(2-4), extracting the powder with a high-voltage pulsed electric field at an electric field strength of 12-17 kV / cm, a pulse width of 18-22 μs, a frequency of 1.5-3 Hz, and a temperature of 28-32° C. for 100-160 s, and centrifuging the powder at 2800-3200 rpm for 3-7 m. in, taking the supernatant, adjusting the pH of the supernatant to 5-6, adsorbing it through a XAD-7HP macroporous resin column at a flow rate of 1.3-1.8 BV / h, eluting it in sequence with 2-4 BV of water (flow rate 1.6-2.3 BV / h), 2-4 BV of a 43%-48% ethanol solution by volume (flow rate 1.3-1.8 BV / h), and 4-6 BV of a 67%-73% ethanol solution by volume (flow rate 1.3-1.8 BV / h), collecting the 67%-73% ethanol eluate by volume, and freeze-drying it in vacuo to obtain a clove extract.

[0023] The invention uses choline chloride as a hydrogen bond acceptor and urea as a hydrogen bond donor to form a deep eutectic solvent, which has high solubility for polyphenol compounds and improves the extraction of polyphenols from clove buds. High-voltage pulses form instantaneous micropores in cell membranes, promoting rapid penetration of the deep eutectic solvent and release of polyphenols, thereby improving the extraction efficiency of polyphenols. XAD-7HP resin has a strong adsorption capacity for phenols, and gradient elution can significantly improve the purity of polyphenols.

[0024] The preparation method of the Sanguisorba officinalis extract of the present invention preferably comprises: crushing the Sanguisorba officinalis root through a 100-150 mesh sieve to obtain Sanguisorba officinalis root powder, mixing it with a volume fraction of 30%-40% ethanol solution at a solid-liquid ratio of 1:12-18 g / mL, adjusting the pH value to 3-4 with citric acid (to prevent polyphenol oxidation), ultrasonically extracting at a power of 270-320 W, a frequency of 38-42 kHz, and 45-50 ° C for 40-50 min, and centrifuging at 3800-4200 rpm for 8-12 min to obtain supernatant 1 and a precipitate; adding anhydrous ethanol to the supernatant 1 to a volume fraction of 67%-73%, precipitating at 4 ° C for 10-14 hours, discarding the precipitate to obtain supernatant 2; mixing the precipitate with a low eutectic solvent (composed of choline chloride and lactic acid at a molar ratio of 1:1.5-2.5) and water at a weight ratio of 1:8 -12:1-2 mixture, ultrasonic extraction at a power of 230-280 W, a frequency of 33-37 kHz, and 38-42°C for 25-35 min, centrifugation at 3800-4200 rpm for 8-12 min to obtain supernatant 3, which was mixed with supernatant 2 and passed through a 0.45 μm membrane. The pH was adjusted to 4.5-5.5, and adsorbed through an HPD-600 macroporous resin at a flow rate of 1-1.5 BV / h. The mixture was eluted in sequence with 4-6 BV of water (flow rate 1.8-2.3 BV / h), 2-4 BV of a volume fraction of 33%-38% ethanol solution (flow rate 1-1.5 BV / h), and 4-6 BV of a volume fraction of 73%-78% ethanol solution (flow rate 1-1.5 BV / h). The eluate with a volume fraction of 73%-78% ethanol was collected and freeze-dried in vacuo to obtain a Sanguisorba officinalis extract.

[0025] The invention first extracts the Sanguisorba officinalis with a 30%-40% ethanol solution, and then performs alcohol precipitation on the supernatant after the extraction, thereby removing the polysaccharide components in the Sanguisorba officinalis and retaining the polyphenol components. According to the invention, based on the characteristics of the Sanguisorba officinalis polyphenols, the invention extracts the precipitate with a low eutectic solvent composed of choline chloride and lactic acid, thereby improving the solubility of the polyphenols in the precipitate and improving the extraction efficiency of the polyphenols. During the extraction process, the pH value is adjusted to acidic to maintain the chemical stability and biological activity of the polyphenols. The Sanguisorba officinalis extract obtained by the invention can inhibit the inflammatory reaction caused by oxidative stress, improve autoimmune inflammation, and regulate immune balance. It can be compounded with clove extract, algae oil, etc. for synergistic effect, thereby improving the anti-inflammatory effect.

