Astaxanthin soft capsule with hypoglycemic function and preparation process thereof

Abstract

The application provides astaxanthin soft capsules with a blood sugar lowering function and a preparation process thereof, and belongs to the technical field of soft capsule preparation; the preparation process comprises the following steps: preparation of mulberry leaf extracellular vesicles; preparation of N-succinyl chitosan; liposome embedding astaxanthin; preparation of astaxanthin soft capsules. The application adopts astaxanthin and allicin compounding, polyethylene glycol is used to modify bovine serum albumin to wrap the liposome of astaxanthin and allicin, and then N-succinyl chitosan / mulberry leaf extracellular vesicles are used for modification, and the synergistic effect of the two can make the liposome embedding astaxanthin compound after N-succinyl chitosan / mulberry leaf extracellular vesicle modification have better targeting and biocompatibility, so as to improve the bioavailability of astaxanthin and improve the blood sugar lowering effect.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of soft capsule preparation, in particular to a soft capsule with blood glucose-lowering function and its preparation process. BACKGROUND

[0002] With the improvement of people's living standards and the change of lifestyle, the incidence of diabetes is increasing year by year, which has become a global public health problem that seriously threatens human health. Diabetes not only causes blood glucose metabolism disorder, but also causes a series of serious complications such as cardiovascular disease, neuropathy, retinopathy, etc., which greatly affects the quality of life and health of patients.

[0003] At present, there are many types of drugs for treating diabetes in clinical practice, mainly including insulin and its analogues, oral hypoglycemic drugs, etc. However, these drugs often have different degrees of side effects in the long-term use process, such as hypoglycemia, weight gain, gastrointestinal discomfort, liver and kidney function damage, etc. Some patients may also have drug resistance, leading to gradually decreased treatment effect. Therefore, developing safe and effective blood glucose-lowering functional products, especially those derived from natural products and having multiple health functions, has become a research hotspot in the field of diabetes prevention and treatment.

[0004] Astaxanthin is a kind of carotenoid with strong antioxidant activity, which exists widely in algae, shrimps, crabs, salmon and other organisms. In recent years, a large number of studies have shown that astaxanthin not only has multiple physiological functions such as antioxidant, anti-inflammatory, anti-fatigue, etc., but also shows potential effects in regulating blood glucose metabolism. Astaxanthin can improve insulin sensitivity, inhibit oxidative stress and inflammatory response, etc., and has certain prevention and improvement effects on diabetes and its complications.

[0005] However, astaxanthin has the problem of low bioavailability. On the one hand, astaxanthin has lipophilicity and very low solubility in water, which makes it difficult to dissolve and disperse in the gastrointestinal tract, thereby affecting absorption. On the other hand, astaxanthin is easily oxidized in the body, and its chemical structure is easily changed in the process of digestion in the gastrointestinal tract and metabolism in the liver, leading to reduced activity and difficulty in being completely absorbed and utilized by the human body.

[0006] Therefore, we propose a soft capsule with blood glucose-lowering function and its preparation process which can improve the bioavailability of astaxanthin. SUMMARY

[0007] In view of the deficiencies in the prior art, the purpose of the present application is to provide a soft capsule with blood glucose-lowering function and its preparation process.

[0008] The preparation process of a soft capsule with blood glucose-lowering function comprises the following steps:

[0009] S1: Preparation of mulberry leaf extracellular vesicles

[0010] Fresh mulberry leaves are added to an enzyme solution, followed by high-voltage pulsed electric field treatment and enzymolysis in the dark, centrifugation, and filtration to obtain mulberry leaf extracellular vesicles;

[0011] S2: Preparation of N-succinyl chitosan

[0012] After mixing deacetylated chitosan and dimethyl sulfoxide, adding succinic anhydride to obtain a precipitate, adjusting the pH after adding the precipitate to deionized water, then adding acetone, centrifuging and washing again, drying and grinding, N-succinyl chitosan is obtained;

[0013] S3: Liposome-embedded astaxanthin

[0014] First, phospholipid, cholesterol and astaxanthin are used to prepare a liposome suspension, then polyethylene glycol is used to modify bovine serum albumin to coat the liposomes, and finally N-succinyl chitosan and mulberry leaf extracellular vesicles are used for modification and coating to obtain N-succinyl chitosan / mulberry leaf extracellular vesicle modified liposome-embedded astaxanthin complex;

[0015] S4: Preparation of astaxanthin soft capsules

[0016] Low-pressure plasma is used to modify dried hydroxypropyl starch, then modified hydroxypropyl starch, glycerol, deionized water and agar are used to prepare a glue solution, and N-succinyl chitosan / mulberry leaf extracellular vesicle modified liposome-embedded astaxanthin complex is used as the capsule content and glue solution into the pill machine to prepare astaxanthin soft capsules.

[0017] Further, step S1 for preparing mulberry leaf extracellular vesicles, specifically comprising the following steps:

[0018] S1.1: Mix 2-3 parts by weight of cellulase, 1-2 parts by weight of pectinase and 1-2 parts by weight of β-glucosidase, then add the enzyme solution to a pH 4.5-5.5 acetic acid-sodium acetate buffer to adjust the pH of the enzyme solution to 4.8-5.2 to obtain the enzyme solution;

[0019] S1.2: Wash fresh mulberry leaves, crush to 20-30 mesh, add to the enzyme solution, then perform high-voltage pulsed electric field treatment at 20-30 kV / cm, 3-4 pulses for 1-2 h in the dark, then enzymolysis at 25-28℃, 200-300 centrifugal force for 10-12 h, stand, centrifuge the upper liquid at 100000-120000 centrifugal force for 150-180 min, discard the precipitate, then centrifuge at 12000-15000 centrifugal force for 2-3 times, 10-12 min each time, combine the two supernatants and reserve for use;

[0020] S1.3: The supernatant combined twice is subjected to ultracentrifugation at 150000-180000 centrifugal force at 4-5℃ for 100-120min using a sucrose pad, and the sucrose layer is collected, washed with a phosphate buffer solution, and subjected to ultrafiltration centrifugation to obtain the mulberry leaf extracellular vesicles.

