Curcumin-loaded zein alcohol-soluble glycosylation protein liposome as well as preparation method and application of curcumin-loaded zein alcohol-soluble glycosylation protein liposome

Abstract

The invention provides zein-soluble glycosylation protein lipidosome loaded with curcumin as well as a preparation method and application thereof, and belongs to the technical field of food and drug processing. According to the preparation method of the curcumin-loaded zein alcohol-soluble glycosylated protein lipidosome, active substances can be effectively embedded through a double embedding technology, the preparation method is simple, easy to operate, mild and safe in condition, simple and efficient in process and low in raw material cost, the prepared finished product is high in embedding rate, and the curcumin-loaded zein alcohol-soluble glycosylated protein lipidosome is suitable for industrial production. The curcumin transport carrier is suitable for large-scale production, and solves the problems of low encapsulation efficiency, low loading capacity, poor stability, poor bioaccessibility, low curcumin utilization rate and the like of the curcumin transport carrier in the prior art. And the zein alcohol-soluble glycosylation protein liposome provided by the invention can be applied to carrying of active substances or drug transportation.

Description

Technical Field

[0001] The invention belongs to the technical field of food and medicine processing, and particularly relates to a method for preparing a curcumin transport carrier. Background Art

[0002] Curcumin is a natural phenolic antioxidant extracted from plant rhizomes. It belongs to the diketone class of compounds and has multiple biological activities. It has potential benefits to human health. However, because it is easily soluble in ethanol solutions, has low solubility in water and poor stability, its absorption rate and utilization in the human body are low.

[0003] Zein has good biocompatibility, degradability, self-assembly and stability, and can be used as a transport carrier for active substances. However, since zein is composed of hydrophobic residues and lacks charged amino acids, it is insoluble in water. This characteristic makes zein face challenges when used as a carrier material for transporting active substances, greatly limiting its application in the field of food and medicine. In order to improve the application prospects of zein, the existing technology modifies zein by methods such as protease hydrolysis, glycosylation reaction, and amino zein, but there are deficiencies in stability and uniformity.

[0004] During the processing of food and medicine, many active substances are very susceptible to degradation and oxidation, and nano-encapsulation technology is used in the prior art to protect them. Liposomes are carrier materials with a lipid bilayer structure, which can encapsulate both hydrophilic and hydrophobic active substances, and can transport active substances to targeted locations. In existing studies, curcumin is generally encapsulated by liposome phospholipid bilayers or by protein composite nanoparticles. This single encapsulation technology has problems such as low encapsulation rate and loading capacity, poor stability and poor bioaccessibility. Therefore, a new type of curcumin transport carrier is urgently needed. Summary of the invention

[0005] The invention solves the problems in the prior art of low encapsulation rate and loading capacity of the curcumin transport carrier, poor stability, poor bioaccessibility and low utilization rate of curcumin.

[0006] A method for preparing zein-soluble glycosylated proteoliposomes loaded with curcumin, the method comprising the following steps:

[0007] S1: dissolving zein in water, adjusting the pH, adding Alcalase for enzymatic hydrolysis for 2 h, then stopping the hydrolysis and centrifuging, and freeze-drying the supernatant after centrifugation to obtain zein hydrolysate powder;

[0008] S2: dissolving the zein hydrolysate powder in water to obtain zein hydrolysate, adding chitosan oligosaccharide and TGase enzyme, reacting for 6 hours at 37°C and pH 7.5, then inactivating the enzyme and dialyzing in deionized water for 2 days, freeze-drying, and obtaining glycosylated zein powder;

[0009] S3: dissolving the glycosylated zein powder in water to obtain a zein glycosylated protein solution, dissolving curcumin in 70% ethanol, and injecting an equal volume of the curcumin solution into the zein glycosylated protein solution to form a complex;

[0010] S4: Soybean lecithin and cholesterol were dissolved in anhydrous ethanol, rotary evaporated at 40°C for 15 min, and hydrated for 1 h by adding 45 mL of PBS solution, and then hydrated for 30 min by adding PBS solution containing the complex, and liposomes were prepared by treating with an ultrasonic probe.