[0026] The algae oil of the present invention is rich in DHA and EPA, which can inhibit the synthesis of inflammatory mediators, regulate the release of inflammatory factors, promote the disappearance of inflammation, and improve chronic inflammatory damage.

[0027] The polyphenol complex of the present invention is compounded with algae oil, and multiple targets are covered throughout the entire inflammation process (the polyphenol complex can inhibit the initiation of inflammation, and the algae oil can promote the resolution of inflammation). The polyphenol complex scavenges free radicals and protects the DHA / EPA in the algae oil from oxidative degradation. The fat-soluble environment of the algae oil improves the bioavailability of the polyphenol complex. The combination of the two can enhance the anti-inflammatory effect.

[0028] The preparation method of the modified resistant dextrin of the present invention preferably includes: mixing resistant dextrin with water at 58-62°C and 300-500rpm for 10-15 minutes to obtain a resistant dextrin solution with a mass fraction of 28%-32%, adjusting the pH value to 8.5-9 (to ensure the reactivity of octenyl succinic anhydride), adding octenyl succinic anhydride (the addition amount is 3%-8% of the weight of the resistant dextrin), reacting at 58-62°C and 250-350rpm for 6-7 hours, adjusting the pH value to 6.3-6.8, centrifuging at 8000-10000rpm for 10-15 minutes, removing the precipitated insoluble matter, passing the supernatant through a polysulfone membrane (molecular weight cutoff 3.5kDa) to remove free octenyl succinic anhydride, and spray drying to obtain modified resistant dextrin.

[0029] The present invention aims to improve the drug loading capacity of resistant dextrin for algae oil (fat-soluble) and polyphenols (partially water-soluble), and enhances its hydrophobicity and embedding capacity by modifying the resistant dextrin with octenylsuccinic anhydride, thereby achieving gastric acid protection and intestinal targeted release.

[0030] The trehalose of the present invention can provide freeze-drying protection and maintain the stability of the preparation; vitamin E succinate acts as an antioxidant to prevent the oxidation of algae oil.

[0031] The present invention also provides a preparation method of the above-mentioned anti-inflammatory composition, comprising: mixing a polyphenol complex, algae oil and vitamin E succinate to obtain an oil phase; mixing modified resistant dextrin, trehalose and water to obtain an aqueous phase; adding the oil phase to the aqueous phase, homogenizing to obtain a nanoemulsion, and freeze-drying to obtain the anti-inflammatory composition.

[0032] The homogenization of the present invention preferably includes: first shearing at 8000-12000 rpm for 4-6 minutes, and then high-pressure homogenization at 45-55 MPa for 8-12 minutes.

[0033] The present invention also provides a use of the anti-inflammatory composition in preparing anti-inflammatory drugs.

[0034] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions of the present invention. Obviously, the embodiments described are only some of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0035] In the following examples, unless otherwise specified, all methods are conventional.

[0036] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0037] The resistant dextrin involved in the examples and comparative examples: resistant dextrin powder, Shandong Bailong Chuangyuan Biotechnology Co., Ltd.; algae oil: Peptide An Biotechnology Co., Ltd., product number TASW-061301.

[0038] Example 1

[0039] (1) Preparation of polyphenol complexes

[0040] 1) Preparation of clove extract

[0041] The clove buds were crushed through a 100-mesh sieve to obtain clove bud powder, which was then mixed with a deep eutectic solvent (composed of choline chloride and urea in a molar ratio of 1:2) and water in a weight ratio of 1:20:3. The mixture was extracted with a high-voltage pulsed electric field at an electric field intensity of 15 kV / cm, a pulse width of 20 μs, a frequency of 2 Hz, and a temperature of 30°C for 120 s. The mixture was centrifuged at 3000 rpm for 5 min, and the supernatant was taken. The pH of the supernatant was adjusted to 5.5, and the mixture was adsorbed on a XAD-7HP macroporous resin column at a flow rate of 1.5 BV / h. The mixture was eluted with 3 BV of water (flow rate 2 BV / h), 3 BV of a 45% ethanol solution by volume (flow rate 1.5 BV / h), and 5 BV of a 70% ethanol solution by volume (flow rate 1.5 BV / h). The 70% ethanol eluate was collected and vacuum freeze-dried to a water content of 2 wt% to obtain a clove extract.