[0021] Further, the ratio of fresh mulberry leaves to enzyme solution in step S1.2 is 5-8g:180mL.

[0022] Further, the preparation of N-succinyl chitosan in step S2 specifically includes the following steps:

[0023] S2.1: 2-3 parts by weight of deacetylated chitosan and 40-50 parts by weight of dimethyl sulfoxide are mixed, 2-3 parts by weight of succinic anhydride is added, and after sealing, stirring is performed at 60-65℃ for 6-8h, then 2-3 times the volume of ethanol is added, the sample is soaked for 2-3h, and then centrifugation is performed at 3500-4000 centrifugal force for 5-10min to obtain a precipitate;

[0024] S2.2: The precipitate is added to 20-30 parts by weight of deionized water, then a sodium hydroxide solution with a concentration of 5-8% is added to adjust the pH to 10-11, then 3-4 times the volume of acetone is added, and then centrifugation is performed at 3500-4000 centrifugal force for 5-10min, i.e., the obtained precipitate is washed with 70% ethanol for 1-2 times and pure acetone for 3-4 times, the washed precipitate is dried at 40-50℃ for 24-25h, and then ground to obtain N-succinyl chitosan.

[0025] Further, the step S3 of embedding astaxanthin in liposomes specifically includes the following steps:

[0026] S3.1: 20-22 parts by weight of phospholipid, 5-8 parts by weight of cholesterol, 4-5 parts by weight of astaxanthin, and 1-2 parts by weight of garlicin are fully dissolved in 50-80 parts by weight of chloroform, and then rotary evaporation is performed on a rotary evaporator to obtain a lipid film, 10-20 parts by weight of the lipid film is added to 30-50 parts by weight of deionized water and vortexed vigorously for 10-12min, then homogenized at 60-63℃ for 15-20min, and then ultrasonic cell disruption is performed on ice at 25-28℃ using a probe ultrasonic cell disruptor at an intensity of 70% for 10-12min to obtain a liposome suspension;

[0027] S3.2: 2-3 parts by weight of bovine serum albumin and 1-2 parts by weight of polyethylene glycol methyl ether-succinimidyl ester are dissolved in 30-40 parts by weight of a PBS solution with a pH of 7.4, and after stirring and reacting at room temperature for 2-3h, the reaction solution is transferred to a dialysis bag and dialyzed in pure water for 48-50h, and then the dialysate is freeze-dried to obtain polyethylene glycol-modified bovine serum albumin.

[0028] S3.3: Polyethylene glycol modified bovine serum albumin is added to deionized water, and then stirred and mixed at 200-300 r / min for 20-30 min to obtain a 10% w / v polyethylene glycol modified bovine serum albumin solution. N-succinyl chitosan is dissolved in deionized water, stirred and mixed for 20-30 min to obtain a 0.5-1% w / v N-succinyl chitosan solution, and then 1-2% mulberry leaf extracellular vesicles are added and mixed to obtain a mixed solution;

[0029] S3.4: The liposome suspension is added to an equal volume of 10% w / v polyethylene glycol modified bovine serum albumin solution, and then incubated at 60-65°C for 60-80 min. Then add to 2-3 times the volume of the mixed solution, incubate at 60-65°C for 20-30 min to obtain N-succinyl chitosan / mulberry leaf extracellular vesicle modified liposome-embedded astaxanthin complex.

[0030] Further, the molecular weight of the polyethylene glycol methyl ether-succinimidyl ester in step S3.2 is 2000.

[0031] Further, the dialysis bag in step S3.2 is a dialysis bag with a molecular weight cut-off of 8000-14000 Da.

[0032] Further, the preparation of astaxanthin soft capsules in step S4 specifically includes the following steps:

[0033] S4.1: 20-30 parts by weight of hydroxypropyl starch is added to 45-50 parts by weight of distilled water, stirred and mixed, and then placed in a dry box at 28-30°C and dried to a water content of 1%. The dried hydroxypropyl starch is placed between the two electrodes of a plasma generator for plasma treatment to obtain modified hydroxypropyl starch.

[0034] S4.2: 28-30 parts by weight of modified hydroxypropyl starch, 20-22 parts by weight of glycerol, and 50-52 parts by weight of deionized water are mixed, and then stirred and mixed at 95-98°C for 30-50 min. Then add 2-3 parts by weight of agar, and then stir and mix again at 95-98°C for 30-50 min. Finally, incubate for 2-3 h to obtain a glue solution.

[0035] S4.3: The N-succinyl chitosan / mulberry leaf extracellular vesicle modified liposome-embedded astaxanthin complex is used as the core content of the capsule and the glue solution is fed into the pill making machine. Pressing, shaping, drying, sorting and packaging are performed to obtain astaxanthin soft capsules.

[0036] Further, in step S4.1, argon is used for plasma treatment at 300-320 W for 4-5 min.

[0037] The astaxanthin soft capsule with the blood sugar lowering function is prepared by the preparation process of the astaxanthin soft capsule with the blood sugar lowering function.