[0011] Preferably, the added amount of Alcalase is 1.2 g.

[0012] Preferably, the mass ratio of the zein enzyme hydrolysate to chitosan oligosaccharide is 3:1.

[0013] Preferably, the TGase enzyme addition amount is 60U / g.

[0014] Preferably, the mass volume ratio of the rice protease hydrolysate powder dissolved in water is 3%.

[0015] Preferably, the mass volume ratio of the glycosylated zein powder dissolved in water is 0.25%.

[0016] Preferably, the mass volume ratio of the curcumin dissolved in 70% ethanol is 0.3%.

[0017] Preferably, the soybean lecithin is 120 mg, the cholesterol is 30 mg, and the anhydrous ethanol is 60 mL.

[0018] A zein-soluble glycosylated proteoliposome loaded with curcumin is prepared by the method.

[0019] Application of zein-soluble glycosylated proteoliposome loaded with curcumin in the preparation of a drug for delivering curcumin.

[0020] Beneficial Effects

[0021] 1. The present invention catalyzes the reaction of zein and chitosan oligosaccharide by TGase enzyme, which is an enzymatic glycosylation reaction. Compared with the reaction temperature required for the Maillard reaction, it is milder, has a short reaction time, and does not produce harmful mutagenic substances. TGase enzyme can catalyze the reaction of glutamine residues in zein molecules with the epsilon-amino group in lysine residues, so that zein molecules undergo intramolecular or intermolecular crosslinking, i.e., zein crosslinked protein. Wherein, lysine residues can be replaced by primary amine substances in chitosan oligosaccharide, i.e., TGase enzyme can catalyze the reaction of zein and chitosan oligosaccharide. With zein as acyl donor and chitosan oligosaccharide as acyl acceptor, the catalytic action of TGase enzyme is utilized to promote enzymatic glycosylation reaction of protein and chitosan oligosaccharide, i.e., zein glycosylated protein. Chitosan oligosaccharide has good solubility and is easier to diffuse than zein. Chitosan oligosaccharide is more active than the N-terminal amine of peptide or the ε-amino group of lysine. Zein glycosylated protein combines the hydrophilicity of chitosan oligosaccharide and the macromolecular characteristics of zein. Therefore, the use of zein glycosylated protein as an embedding material has more advantages than zein cross-linked protein.

[0022] 2. The present invention provides a liposome with double-embedded curcumin, wherein enzymatically glycosylated zein and liposome vesicles are used to double-embed curcumin, wherein the first layer of embedding wall material of the liposome with double-embedded curcumin is an enzymatically glycosylated chitosan oligosaccharide-zein complex, and the second layer of embedding wall material is a liposome vesicle. The zein liposome loaded with curcumin prepared by the present invention has high stability, good bioaccessibility and good in vitro digestion experiment effect, and can be used for the processing of drugs and foods.

[0023] 3. The present invention utilizes the good biocompatibility of liposomes to perform secondary encapsulation of curcumin. Compared with single chitosan oligosaccharide-zein encapsulation, the intestinal fluid release rate and bioaccessibility of curcumin can be significantly improved. The intestinal fluid release rate is increased from 31.36% to 73.6% at 120 min, and the bioaccessibility is increased from 28.6% to 70.1%. The improvement in bioaccessibility indicates that the double encapsulation technology has good encapsulation ability.

[0024] 4. The liposomes of zein-soluble glycosylated protein loaded with curcumin prepared by the present invention can better retain the biological activity of curcumin. The particle size of the liposomes is 269nm, the PDI is 0.3, the potential is -11.9mv, the distribution is uniform, and the stability is good. In the in vitro digestion experiment simulation, the gastric juice release rate is only 8.1% in 120min, the intestinal juice release rate is as high as 73.6%, and the bioaccessibility is 70.1%, which improves the biological activity of curcumin and the survival rate in gastric juice and the release rate in intestinal juice.