[0042] 2) Preparation of Sanguisorba officinalis extract

[0043] Sanguisorba officinalis root powder was ground through a 120-mesh sieve to obtain Sanguisorba officinalis root powder. The powder was mixed with a 35% ethanol solution at a solid-liquid ratio of 1:15 g / mL. The pH was adjusted to 3.5 with citric acid. Ultrasonic extraction was performed at 300 W, 40 kHz, and 48°C for 45 minutes. The solution was centrifuged at 4000 rpm for 10 minutes to obtain supernatant 1 and a precipitate. Anhydrous ethanol was added to supernatant 1 to a 70% volume fraction, and the solution was allowed to precipitate at 4°C for 12 hours. The precipitate was discarded to obtain supernatant 2. The precipitate was then mixed with a deep eutectic solvent (composed of choline chloride and lactic acid in a molar ratio of 1:1.5-2.5) and water in a weight ratio of 1:10:1. 5, ultrasonic extraction was carried out at a power of 250 W, a frequency of 35 kHz, and 40°C for 30 min, and supernatant 3 was obtained by centrifugation at 4000 rpm for 10 min to obtain supernatant 3. The supernatant 3 was mixed with supernatant 2 and passed through a 0.45 μm membrane. The pH was adjusted to 5, and the supernatant was adsorbed on a HPD-600 macroporous resin at a flow rate of 1.3 BV / h. The supernatant was eluted in sequence with 5 BV of water (flow rate 3 BV / h), 3 BV of 35% ethanol solution by volume (flow rate 1.3 BV / h), and 5 BV of 75% ethanol solution by volume (flow rate 1.3 BV / h). The 75% ethanol eluate was collected and freeze-dried in vacuum to obtain the Sanguisorba officinalis extract.

[0044] 3) Preparation of polyphenol complexes

[0045] The clove extract and the sanguisorba officinalis extract are mixed in a weight ratio of 2:1 to obtain a polyphenol complex.

[0046] (2) Preparation of modified resistant dextrin

[0047] Resistant dextrin was mixed with water at 60°C and 400 rpm for 12 minutes to obtain a 30% mass fraction of resistant dextrin solution, the pH value was adjusted to 8.7, octenylsuccinic anhydride was added (the addition amount was 5% of the weight of the resistant dextrin), and the mixture was reacted at 60°C and 300 rpm for 6.5 hours. The pH value was adjusted to 6.5, and the mixture was centrifuged at 9000 rpm for 12 minutes to remove the precipitated insoluble matter. The supernatant was passed through a polysulfone membrane (molecular weight cutoff 3.5 kDa) to remove free octenylsuccinic anhydride, and the retained portion was dried to obtain modified resistant dextrin.

[0048] (3) Preparation of anti-inflammatory composition

[0049] Accurately weigh 25 parts of polyphenol complex, 35 parts of algae oil, 20 parts of modified resistant dextrin, 20 parts of trehalose, and 2 parts of vitamin E succinate;

[0050] The polyphenol complex, algal oil and vitamin E succinate were mixed, mixed at 55°C and 250 rpm for 5 minutes, and then homogenized at 8000 rpm for 5 minutes to obtain an oil phase; the modified resistant dextrin, trehalose and water were mixed at 60°C and 400 rpm for 12 minutes to obtain an aqueous phase with a solid content of 20 wt%; the oil phase was added to the aqueous phase, first sheared at 10000 rpm for 5 minutes, and then high-pressure homogenized at 50 MPa twice, each time for 5 minutes, to obtain a nanoemulsion, which was freeze-dried to a water content of 3 wt% to obtain an anti-inflammatory composition.

[0051] Example 2

[0052] (1) Preparation of polyphenol complexes

[0053] 1) Preparation of clove extract

[0054] The clove buds were crushed through an 80-mesh sieve to obtain clove bud powder, which was mixed with a deep eutectic solvent (composed of choline chloride and urea in a molar ratio of 1:1.5) and water in a weight ratio of 1:18:2. The mixture was extracted with a high-voltage pulsed electric field at an electric field intensity of 12 kV / cm, a pulse width of 18 μs, a frequency of 1.5 Hz, and a temperature of 28°C for 160 s. The mixture was centrifuged at 2800 rpm for 7 min, and the supernatant was taken. The pH of the supernatant was adjusted to 5, and the mixture was adsorbed on a XAD-7HP macroporous resin column at a flow rate of 1.3 BV / h. The mixture was eluted with 2 BV of water (flow rate 1.6 BV / h), 2 BV of 43% ethanol solution by volume (flow rate 1.3 BV / h), and 4 BV of 67% ethanol solution by volume (flow rate 1.3 BV / h). The 67% ethanol eluate was collected and vacuum freeze-dried to a water content of 3 wt% to obtain a clove extract.