[0038] Compared with the prior art, the present application has at least the following beneficial effects:

[0039] 1、The liposome of the present application is prepared by polyethylene glycol modified bovine serum albumin coated astaxanthin and allicin, which increases the stability and blood circulation time of the liposome, and these components have relatively high biological safety, and the polyethylene glycol modified bovine serum albumin coated liposome can reduce the recognition and removal of the liposome by the immune system, prolong the action time in the body, thereby facilitating absorption and improving bioavailability, and then N-succinyl chitosan / mulberry leaf extracellular vesicle modification can produce synergistic effect, N-succinyl chitosan can realize covalent binding with the specific expression protein ligand of intestinal epithelial cells through amidation reaction, so as to accurately target the intestinal epithelial cells, and the membrane structure of the mulberry leaf extracellular vesicle is similar to that of the cell membrane, which can better fuse with the cell and promote the components such as astaxanthin to enter the cell to play a role, thereby enhancing the regulation function on blood glucose metabolism related cells, and the synergistic effect of the two can make the N-succinyl chitosan / mulberry leaf extracellular vesicle modified liposome embedded astaxanthin complex have better targeting and biocompatibility, thereby improving the bioavailability of astaxanthin and the blood sugar lowering effect.

[0040] 2、The active ingredients in the capsule contain astaxanthin and allicin, astaxanthin itself has various physiological activities such as antioxidant and anti-inflammatory, and can reduce blood sugar through the pathways of regulating insulin signaling pathway and improving pancreatic beta cell function, allicin also has certain blood sugar lowering effect, it can promote the secretion of insulin and improve the uptake and utilization of glucose by tissues, at the same time, the mulberry leaf extracellular vesicle contains various components with blood sugar lowering activity, such as flavones and polysaccharides, which synergistically act with astaxanthin and allicin to further enhance the blood sugar lowering effect.

[0041] 3. This invention utilizes argon plasma treatment to induce cross-linking reactions between hydroxypropyl starch molecules. The argon plasma contains numerous high-energy particles and active groups, which interact with the hydroxypropyl starch molecules, triggering the formation of new chemical bonds between the molecular chains. This allows the modified hydroxypropyl starch to construct a more stable and compact three-dimensional network structure when forming capsule shells, resulting in capsule shells with better mechanical strength and stability, making them less prone to breakage and better protecting the capsule contents during storage and transportation. Furthermore, after plasma treatment, tiny gaps are formed between the molecular chains of hydroxypropyl starch, allowing water molecules to more easily enter these gaps. When the capsule enters the gastrointestinal tract, a large number of water molecules are attracted to and bind to the starch, accelerating the water absorption process. This allows the capsule shell to expand and disintegrate rapidly, releasing the capsule contents more quickly and increasing the dissolution rate of astaxanthin in the gastrointestinal tract, thus facilitating its absorption in the body and better exerting its physiological effects such as lowering blood sugar. Attached Figure Description

[0042] The accompanying drawings, which are incorporated herein and form part of the specification, illustrate embodiments of the invention and, together with the specification, further serve to explain the principles of the invention and enable those skilled in the art to practice and use the invention.

[0043] Figure 1 This is a flowchart illustrating the preparation process of an astaxanthin soft capsule with hypoglycemic function used in an embodiment of the present invention. Detailed Implementation

[0044] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0045] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Every smaller range between any stated value or intermediate value within a stated range, and any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0046] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar to or equivalent to those described herein may be used in the implementation or testing of this invention.

[0047] Many modifications and variations of the specific embodiments of the application can be made without departing from the scope or spirit of the application. Other embodiments of the application will be apparent to those skilled in the art from consideration of the specification and practice of the application. The specification and examples are illustrative only.

[0048] As used herein, "comprise", "comprising", "containing", "include", "including", "have" and "having", "contain", "containing", "characterized by" and the like are open-ended language that means "including but not limited to", and the numerical order of S1-S4 and the like in the steps of the present application are only used to distinguish the preparation methods of different materials, and are not used to limit the sequence of the steps.

[0049] Example 1: Preparation process of astaxanthin soft capsules with hypoglycemic function, as shown in Figure 1 The preparation process comprises the following steps:

[0050] S1: Preparation of mulberry leaf extracellular vesicles

[0051] S1.1: Mix 2 parts by weight of cellulase, 1 part by weight of pectinase and 1 part by weight of β-glucosidase, then add the enzyme solution to the acetic acid-sodium acetate buffer with pH value of 4.5 to adjust the pH value of the enzyme solution to 4.8, and obtain the enzyme solution;

[0052] S1.2: Wash fresh mulberry leaves, crush to 20 mesh, and then add the enzyme solution according to the solid-liquid ratio of 5g:180mL, then perform high-voltage pulsed electric field treatment under dark conditions at 20kV / cm for 3 pulses for 1h, then perform enzymolysis at 25℃ for 10h under 200 centrifugal force, stand still, centrifuge the upper liquid at 100000 centrifugal force for 150min, discard the precipitate, then centrifuge twice at 12000 centrifugal force for 10min each time, combine the two supernatants, and reserve for use;

[0053] S1.3: The combined supernatant is subjected to ultracentrifugation at 4℃ under 150000 centrifugal force for 100min using sucrose pad, the sucrose layer is washed with phosphate buffer, and the mulberry leaf extracellular vesicles are obtained after ultrafiltration centrifugation of the ultrafiltration membrane;

[0054] S2: Preparation of N-succinyl chitosan

[0055] S2.1: Mix 2 parts by weight of deacetylated chitosan and 40 parts by weight of dimethyl sulfoxide, then add 2 parts by weight of succinic anhydride, seal and stir at 60℃ for 6h, then soak the sample in 2 times the volume of ethanol for 2h, and then centrifuge at 3500 centrifugal force for 5min to obtain the precipitate;