[0025] 5. The double encapsulation technology provided by the present invention can effectively encapsulate active substances. The preparation method is simple, easy to operate, the conditions are mild and safe, the process is simple and efficient, the raw material cost is low, the finished product has a high encapsulation rate, and is suitable for large-scale production. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 is the FESEM image of zein liposomes loaded with curcumin;

[0027] Figure 2 This is the FESEM image of zein cross-linked proteoliposomes loaded with curcumin;

[0028] Figure 3 This is the FESEM image of zein-soluble glycosylated proteoliposomes loaded with curcumin;

[0029] Figure 4 It is a comparison result of particle size and PDI, wherein Us-Cur-Zein-Lip is zein liposome loaded with curcumin, Cl-Cur-Zein-Lip is zein glycosylated protein liposome loaded with curcumin, and Gly-Cur-Zein-Lip is zein glycosylated protein liposome loaded with curcumin;

[0030] Figure 5 The potential comparison result of the liposomes in Examples 1-3 is shown, wherein Us-Cur-Zein-Lip is a zein hydrolyzed protein liposome loaded with curcumin, Cl-Cur-Zein-Lip is a zein collagen liposome loaded with curcumin, and Gly-Cur-Zein-Lip is a zein glycosylated protein liposome loaded with curcumin;

[0031] Figure 6 : is a graph showing the results of gastric fluid release rate, wherein Us-Cur-Zein is zein alcohol-soluble hydrolyzed protein nanoparticles loaded with curcumin, Us-Cur-Zein-Lip is zein alcohol-soluble hydrolyzed protein liposomes loaded with curcumin, Cl-Cur-Zein is zein alcohol-soluble cross-linked protein nanoparticles loaded with curcumin, Cl-Cur-Zein-Lip is zein alcohol-soluble collagen liposomes loaded with curcumin, Gly-Cur-Zein is zein alcohol-soluble glycosylated protein nanoparticles loaded with curcumin, and Gly-Cur-Zein-Lip is zein alcohol-soluble glycosylated protein liposomes loaded with curcumin;

[0032] Figure 7: is the intestinal fluid release rate result graph, wherein Us-Cur-Zein is zein alcohol-soluble hydrolyzed protein nanoparticles loaded with curcumin, Us-Cur-Zein-Lip is zein alcohol-soluble hydrolyzed protein liposomes loaded with curcumin, Cl-Cur-Zein is zein alcohol-soluble cross-linked protein nanoparticles loaded with curcumin, Cl-Cur-Zein-Lip is zein alcohol-soluble collagen liposomes loaded with curcumin, Gly-Cur-Zein is zein alcohol-soluble glycosylated protein nanoparticles loaded with curcumin, and Gly-Cur-Zein-Lip is zein alcohol-soluble glycosylated protein liposomes loaded with curcumin;

[0033] Figure 8 is the encapsulation efficiency and loading capacity of the nanoparticles in Comparative Examples 1-3, wherein Us-Cur-Zein is a zein alcohol-soluble hydrolyzed protein nanoparticle loaded with curcumin, Cl-Cur-Zein is a zein alcohol-soluble cross-linked protein nanoparticle loaded with curcumin, and Gly-Cur-Zein is a zein alcohol-soluble glycosylated protein nanoparticle loaded with curcumin;

[0034] Fig. 9 It is a graph of bioaccessibility test results, wherein Us-Cur-Zein is zein alcohol-soluble hydrolyzed protein nanoparticles loaded with curcumin, Us-Cur-Zein-Lip is zein alcohol-soluble hydrolyzed protein liposomes loaded with curcumin, Cl-Cur-Zein is zein alcohol-soluble cross-linked protein nanoparticles loaded with curcumin, Cl-Cur-Zein-Lip is zein alcohol-soluble collagen liposomes loaded with curcumin, Gly-Cur-Zein is zein alcohol-soluble glycosylated protein nanoparticles loaded with curcumin, and Gly-Cur-Zein-Lip is zein alcohol-soluble glycosylated protein liposomes loaded with curcumin. DETAILED DESCRIPTION

[0035] Example 1. Preparation of zein hydrolyzate liposomes loaded with curcumin.