[0055] 2) Preparation of Sanguisorba officinalis extract

[0056] The root of Sanguisorba officinalis was crushed through a 100-mesh sieve to obtain Sanguisorba officinalis root powder, which was mixed with a 30% ethanol solution by volume at a solid-liquid ratio of 1:12 g / mL, and the pH value was adjusted to 3 with citric acid. The mixture was ultrasonically extracted at a power of 270 W, a frequency of 38 kHz, and 45°C for 50 min, and centrifuged at 3800 rpm for 12 min to obtain supernatant 1 and a precipitate. Anhydrous ethanol was added to the supernatant 1 to a volume fraction of 67%, and the mixture was precipitated at 4°C for 10 h. The precipitate was discarded to obtain supernatant 2. The precipitate was mixed with a low eutectic solvent (composed of choline chloride and lactic acid at a molar ratio of 1:1.5) and water at a weight ratio of 1:8:1. , ultrasonic extraction at 230W, frequency 33kHz, 38℃ for 35min, centrifugation at 3800rpm for 12min, obtaining supernatant 3, mixing with supernatant 2 and passing through a 0.45μm membrane, adjusting the pH to 4.5, and adsorbing through HPD-600 macroporous resin at a flow rate of 1BV / h, eluting with 4BV water (flow rate 1.8BV / h), 2BV volume fraction 33% ethanol solution (flow rate 1BV / h), and 4BV volume fraction 73% ethanol solution (flow rate 1BV / h), collecting the 73% volume fraction ethanol eluate, and vacuum freeze-drying to obtain the Sanguisorba officinalis extract.

[0057] 3) Preparation of polyphenol complexes

[0058] The clove extract and the sanguisorba officinalis extract are mixed in a weight ratio of 1:1 to obtain a polyphenol complex.

[0059] (2) Preparation of modified resistant dextrin

[0060] Resistant dextrin was mixed with water at 58°C and 300 rpm for 15 minutes to obtain a resistant dextrin solution with a mass fraction of 28%. The pH value was adjusted to 8.5, and octenylsuccinic anhydride was added (the addition amount was 3% of the weight of the resistant dextrin). After reacting at 58°C and 250 rpm for 7 hours, the pH value was adjusted to 6.3, and the mixture was centrifuged at 8000 rpm for 15 minutes to remove the precipitated insoluble matter. The supernatant was passed through a polysulfone membrane (molecular weight cutoff 3.5 kDa) to remove free octenylsuccinic anhydride, and the retained portion was dried to obtain modified resistant dextrin.

[0061] (3) Preparation of anti-inflammatory composition

[0062] Accurately weigh 20 parts of polyphenol complex, 30 parts of algae oil, 15 parts of modified resistant dextrin, 15 parts of trehalose, and 1 part of vitamin E succinate;

[0063] The polyphenol complex, algal oil and vitamin E succinate were mixed, mixed at 50°C and 200 rpm for 8 minutes, and then homogenized at 7000 rpm for 8 minutes to obtain an oil phase; the modified resistant dextrin, trehalose and water were mixed at 62°C and 300 rpm for 15 minutes to obtain an aqueous phase with a solid content of 18 wt%; the oil phase was added to the aqueous phase, first sheared at 8000 rpm for 6 minutes, and then homogenized at 45 MPa under high pressure for 12 minutes to obtain a nanoemulsion, which was freeze-dried to a water content of 4 wt% to obtain an anti-inflammatory composition.

[0064] Example 3

[0065] (1) Preparation of polyphenol complexes

[0066] 1) Preparation of clove extract

[0067] The clove buds were crushed through a 120-mesh sieve to obtain clove bud powder, which was then mixed with a deep eutectic solvent (composed of choline chloride and urea in a molar ratio of 1:2.5) and water in a weight ratio of 1:23:4. The mixture was extracted with a high-voltage pulsed electric field at an electric field intensity of 17 kV / cm, a pulse width of 22 μs, a frequency of 3 Hz, and a temperature of 32°C for 100 s. The mixture was centrifuged at 3200 rpm for 3 min, and the supernatant was taken. The pH of the supernatant was adjusted to 6, and the mixture was adsorbed on a XAD-7HP macroporous resin column at a flow rate of 1.8 BV / h. The mixture was eluted with 4 BV of water (flow rate 2.3 BV / h), 4 BV of 48% ethanol solution (flow rate 1.8 BV / h), and 6 BV of 73% ethanol solution (flow rate 1.8 BV / h). The 73% ethanol eluate was collected and vacuum freeze-dried to a water content of 4 wt% to obtain a clove extract.