[0056] S2.2: The precipitate was added to 20 parts by weight of deionized water, then a 5% concentration of sodium hydroxide solution was added to adjust the pH to 10, 3 times the volume of acetone was added, and then centrifuged at 3500 centrifugal force for 5 min, i.e. the obtained precipitate was washed once with 70% ethanol and 3 times with pure acetone, and the washed precipitate was dried at 40°C for 24 h, and then ground to obtain N-succinyl chitosan;

[0057] S3: Liposome-embedded astaxanthin

[0058] S3.1: 20 parts by weight of phospholipid, 5 parts by weight of cholesterol, 4 parts by weight of astaxanthin and 1 part by weight of allicin were dissolved in 50 parts by weight of chloroform, and then rotary evaporation was performed on a rotary evaporator to obtain a lipid film, 10 parts by weight of the lipid film was added to 30 parts by weight of deionized water and vortexed vigorously for 10 min, then homogenized at 60°C for 15 min, and then ultrasonicated at 70% intensity on ice for 10 min using a probe ultrasonic cell disruptor at 25°C to obtain a liposome suspension;

[0059] S3.2: 2 parts by weight of bovine serum albumin and 1 part by weight of polyethylene glycol methyl ether-succinimidyl ester with a molecular weight of 2000 were dissolved in 30 parts by weight of PBS solution with pH 7.4, and after stirring and reacting at room temperature for 2 h, the reaction solution was transferred to a dialysis bag with a molecular weight cutoff of 8000 Da, and dialysis purification was performed in pure water for 48 h, and the dialysate was freeze-dried to obtain polyethylene glycol-modified bovine serum albumin;

[0060] S3.3: The polyethylene glycol-modified bovine serum albumin was added to deionized water, and then stirred and mixed at 200 r / min for 20 min to obtain a 10% w / v polyethylene glycol-modified bovine serum albumin solution, N-succinyl chitosan was dissolved in deionized water and stirred and mixed for 20 min to obtain a 0.5% w / v N-succinyl chitosan solution, and then 1% mulberry leaf extracellular vesicles were added and mixed to obtain a mixed solution;

[0061] S3.4: The liposome suspension was added to an equal volume of 10% w / v polyethylene glycol-modified bovine serum albumin solution, and then incubated at 60°C for 60 min, and then added to 2 times the volume of the mixed solution and incubated at 60°C for 20 min to obtain N-succinyl chitosan / mulberry leaf extracellular vesicle-modified liposome-embedded astaxanthin complex;

[0062] S4: Preparation of astaxanthin soft capsules

[0063] S4.1: 20 parts by weight of hydroxypropyl starch was added to 45 parts by weight of distilled water, after stirring and mixing, it was put into a drying oven at 28℃, and dried to a water content of 1%, then the dried hydroxypropyl starch was put between the two electrodes of a plasma generator, and treated at 300W for 4min using argon to obtain modified hydroxypropyl starch;

[0064] S4.2: 28 parts by weight of modified hydroxypropyl starch, 20 parts by weight of glycerol and 50 parts by weight of deionized water were mixed, then stirred and mixed at 95℃ for 30min, then 2 parts by weight of agar was added, and stirred and mixed again at 95℃ for 30min, and finally incubated for 2h to obtain a glue solution;

[0065] S4.3: The N-succinyl chitosan / silkworm leaf extracellular vesicle modified liposome-embedded astaxanthin complex was used as the core content and the glue solution into the pill making machine, and the pill was pressed, shaped, dried, sorted and packaged to obtain astaxanthin soft capsules with a loading capacity of 500mg / pill and a glue skin thickness of 0.85mm.

[0066] Example 2: A preparation process of astaxanthin soft capsules with hypoglycemic function, as shown in Figure 1 , comprising the following steps:

[0067] S1: Preparation of silkworm leaf extracellular vesicle

[0068] S1.1: 2 parts by weight of cellulase, 1 part by weight of pectinase and 1 part by weight of β-glucosidase were mixed, then added to a pH 4.5 acetic acid-sodium acetate buffer to adjust the pH of the enzyme solution to 4.8 to obtain an enzyme solution;

[0069] S1.2: Fresh silkworm leaves were washed, crushed to 30 mesh, and then added to the enzyme solution according to the solid-liquid ratio of 5g:180mL, then treated by high-voltage pulsed electric field under dark conditions at 30kV / cm and 4 pulses for 2h, then enzymolyzed at 28℃ and 300 centrifugal force for 12h, and then centrifuged at 120000 centrifugal force for 180min, and the precipitate was discarded, then centrifuged at 15000 centrifugal force for 3 times, each time for 12min, and then the supernatant was combined and reserved;

[0070] S1.3: The combined supernatant was ultracentrifuged at 5℃, 180000 centrifugal force using sucrose pad, and then centrifuged at 120min, and then washed with phosphate buffer, and then ultrafiltered by ultrafiltration membrane to obtain silkworm leaf extracellular vesicles;

[0071] S2: Preparation of N-succinyl chitosan

[0072] S2.1: 2 parts by weight of chitosan and 40 parts by weight of dimethyl sulfoxide were mixed, then 2 parts by weight of succinic anhydride was added, and after being sealed, it was stirred at 65°C for 8h, then 2 times volume of ethanol was added, the sample was soaked for 3h, then centrifuged at 4000 centrifugal force for 10min to obtain the precipitate;