[0036] S1: Preparation of zein hydrolysate. Dissolve 40 g zein in 800 mL water, adjust the pH to 9, add 1.2 g Alcalase and perform enzymatic hydrolysis in a magnetic stirring water bath for 2 h. Then, heat in a water bath at 85°C for 5 min to stop the hydrolysis, obtain the product, centrifuge the product at 5000 rpm for 20 min, and freeze-dry the supernatant after centrifugation to obtain zein hydrolysate powder.

[0037] S2: Preparation of a zein-loaded hydrolyzed protein complex. A certain mass of zein hydrolyzate powder was weighed and dissolved in water at a mass concentration of 0.25% (m / v), and then subjected to magnetic stirring at 600 r / min for 30 min to obtain a zein solution. Curcumin was dissolved in 70% (v / v) ethanol at a mass concentration of 0.3% (m / v) to obtain a curcumin solution. An equal volume of curcumin solution was slowly injected into the zein solution using a syringe with a fine needle, and the mixture was continuously stirred for 2 h using a magnetic stirring at 600 r / min to form a complex.

[0038] S3: Preparation of liposomes. 120 mg of soybean lecithin and 30 mg of cholesterol were dissolved in 60 mL of anhydrous ethanol, and the anhydrous ethanol was removed by rotary evaporation at 40°C for 15 min to obtain a phospholipid membrane, and 45 mL of PBS solution was added to hydrate for 1 h to remove the phospholipid membrane. Then, 15 mL of PBS solution (containing 15 mg of zein complex loaded with curcumin) was added thereto and hydrated for 30 min. After ultrasonic probe was used for 5 min, uniformly dispersed liposomes were obtained.

[0039] Example 2. Preparation of zein liposomes loaded with curcumin.

[0040] S1: Preparation of zein hydrolysate. Dissolve 40 g zein in 800 mL water, adjust the pH to 9, add 1.2 g Alcalase and perform enzymatic hydrolysis in a magnetic stirring water bath for 2 h. After the reaction is completed, heat in a water bath at 85°C for 5 min to stop the hydrolysis, obtain the product, centrifuge the product at 5000 rpm for 20 min, and freeze-dry the supernatant after centrifugation to obtain zein hydrolysate powder.

[0041] S2: TGase enzyme cross-linked zein. The zein hydrolysate powder was dissolved in water with a mass concentration of 3% (m / v), and 60U / g of TGase enzyme was added. The reaction was carried out at 37°C and pH 7.5 for 6 hours. After the reaction, the enzyme was inactivated in an 85°C water bath for 5 minutes, cooled to room temperature, dialyzed in deionized water at 4°C for 2 days, and finally freeze-dried to obtain TGase enzyme cross-linked zein powder.

[0042] S3: Preparation of a curcumin-loaded corn alcohol-soluble cross-linked protein complex. The TGase enzyme cross-linked corn alcohol-soluble protein powder was dissolved in water at a mass concentration of 0.25% (m / v), and then magnetic stirring (600r / min) was performed for 30 minutes to obtain a TGase enzyme cross-linked corn alcohol-soluble protein solution. Curcumin was dissolved in 70% (v / v) ethanol at a mass concentration of 0.3% (m / v) to obtain a curcumin solution. Then, an equal volume of curcumin solution was slowly injected into the TGase enzyme cross-linked corn alcohol-soluble protein solution using a syringe with a fine needle, and stirred continuously for 2 hours (600r / min) with a magnetic stirrer to form a complex.

[0043] S4: Preparation of liposomes. 120 mg of soybean lecithin and 30 mg of cholesterol were dissolved in 60 mL of anhydrous ethanol, and the anhydrous ethanol was removed by rotary evaporation at 40°C for 15 min to obtain a phospholipid membrane, and 45 mL of PBS solution was added to hydrate for 1 h to make the phospholipid membrane fall off. Then 15 mL of PBS solution (containing 15 mg of the complex) was added to hydrate for 30 min, and after ultrasonic probe for 5 min, uniformly dispersed liposomes were obtained.