[0068] 2) Preparation of Sanguisorba officinalis extract

[0069] The root of Sanguisorba officinalis was crushed through a 150-mesh sieve to obtain Sanguisorba officinalis root powder, which was mixed with a 40% ethanol solution by volume at a solid-liquid ratio of 1:18 g / mL, and the pH value was adjusted to 4 (with citric acid, and ultrasonically extracted at a power of 320 W, a frequency of 42 kHz, and 50° C. for 40 min, and centrifuged at 4200 rpm for 8 min to obtain supernatant 1 and a precipitate; anhydrous ethanol was added to the supernatant 1 to a volume fraction of 73%, and the mixture was precipitated at 4° C. for 14 h, and the precipitate was discarded to obtain supernatant 2; the precipitate was mixed with a deep eutectic solvent (composed of choline chloride and lactic acid at a molar ratio of 1:2.5) and water at a weight ratio of 1:12:2, and the mixture was stirred at room temperature for 2 hours. Ultrasonic extraction was performed at 280 W, a frequency of 37 kHz, and 42° C. for 25 min, and the supernatant was centrifuged at 4200 rpm for 8 min to obtain supernatant 3. The supernatant 3 was mixed with supernatant 2 and passed through a 0.45 μm membrane. The pH was adjusted to 5.5, and the supernatant was adsorbed on a HPD-600 macroporous resin at a flow rate of 1.5 BV / h. The supernatant was eluted in sequence with 6 BV of water (flow rate 2.3 BV / h), 4 BV of 38% ethanol solution (flow rate 1.5 BV / h), and 6 BV of 78% ethanol solution (flow rate 1.5 BV / h). The 78% ethanol eluate was collected and freeze-dried in vacuo to obtain the Sanguisorba officinalis extract.

[0070] 3) Preparation of polyphenol complexes

[0071] The clove extract and the sanguisorba officinalis extract were mixed in a weight ratio of 3:1 to obtain a polyphenol complex.

[0072] (2) Preparation of modified resistant dextrin

[0073] Resistant dextrin was mixed with water at 62°C and 500 rpm for 10 minutes to obtain a resistant dextrin solution with a mass fraction of 32%. The pH value was adjusted to 9, and octenylsuccinic anhydride was added (the addition amount was 8% of the weight of the resistant dextrin). After reacting at 62°C and 350 rpm for 6 hours, the pH value was adjusted to 6.8, and the mixture was centrifuged at 10,000 rpm for 10 minutes to remove the precipitated insoluble matter. The supernatant was passed through a polysulfone membrane (molecular weight cutoff 3.5 kDa) to remove free octenylsuccinic anhydride, and the retained portion was dried to obtain modified resistant dextrin.

[0074] (3) Preparation of anti-inflammatory composition

[0075] Accurately weigh 30 parts of polyphenol complex, 40 parts of algae oil, 25 parts of modified resistant dextrin, 25 parts of trehalose, and 3 parts of vitamin E succinate;

[0076] The polyphenol complex, algal oil and vitamin E succinate were mixed, mixed at 60°C and 300 rpm for 3 minutes, and then homogenized at 9000 rpm for 3 minutes to obtain an oil phase; the modified resistant dextrin, trehalose and water were mixed at 62°C and 450 rpm for 10 minutes to obtain an aqueous phase with a solid content of 23 wt%; the oil phase was added to the aqueous phase, first sheared at 12000 rpm for 4 minutes, and then high-pressure homogenized at 55 MPa for 3 times, each time for 5 minutes to obtain a nanoemulsion, which was freeze-dried to a water content of 3 wt% to obtain an anti-inflammatory composition.

[0077] Comparative Example 1

[0078] The specific implementation is the same as that of Example 1, except that "25 parts of polyphenol complex and 35 parts of algae oil" in step (3) are replaced by "60 parts of polyphenol complex".

[0079] Comparative Example 2

[0080] The specific implementation method is the same as that of Example 1, except that "25 parts of polyphenol complex and 35 parts of algae oil" in step (3) is replaced by "60 parts of algae oil" and step (1) is deleted.