[0073] S2.2: The precipitate was added to 20 parts by weight of deionized water, then a 5% concentration of sodium hydroxide solution was added to adjust the pH to 10, then 3 times volume of acetone was added, then centrifuged at 4000 centrifugal force for 10min, the obtained precipitate was washed with 70% ethanol for 2 times, and pure acetone for 4 times, the washed precipitate was dried at 50°C for 25h, then ground to obtain N-succinyl chitosan;

[0074] S3: Liposome-embedded astaxanthin

[0075] S3.1: 20 parts by weight of phospholipid, 5 parts by weight of cholesterol, 4 parts by weight of astaxanthin and 1 part by weight of garlicin were dissolved in 50 parts by weight of chloroform, then rotary evaporation was performed on a rotary evaporator to obtain a lipid film, 10 parts by weight of the lipid film was added to 30 parts by weight of deionized water and vortexed vigorously for 12min, then homogenized at 63°C for 20min, and then ultrasonic cell disruptor at 28°C with a probe at 70% intensity for 12min on ice to obtain a liposome suspension;

[0076] S3.2: 2 parts by weight of bovine serum albumin and 1 part by weight of polyethylene glycol methyl ether-succinimidyl ester with a molecular weight of 2000 were dissolved in 30 parts by weight of PBS solution with pH 7.4, and after stirring at room temperature for 3h, the reaction solution was transferred to a dialysis bag with a molecular weight cutoff of 14000Da, and dialysis purification was performed in pure water for 50h, and the dialysate was freeze-dried to obtain polyethylene glycol modified bovine serum albumin;

[0077] S3.3: The polyethylene glycol modified bovine serum albumin was added to deionized water, then stirred and mixed at 300r / min for 30min to obtain a 10%w / v polyethylene glycol modified bovine serum albumin solution, N-succinyl chitosan was dissolved in deionized water, stirred and mixed for 30min to obtain a 0.5%w / v N-succinyl chitosan solution, then 1% mulberry leaf extracellular vesicles were added and mixed to obtain a mixture;

[0078] S3.4: The liposome suspension was added to an equal volume of 10%w / v polyethylene glycol modified bovine serum albumin solution, then incubated at 65°C for 80min, then added to 2 times volume of the mixture, and incubated at 65°C for 30min to obtain N-succinyl chitosan / mulberry leaf extracellular vesicle modified liposome-embedded astaxanthin complex;

[0079] S4: Preparation of astaxanthin soft capsules

[0080] S4.1: 20 parts by weight of hydroxypropyl starch was added to 45 parts by weight of distilled water, after stirring and mixing, it was put into a drying oven at 30℃, and dried to a water content of 1%, then the dried hydroxypropyl starch was placed between the two electrodes of a plasma generator, and treated with argon at 320W for 5min to obtain modified hydroxypropyl starch;

[0081] S4.2: 28 parts by weight of modified hydroxypropyl starch, 20 parts by weight of glycerol and 50 parts by weight of deionized water were mixed, then stirred and mixed at 98℃ for 50min, then 2 parts by weight of agar was added, and stirred and mixed again at 98℃ for 50min, and finally incubated for 3h to obtain a glue solution;

[0082] S4.3: The N-succinyl chitosan / silkworm leaf extracellular vesicle modified liposome-embedded astaxanthin complex was used as the core content and the glue solution, and was sent into a pill making machine to perform pill pressing, shaping, drying, pill sorting and packaging to obtain astaxanthin soft capsules with a loading amount of 500mg / pill and a glue skin thickness of 0.85mm.

[0083] Example 3: A preparation process of astaxanthin soft capsules with hypoglycemic function, as shown in Figure 1 , comprising the following steps:

[0084] S1: Preparation of silkworm leaf extracellular vesicle

[0085] S1.1: 3 parts by weight of cellulase, 2 parts by weight of pectinase and 2 parts by weight of β-glucosidase were mixed, then added to a pH 5.5 acetic acid-sodium acetate buffer solution to adjust the pH of the enzyme solution to 5.2 to obtain an enzyme solution;

[0086] S1.2: Fresh silkworm leaves were washed, crushed to 20 mesh, and then added to the enzyme solution according to a solid-liquid ratio of 8g:180mL, then subjected to high-voltage pulsed electric field treatment under dark conditions at 20kV / cm for 3 pulses for 1h, then subjected to enzymolysis at 25℃ for 10h under a centrifugal force of 200, and then centrifuged at 100000 for 150min, the precipitate was discarded, and then centrifuged twice at 12000 for 10min each time, and the two supernatants were combined for standby;

[0087] S1.3: The combined supernatant was subjected to ultracentrifugation at 150000 under a centrifugal force at 4℃ for 100min using a sucrose pad, the sucrose layer was washed with a phosphate buffer solution, and then subjected to ultrafiltration centrifugation to obtain silkworm leaf extracellular vesicles;

[0088] S2: Preparation of N-succinyl chitosan

[0089] S2.1: 3 parts by weight of chitosan and 50 parts by weight of dimethyl sulfoxide were mixed, then 3 parts by weight of succinic anhydride was added, and after being sealed, it was stirred at 60°C for 6h, then 3 times the volume of ethanol was added, the sample was soaked for 2h, then centrifuged at 3500 centrifugal force for 5min to obtain a precipitate;

[0090] S2.2: The precipitate was added to 30 parts by weight of deionized water, then a sodium hydroxide solution with a concentration of 8% was added to adjust the pH to 11, then 4 times the volume of acetone was added, then centrifuged at 3500 centrifugal force for 5min, the obtained precipitate was washed once with 70% ethanol and 3 times with pure acetone, and the washed precipitate was dried at 40°C for 24h, then ground and crushed to obtain N-succinyl chitosan;