[0044] Example 3. Preparation of zein-loaded liposomes containing curcumin.

[0045] S1: Preparation of zein hydrolysate. Dissolve 40 g zein in 800 mL water, adjust the pH to 9, add 1.2 g Alcalase and perform enzymatic hydrolysis in a magnetic stirring water bath for 2 h. After the reaction is completed, heat in a water bath at 85°C for 5 min to stop the hydrolysis, obtain the product, centrifuge the product at 5000 rpm for 20 min, and freeze-dry the supernatant after centrifugation to obtain zein hydrolysate powder.

[0046] S2: Glycosylation of zein. The zein hydrolysate powder was dissolved in water at a mass concentration of 3% (m / v) to obtain zein hydrolysate, chitosan oligosaccharide (the mass ratio of zein hydrolysate to chitosan oligosaccharide was 3:1) and 60U / g TGase enzyme were added, and the reaction was carried out at 37°C and pH 7.5 for 6h for enzymatic glycosylation. After the reaction, the enzyme was inactivated in an 85°C water bath for 5min, cooled to room temperature, dialyzed in deionized water at 4°C for 2 days to remove unreacted chitosan oligosaccharide, and finally freeze-dried to obtain glycosylated zein powder.

[0047] S3: Preparation of a zein-glycosylated protein complex loaded with curcumin. Glycosylated zein powder was dissolved in water at a mass concentration of 0.25% (m / v), and then subjected to magnetic stirring at 600 r / min to obtain a zein-glycosylated protein solution. Curcumin was dissolved in 70% (v / v) ethanol at a mass concentration of 0.3% (m / v) to obtain a curcumin solution. Then, an equal volume of curcumin solution was slowly injected into the zein-glycosylated protein solution using a syringe with a fine needle, and the mixture was continuously stirred for 2 h using a magnetic stirring at 600 r / min to form a complex.

[0048] S4: Preparation of liposomes. Dissolve 120 mg of soybean lecithin and 30 mg of cholesterol in 60 mL of anhydrous ethanol, remove the anhydrous ethanol by rotary evaporation at 40°C for 15 min to obtain a phospholipid membrane, and add 45 mL of PBS solution to hydrate for 1 h to remove the phospholipid membrane. Then add 15 mL of PBS solution (containing 15 mg of the complex) to hydrate for 30 min, and after ultrasonic probe for 5 min, uniformly dispersed liposomes were obtained.

[0049] Comparative Example 1. Preparation of curcumin-loaded corn alcohol-soluble hydrolyzed protein nanoparticles.

[0050] The preparation method of this comparative example is consistent with S1 and S2 in Example 1, except that liposome embedding is not used, that is, S3 is missing.

[0051] Comparative Example 2. Preparation of curcumin-loaded zein nanoparticles.

[0052] The preparation method of this comparative example is consistent with S1, S2 and S3 in Example 2, except that liposome embedding is not used, that is, S4 is missing.

[0053] Comparative Example 3. Preparation of curcumin-loaded zein protein nanoparticles.

[0054] The preparation method of this comparative example is consistent with S1, S2 and S3 in Example 3, except that liposome embedding is not used, that is, S4 is missing.

[0055] Example 4. Determination of the particle morphology of zein liposomes loaded with curcumin.

[0056] The frozen samples of liposomes in Examples 1-3 were transferred to the product chamber under vacuum, chopped with a cooling knife, sublimated at -95°C for 5 min, and sputtered with a platinum coating using an ion sputtering device. The samples were then transferred to a scanning electron microscope and observed at -130°C. An accelerating voltage of 5 kV was used and images were taken at 60,000 times to observe changes in the microstructure of the samples. The experimental results are shown in Figure 1-3 .