[0081] Comparative Example 3

[0082] The specific implementation is the same as that of Example 1, except that the "modified resistant dextrin" in step (3) is replaced by "resistant dextrin".

[0083] Comparative Example 4

[0084] The specific implementation method is the same as that of Example 1, except that "20 parts of modified resistant dextrin and 20 parts of trehalose" in step (3) is replaced by "10 parts of modified resistant dextrin and 30 parts of trehalose".

[0085] Comparative Example 5

[0086] The specific implementation method is the same as that of Example 1, except that "20 parts of modified resistant dextrin and 20 parts of trehalose" in step (3) is replaced by "30 parts of modified resistant dextrin and 10 parts of trehalose".

[0087] Comparative Example 6

[0088] The specific implementation is the same as that of Example 1, except that in step (1), "mixed with a deep eutectic solvent (composed of choline chloride and urea in a molar ratio of 1:2) and water in a weight ratio of 1:20:3" is replaced by "mixed with a 70% by volume ethanol solution in a weight ratio of 1:23".

[0089] Comparative Example 7

[0090] The specific implementation method is the same as that of Example 1, except that in step (1), 2) the Sanguisorba officinalis extract is prepared as follows: the Sanguisorba officinalis root is crushed through a 120-mesh sieve to obtain Sanguisorba officinalis root powder, which is mixed with a 75% volume fraction ethanol solution at a solid-liquid ratio of 1:15 g / mL, and ultrasonically extracted at a power of 300 W, a frequency of 40 kHz, and 48°C for 45 min, followed by centrifugation at 4000 rpm for 10 min. The supernatant is passed through a 0.45 μm membrane, the pH is adjusted to 5, and adsorbed on an HPD-600 macroporous resin at a flow rate of 1.3 BV / h. The mixture is eluted in sequence with 5 BV of water (flow rate 3 BV / h), 3 BV of a 35% volume fraction ethanol solution (flow rate 1.3 BV / h), and 5 BV of a 75% volume fraction ethanol solution (flow rate 1.3 BV / h). The 75% volume fraction ethanol eluate is collected and freeze-dried in vacuo to obtain the Sanguisorba officinalis extract.

[0091] Test Example 1

[0092] Anti-inflammatory animal experiments

[0093] (1) Mouse ear swelling model

[0094] SPF male 18-20 g BALB / c mice were selected and, after adaptive feeding for 7 days, were evenly divided into a model group, a positive control group, and 10 experimental groups (Example 1 group, Example 2 group, Example 3 group, Comparative Example 1 group, Comparative Example 2 group, Comparative Example 3 group, Comparative Example 4 group, Comparative Example 5 group, Comparative Example 6 group, and Comparative Example 7 group, 10 mice in each group).

[0095] The experimental group was orally administered with the corresponding anti-inflammatory composition at a dose of 50 mg / kg, the model group was orally administered with an equal volume of normal saline, and the positive control group was intraperitoneally injected with dexamethasone at a dose of 5 mg / kg for 7 consecutive days.

[0096] One hour after the last dose on day 7, 20 μL of xylene was evenly applied to both sides of the right ear of the mice. The left ear was left untreated (self-control). One hour after xylene application, the mice were sacrificed by cervical dislocation. The left and right ear pieces were removed using a 6 mm punch and weighed separately. Ear swelling (%) was calculated as (right ear weight - left ear weight) / left ear weight × 100%.

[0097] The right ear tissues of mice in each group were added into RIPA lysis buffer (50 mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, 1 mM EDTA, 1× protease inhibitor) at a weight ratio of 1:9. The cells were mechanically homogenized on ice at 1,000 rpm for 3 times, each time for 10 s. The cells were centrifuged at 4°C and 12,000 × g for 15 min, and the precipitate was discarded. The TNF-α and IL-6 levels in the supernatant of each group were determined using ELISA kits.

[0098] The specific results of ear swelling and the contents of TNF-α and IL-6 in ear tissues of each group are shown in Table 1.