[0091] S3: Liposome-embedded astaxanthin

[0092] S3.1: 22 parts by weight of phospholipid, 8 parts by weight of cholesterol, 5 parts by weight of astaxanthin and 2 parts by weight of garlicin were dissolved in 80 parts by weight of chloroform, then rotary evaporation was performed on a rotary evaporator to obtain a lipid film, 20 parts by weight of the lipid film was added to 50 parts by weight of deionized water and vortexed vigorously for 10min, then homogenized at 60°C for 15min, and then ultrasonic cell disruptor was used at 70% intensity on ice for 10min at 25°C to obtain a liposome suspension;

[0093] S3.2: 3 parts by weight of bovine serum albumin and 2 parts by weight of polyethylene glycol methyl ether-succinimidyl ester with a molecular weight of 2000 were dissolved in 40 parts by weight of PBS solution with pH 7.4, and after stirring and reacting at room temperature for 2h, the reaction solution was transferred to a dialysis bag with a molecular weight cutoff of 8000Da, and dialysis purification was performed in pure water for 48h, and the dialysate was freeze-dried to obtain polyethylene glycol modified bovine serum albumin;

[0094] S3.3: The polyethylene glycol modified bovine serum albumin was added to deionized water, then stirred and mixed at 200r / min for 20min to obtain a 10%w / v polyethylene glycol modified bovine serum albumin solution, N-succinyl chitosan was dissolved in deionized water, stirred and mixed for 20min to obtain a 1%w / v N-succinyl chitosan solution, then 2% mulberry leaf extracellular vesicles were added and mixed to obtain a mixture;

[0095] S3.4: The liposome suspension was added to an equal volume of 10%w / v polyethylene glycol modified bovine serum albumin solution, then incubated at 60°C for 60min, then added to 3 times the volume of the mixture, and incubated at 60°C for 20min to obtain N-succinyl chitosan / mulberry leaf extracellular vesicle modified liposome-embedded astaxanthin complex;

[0096] S4: Preparation of astaxanthin soft capsules

[0097] S4.1: 30 parts by weight of hydroxypropyl starch was added to 50 parts by weight of distilled water, after stirring and mixing, it was put into a drying oven at 28℃, and dried to a water content of 1%, then the dried hydroxypropyl starch was put between the two electrodes of a plasma generator, and treated with argon at 300W for 4min to obtain modified hydroxypropyl starch;

[0098] S4.2: 30 parts by weight of modified hydroxypropyl starch, 22 parts by weight of glycerol and 52 parts by weight of deionized water were mixed, then stirred and mixed at 95℃ for 30min, then 3 parts by weight of agar was added, and stirred and mixed again at 95℃ for 30min, and finally incubated for 2h to obtain a glue solution;

[0099] S4.3: The N-succinyl chitosan / silkworm leaf extracellular vesicle modified liposome-embedded astaxanthin complex was used as the core content and the glue solution was fed into the pill making machine, and the astaxanthin soft capsule with a loading capacity of 500mg / pill and a glue thickness of 0.85mm was obtained after pressing, shaping, drying, picking and packaging.

[0100] Comparative Example 1: Compared with Example 1, the difference between Comparative Example 1 and Example 1 is that the garlicin in step S3.1 is removed, and the rest of the steps remain unchanged to prepare the astaxanthin soft capsule, which is recorded as Comparative Example 1.

[0101] Comparative Example 2: Compared with Example 1, the difference between Comparative Example 2 and Example 1 is that the astaxanthin in step S3.1 is removed, and the rest of the steps remain unchanged to prepare the soft capsule, which is recorded as Comparative Example 2.

[0102] Comparative Example 3: Compared with Example 1, the difference between Comparative Example 3 and Example 1 is that the mixed solution in steps S1 and S3.3 is removed, and the mixed solution in S3.4 is replaced with 0.5-1%w / v N-succinyl chitosan solution, and the rest of the steps remain unchanged to prepare the astaxanthin soft capsule, which is recorded as Comparative Example 3.

[0103] Comparative Example 4: Compared with Example 1, the difference between Comparative Example 4 and Example 1 is that the mixed solution in steps S2 and S3.3 is removed, and the mixed solution in S3.4 is replaced with 1-2%w / v silkworm leaf extracellular vesicle solution, and the rest of the steps remain unchanged to prepare the astaxanthin soft capsule, which is recorded as Comparative Example 4.

[0104] Comparative Example 5: Compared with Example 1, the difference between Comparative Example 5 and Example 1 is that step S4.1 is removed, and the modified hydroxypropyl starch in S4.2 is replaced with hydroxypropyl starch, and the rest of the steps remain unchanged to prepare the astaxanthin soft capsule, which is recorded as Comparative Example 5.

[0105] Hypoglycemic effect of the soft capsule:

[0106] ICR mice, half male and half female, weight 20-22g. Randomly grouped, 10 in each group, the mice were fasted for 18h before the test, the next morning intraperitoneal injection of STZ buffer 100mg / Kg (the blank group injected with the same amount of 0.1mol / L citric acid-sodium citrate buffer), the soft capsules prepared in Examples 1-3 and Comparative Examples 1-5 were administered by gavage, once a day, 1 capsule each time, for 14 consecutive days, the blank group and the model group were administered with purified water by gavage, at the same time, 0.4ml of high-fat high-sugar high-salt emulsion was given to each mouse to copy the type 2 diabetes mouse model, the blank group was fed with basic feed. 40min after the last administration on the 14th day, blood was taken, the fasting blood glucose of the mice was determined, the average value was taken, and the results are shown in Table 1.

[0107] Table 1. Results of determination of fasting blood glucose of mice

[0108] Blood glucose (mmol / L) blank group 5.84 model group 24.65 Example 1 12.87 Example 2 12.34 Example 3 12.42 Comparative Example 1 14.72 Comparative Example 2 19.87 Comparative Example 3 15.92 Comparative Example 4 14.78 Comparative Example 5 13.91

[0109] As can be seen from the data in Table 1, compared with the model group, the soft capsules prepared in the application can effectively reduce the fasting blood glucose value of the type 2 diabetes model, indicating that the soft capsules prepared in the application have a significant hypoglycemic effect; as can be seen from the data of Comparative Example 1 and Comparative Example 2, the active ingredients astaxanthin and allicin in the capsules of the application can produce a synergistic effect, further enhancing the hypoglycemic effect; as can be seen from the data of Comparative Example 2 and Comparative Example 3, the hypoglycemic effect is reduced, indicating that the hypoglycemic effect of N-succinyl chitosan / silkworm leaf extracellular vesicles after modification is better, because N-succinyl chitosan realizes covalent combination with the specific expression protein ligand of intestinal epithelial cells through amidation reaction, thereby accurately targeting intestinal epithelial cells, and the membrane structure of silkworm leaf extracellular vesicles is similar to that of cell membranes, which can better fuse with cells, thereby improving the bioavailability of astaxanthin and effectively improving the hypoglycemic effect; as can be seen from the data of Examples and Comparative Examples 3-4, the synergistic effect of the two can effectively improve the hypoglycemic effect of the soft capsules; as can be seen from the data of Comparative Example 5, using modified hydroxypropyl starch to prepare the capsule shell can also increase the hypoglycemic effect, because using modified hydroxypropyl starch to prepare the capsule shell can improve the dissolution rate of astaxanthin in the gastrointestinal tract, which is conducive to its absorption in the body, thereby improving the hypoglycemic effect.

[0110] The above examples only illustrate the principles and effects of the application, and are not intended to limit the application. Any person skilled in the art can modify or change the above examples without departing from the spirit and scope of the application. Therefore, all equivalent modifications or changes made by those skilled in the art without departing from the spirit and technical ideas disclosed by the application should be covered by the claims of the application.

Claims

1. A process for preparing astaxanthin soft capsules having a blood sugar-lowering function, characterized by, Comprising the following steps: S1: Preparation of mulberry leaf extracellular vesicles Fresh mulberry leaves are added to an enzyme solution, and then subjected to high-voltage pulsed electric field treatment and enzymolysis in the dark. After centrifugation, filtration is performed to obtain mulberry leaf extracellular vesicles. S2: Preparation of N-succinyl chitosan After mixing deacetylated chitosan and dimethyl sulfoxide, succinic anhydride is added for reaction to obtain precipitate. The precipitate is added to deionized water to adjust the pH, and then acetone is added. After centrifugal washing and drying, N-succinyl chitosan is obtained by grinding and crushing. S3: Liposome-embedded astaxanthin First, a liposome suspension is prepared using phospholipids, cholesterol, astaxanthin, and allicin. Then, the liposome is wrapped with polyethylene glycol modified bovine serum albumin. Finally, the N-succinyl chitosan / mulberry leaf extracellular vesicle modified liposome-embedded astaxanthin complex is obtained by wrapping with N-succinyl chitosan and mulberry leaf extracellular vesicles. S4: Preparation of astaxanthin soft capsules Low-pressure plasma is used to modify dried hydroxypropyl starch. Then, a gel solution is prepared using modified hydroxypropyl starch, glycerol, deionized water, and agar. The N-succinyl chitosan / mulberry leaf extracellular vesicle modified liposome-embedded astaxanthin complex is used as the capsule core content and the gel solution is fed into the pill-making machine to prepare astaxanthin soft capsules. The specific preparation process of modified hydroxypropyl starch is as follows: 20-30 parts by weight of hydroxypropyl starch is added to 45-50 parts by weight of distilled water, mixed and stirred, and then placed in a drying box at 28-30°C. After drying to a moisture content of 1%, the dried hydroxypropyl starch is placed between the two electrodes of a plasma generator, and argon gas is used to generate plasma at 300-320W for 4-5min to obtain modified hydroxypropyl starch.

2. The process for preparing astaxanthin soft capsules having a blood sugar lowering function according to claim 1, characterized in that, Step S1: Preparation of mulberry leaf extracellular vesicles, comprising the following steps: S1.1: Mix 2-3 parts by weight of cellulase, 1-2 parts by weight of pectinase, and 1-2 parts by weight of β-glucosidase, and then add a pH 4.5-5.5 acetic acid-sodium acetate buffer solution to adjust the pH of the enzyme solution to 4.8-5.2 to obtain the enzyme solution; S1.2: Wash fresh mulberry leaves, crush them to 20-30 mesh, and then add the enzyme solution. Then, under dark conditions, perform high-voltage pulsed electric field treatment at 20-30kV / cm and 3-4 pulses for 1-2h. Then, perform enzymolysis at 25-28°C and 200-300 centrifugal force for 10-12h. After standing, centrifuge the upper liquid at 100000-120000 centrifugal force for 150-180min, discard the precipitate, and then centrifuge at 12000-15000 centrifugal force for 2-3 times, each time for 10-12min. Combine the two supernatants and reserve for use; S1.3: The combined supernatant is subjected to ultracentrifugation at 4-5°C, 150000-180000 centrifugal force using a sucrose pad. After 100-120min, take the sucrose layer, wash it with a phosphate buffer solution, and then perform ultrafiltration centrifugation to obtain mulberry leaf extracellular vesicles.