[0099] Table 1 Ear swelling and TNF-α and IL-6 levels in mice in each experimental group

[0100]

[0101]

[0102] The results in Table 1 show that the ear swelling of mice in the model group was high, indicating that xylene successfully induced an acute inflammatory model. In the positive control group (dexamethasone), ear swelling was reduced to 28.43%, and TNF-α and IL-6 levels were reduced, respectively, verifying the reliability of the experimental system. The ear swelling, TNF-α, and IL-6 levels of Examples 1-3 were similar to those of the positive control group, indicating that the core formula of the polyphenol-algae oil compound of the present invention can effectively inhibit acute inflammatory reactions and the release of inflammatory factors. The ear swelling and inflammatory factor levels of Comparative Examples 1 and 2 were significantly higher than those of the Example group, demonstrating that the combination of polyphenols and algae oil has a synergistic anti-inflammatory effect, while the effect of a single ingredient is less effective. The effects of Comparative Examples 3-5 were weaker than those of the Example group, indicating that the structure of the modified resistant dextrin and its ratio with trehalose are key to maintaining the stability and anti-inflammatory activity of the composition. The inflammatory indicators of Comparative Examples 6-7 were higher than those of the Example group, indicating that the optimized preparation process of the polyphenol extract (such as deep eutectic solvent-assisted extraction and step-by-step purification) can improve the retention rate of active ingredients and enhance the anti-inflammatory effect.

[0103] (2) Mouse colitis model

[0104] SPF male 18-20 g BALB / c mice were selected and, after adaptive feeding for 7 days, were evenly divided into a negative group, a model group, a positive group, and 10 experimental groups (Example 1 group, Example 2 group, Example 3 group, Comparative Example 1 group, Comparative Example 2 group, Comparative Example 3 group, Comparative Example 4 group, Comparative Example 5 group, Comparative Example 6 group, and Comparative Example 7 group, 10 mice in each group).

[0105] Except for the negative group, dextran sulfate sodium was added to the drinking water to a 3% mass fraction (to induce acute colitis). The negative group received regular water for 10 consecutive days. Starting on the fourth day of drinking, the experimental group received a 50 mg / kg dose of the corresponding anti-inflammatory composition by gavage, the model group received an equal volume of normal saline, and the positive group received a 50 mg / kg dose of sulfasalazine by gavage for 7 consecutive days.

[0106] On the 7th day after oral administration, the disease activity index was tested. The DAI scoring standard is shown in Table 2.

[0107] After the last administration, the mice were fasted for 12 h before being sacrificed, and the colon was separated from the cecum to the anus, and the colon length was measured.

[0108] 1 cm of the distal colon was obtained, fixed with 4% paraformaldehyde, embedded in paraffin, and stained with HE. Pathological scoring criteria (0-12 points) included: inflammatory infiltration (0-3), crypt destruction (0-3), ulcer extent (0-3), and edema (0-3).

[0109] The colon of each mouse group was added to RIPA lysis buffer at a weight ratio of 1:9, and mechanically homogenized on ice at 1000 rpm for 3 times, each time for 10 s. The mixture was centrifuged at 4°C and 12000×g for 15 min, and the precipitate was discarded. The levels of TNF-α and IL-6 in the supernatant of each group were determined using ELISA kits.

[0110] The DAI scores, colon lengths, pathological scores, and TNF-α and IL-6 levels of mice in each group are shown in Table 3 .

[0111] Table 2 Disease Activity Index (DAI) scoring criteria

[0112] Weight loss (%) blood in the stool Stool characteristics score none Negative normal 0 1-5％ Occult blood positive soft stools 1 5-10％ Visible blood in stool diarrhea 2 >10％ severe bleeding bloody diarrhea 3

[0113] Table 3 Experimental results of mice in each group

[0114]

[0115]

[0116] Table 3 data show that the model group mice DAI score is high, colon length is significantly shortened, pathological score is high, TNF-α and IL-6 content are significantly increased, indicating that sodium dextran sulfate successfully induced acute colitis model. The positive control group (sulfasalazine) DAI score dropped to 1.0, colon length returned to 7.02cm, pathological score 3.2, and inflammatory factor level was significantly reduced, verifying the reliability of the experimental system. The DAI score, colon length, pathological score, TNF-α and IL-6 content of Example 1-3 group were close to the positive control group effect, indicating that the composition can effectively alleviate colitis symptoms, reduce intestinal inflammatory damage and inhibit the release of inflammatory factors. The DAI score, colon length, pathological score and inflammatory factor level of Comparative Example 1-2 were significantly worse than those of the Example group, proving again that the composite synergistic effect of polyphenols and algae oil is crucial to the improvement of chronic inflammation. The intestinal damage index of Comparative Example 3-5 (carrier component change) was better than that of the single component group, but weaker than that of the Example group, indicating that the optimized ratio of modified resistant dextrin and trehalose can enhance the intestinal targeting and stability of the composition and enhance the anti-inflammatory effect. The inflammatory indicators of Comparative Examples 6-7 (extract process changes) were better than those of the other comparative examples, but still worse than those of the embodiment group, indicating that the optimization of the preparation process of the extract can improve the bioavailability of the active ingredients and enhance the intervention effect on chronic intestinal inflammation.