3. The process for preparing astaxanthin soft capsules having a blood sugar lowering function according to claim 2, characterized in that, In step S1.2, the ratio of fresh mulberry leaves to enzyme solution is 5-8g:180mL.

4. The process for preparing astaxanthin soft capsules having a blood sugar lowering function according to claim 2, characterized in that, Step S2: Preparation of N-succinyl chitosan, comprising the following steps: S2.1: 2-3 parts by weight of deacetylated chitosan and 40-50 parts by weight of dimethyl sulfoxide are mixed, then 2-3 parts by weight of succinic anhydride is added, after sealing, stirring at 60-65 DEG C for 6-8 h, then 2-3 times volume of ethanol is added, the sample is soaked for 2-3 h, then centrifuged at 3500-4000 centrifugal force for 5-10 min to obtain a precipitate; S2.2: the precipitate is added to 20-30 parts by weight of deionized water, then a sodium hydroxide solution with a concentration of 5-8% is added to adjust the pH to 10-11, then 3-4 times volume of acetone is added, then centrifuged at 3500-4000 centrifugal force for 5-10 min, the obtained precipitate is washed with 70% ethanol for 1-2 times, and pure acetone for 3-4 times, the washed precipitate is dried at 40-50 DEG C for 24-25 h, then ground to obtain N-succinyl chitosan.

5. The process for preparing astaxanthin soft capsules having a blood sugar lowering function according to claim 4, wherein the astaxanthin is dissolved in a solvent, and the solution is filled into a soft capsule. Step S3 liposome embedding astaxanthin, specifically comprising the following steps: S3.1: 20-22 parts by weight of phospholipid, 5-8 parts by weight of cholesterol, 4-5 parts by weight of astaxanthin and 1-2 parts by weight of garlicin are dissolved in 50-80 parts by weight of chloroform, then rotary evaporation is performed on a rotary evaporator to obtain a lipid film, 10-20 parts by weight of the lipid film is added to 30-50 parts by weight of deionized water and vortexed vigorously for 10-12 min, then homogenized at 60-63 DEG C for 15-20 min, and then ultrasonic cell disrupter is used for ultrasonic treatment at 70% intensity on ice at 25-28 DEG C for 10-12 min to obtain a liposome suspension; S3.2: 2-3 parts by weight of bovine serum albumin and 1-2 parts by weight of polyethylene glycol methyl ether-succinimidyl ester are dissolved in 30-40 parts by weight of PBS solution with pH 7.4, after stirring and reacting at room temperature for 2-3 h, the reaction solution is transferred to a dialysis bag and dialyzed in pure water for 48-50 h, and then the dialysate is freeze-dried to obtain polyethylene glycol modified bovine serum albumin; S3.3: the polyethylene glycol modified bovine serum albumin is added to deionized water, then stirred and mixed at 200-300 r / min for 20-30 min to obtain a 10% w / v polyethylene glycol modified bovine serum albumin solution, N-succinyl chitosan is dissolved in deionized water, stirred and mixed for 20-30 min to obtain a 0.5-1% w / v N-succinyl chitosan solution, then 1-2% mulberry leaf extracellular vesicles are added and mixed to obtain a mixed solution; S3.4: the liposome suspension is added to an equal volume of 10% w / v polyethylene glycol modified bovine serum albumin solution, then incubated at 60-65 DEG C for 60-80 min, then added to 2-3 times volume of the mixed solution, and incubated at 60-65 DEG C for 20-30 min to obtain N-succinyl chitosan / mulberry leaf extracellular vesicle modified liposome embedding astaxanthin complex.

6. The process for preparing astaxanthin soft capsules having a blood sugar lowering function according to claim 5, wherein the astaxanthin is dissolved in a solvent, and the solution is filled into a soft capsule. The polyethylene glycol methyl ether-succinimidyl ester in step S3.2 has a molecular weight of 2000.

7. The process for preparing astaxanthin soft capsules having a blood sugar lowering function according to claim 5, wherein the astaxanthin is dissolved in a solvent, and the solution is filled into a soft capsule. The dialysis bag in step S3.2 is a dialysis bag with a molecular weight cut-off of 8000-14000 Da.

8. The process for preparing astaxanthin soft capsules having a blood sugar lowering function according to claim 7, characterized in that, Step S4 preparation of astaxanthin soft capsules, specifically comprising the following steps: S4.1: 20-30 parts by weight of hydroxypropyl starch is added to 45-50 parts by weight of distilled water, after stirring and mixing, it is placed in a drying box at 28-30℃, and dried to a water content of 1%, then the dried hydroxypropyl starch is placed between the two electrodes of a plasma generator for plasma treatment to obtain modified hydroxypropyl starch; S4.2: 28-30 parts by weight of modified hydroxypropyl starch, 20-22 parts by weight of glycerol and 50-52 parts by weight of deionized water are mixed, then stirred and mixed at 95-98℃ for 30-50 min, then 2-3 parts by weight of agar is added, and stirred and mixed again at 95-98℃ for 30-50 min, and finally incubated for 2-3 h to obtain a glue solution; S4.3: the N-succinyl chitosan / san leaf extracellular vesicle modified liposome-embedded astaxanthin compound is used as the capsule core content and the glue solution, and is sent into a pill making machine for pill pressing, shaping, drying, pill sorting and packaging to obtain astaxanthin soft capsules.

9. Astaxanthin soft capsule having a blood sugar lowering function, characterized by, It is prepared by the preparation process of the astaxanthin soft capsule with blood glucose lowering function according to any one of claims 1-8.

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