[0117] In summary, the polyphenol compound algae oil anti-inflammatory composition of the present invention has significant anti-inflammatory activity to acute inflammation (ear swelling) and chronic inflammation (colitis), and the effect is close to that of positive drugs (dexamethasone, sulfasalazine). Experiments show that the compounding of the polyphenol complex of the present invention and algae oil, the optimized ratio of modified resistant dextrin and trehalose, and the deep eutectic solvent-assisted extraction process of the polyphenol extract are the key technical features to ensure the anti-inflammatory effect of the composition, and the change of any link (such as single component replacement, process simplification, carrier ratio deviation) will cause the anti-inflammatory activity to decline. Experimental data confirm that the composition plays an anti-inflammatory role by suppressing the release of inflammatory factors (TNF-α, IL-6) and alleviating tissue damage, and has good application potential.

[0118] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. An anti-inflammatory composition of polyphenols compounded with algae oil, characterized in that: The invention comprises the following raw materials in parts by weight: 20-30 parts of polyphenol complex, 30-40 parts of algae oil, 15-25 parts of modified resistant dextrin, 15-25 parts of trehalose and 1-3 parts of vitamin E succinate; The polyphenol complex consists of clove extract and sanguisorba officinalis extract in a weight ratio of 1-3:

1.

2. The anti-inflammatory composition according to claim 1, characterized in that The preparation method of the clove extract comprises: mixing clove buds, a low eutectic solvent and water, extracting the mixture through a high-voltage pulse electric field, centrifuging the mixture, and purifying the supernatant through a macroporous resin to obtain the clove extract.

3. The anti-inflammatory composition according to claim 2, characterized in that The deep eutectic solvent is composed of choline chloride and urea in a molar ratio of 1:1.5-2.5, and the weight ratio of the clove buds, the deep eutectic solvent and water is 1:(18-23):(2-4).

4. The anti-inflammatory composition according to claim 1, characterized in that The preparation method of the Sanguisorba officinalis extract comprises: mixing Sanguisorba officinalis roots with a 30%-40% ethanol solution by volume, subjecting the mixture to ultrasonic treatment for 40-50 minutes, centrifuging, mixing a precipitate, a low eutectic solvent and water, subjecting the mixture to ultrasonic extraction, centrifuging, and purifying the supernatant with a macroporous resin to obtain the Sanguisorba officinalis extract.

5. The anti-inflammatory composition according to claim 4, characterized in that The eutectic solvent is composed of choline chloride and lactic acid in a molar ratio of 1:1.5-2.5, and the weight ratio of the precipitate, the deep eutectic solvent and water is 1:8-12:1-2.

6. The anti-inflammatory composition according to claim 1, characterized in that The preparation method of the modified resistant dextrin preferably comprises: mixing resistant dextrin with water, adjusting the pH value to 8.5-9, adding octenyl succinic anhydride for reaction, and centrifuging to precipitate the modified resistant dextrin.

7. The anti-inflammatory composition according to claim 6, characterized in that The reaction temperature is 58-62° C., the rotation speed is 250-350 rpm, and the reaction time is 6-7 h.

8. A method for preparing the anti-inflammatory composition according to any one of claims 1 to 7, comprising: mixing the polyphenol complex, algae oil, and vitamin E succinate to obtain an oil phase; mixing the modified resistant dextrin, trehalose and water to obtain an aqueous phase; The oil phase is added to the water phase, homogenized to obtain a nanoemulsion, and freeze-dried to obtain an anti-inflammatory composition.

9. The anti-inflammatory composition according to claim 8, characterized in that The homogenization includes: first shearing at 8000-12000 rpm for 4-6 minutes, and then high-pressure homogenization at 45-55 MPa for 8-12 minutes.

10. Use of the anti-inflammatory composition according to any one of claims 1 to 7 in the preparation of anti-inflammatory drugs.

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