[0057] like Figure 1-3 As shown, compared with the liposomes of corn alcohol-soluble glycosylated protein loaded with curcumin in Example 3, the liposomes of corn alcohol-soluble hydrolyzed protein loaded with curcumin in Example 1 present an irregular shape, while the liposomes of corn alcohol-soluble cross-linked protein loaded with curcumin in Example 2 are more uniform in shape, because corn alcohol-soluble protein is cross-linked under the action of TGase enzyme to form protein-protein polymers, and hydrophobic amino acids are wrapped with curcumin inside, and liposomes are wrapped with corn alcohol-soluble cross-linked protein outside, presenting an ellipsoidal shape. In Example 3, the liposomes of corn alcohol-soluble protein embedded curcumin after glycosylation and cross-linking are because corn alcohol-soluble protein and chitosan oligosaccharide undergo glycosylation reaction under the action of TGase enzyme, and the acyl group of corn alcohol-soluble protein and chitosan oligosaccharide combine to form a protein-sugar complex. The hydrophobic amino acids of corn alcohol-soluble protein self-assemble in the interior of the liposome to wrap curcumin, and the hydrophilic chitosan oligosaccharide is distributed in the outer layer of the protein-sugar complex, and the complex is embedded in the two layers of liposomes and is spherical. Comparing the three, the zein-soluble glycosylated proteoliposomes loaded with curcumin in Example 3 have a uniform structure, a round spherical shape, and a better embedding effect.

[0058] Example 5. Detection of liposome particle size, Zeta potential and PDI.

[0059] The three liposome samples in Examples 1-3 were diluted to 1 mg / mL with phosphate buffer (PBS buffer, pH 7, concentration 0.01 mol / L), and then the particle size, Zeta potential, and PDI were measured using a particle size analyzer. The experimental results are shown in Figure 4-5 .

[0060] The particle size of the liposome of the zein glycosylated protein loaded with curcumin in Example 3 is 269nm, the PDI is 0.3, and the Zeta potential is -11.9mv, which is smaller than the other two liposomes in particle size and PDI, and has better dispersibility. At this time, the absolute value of the Zeta potential is large, the electrostatic repulsion is large, and the aggregation of the liposomes can be effectively prevented, and the liposomes are more stable. The particle size distribution of the glycosylated zein liposome is uniform, and the system is stable.

[0061] Example 6. Detection of release rate in gastrointestinal fluid.

[0062] Zein nanoparticles and liposomes loaded with curcumin were digested in vitro using a static in vitro digestion model. The liposomes and nanoparticles in Examples 1-3 and Comparative Examples 1-3 were mixed with simulated gastric fluid (SGF) in equal volumes, stirred at 100 r / min for 2 hours at 37°C to simulate the gastric digestion process, and samples were taken every 30 minutes. After the simulated gastric digestion, an equal volume of simulated intestinal fluid (SIF) was mixed into the above-mentioned gastric digestion solution, and the pH value of the solution was adjusted to 7 with NaOH. The solution was incubated at 37°C for 120 minutes with shaking to simulate the intestinal digestion process, and samples were taken at 15, 30, 60, 90 and 120 minutes of digestion, respectively. The experimental results are shown in Table 1. Figure 6-7 .

[0063] In the first two hours of digestion, the curcumin gastric juice release rates in Examples 1, 2, 3 and Comparative Examples 1, 2, 3 were 8.8%, 8.3%, 8.1% and 17.27%, 16.31%, 16.04% respectively. The gastric juice release rates corresponding to the nanoparticles in the comparative examples were all unsatisfactory, that is, the nanoparticles in Comparative Examples 1-3 were destroyed by gastric juice. In comparison, the double-embedding system in Examples 1-3 had a better embedding effect in gastric juice than the single-embedding system in Comparative Examples 1-3, and better retained the biological activity of curcumin. The gastric juice release rates of the double-embedding system liposomes in Examples 2 and 3 were not much different, indicating that these two double-embedded liposomes can protect curcumin in gastric juice, but the gastric juice release rate of the zein glycosylated protein liposome loaded with curcumin in Example 3 was the lowest, indicating that more curcumin can be encapsulated by double embedding of glycosylated zein and liposomes. After 2 hours in the intestinal fluid, the intestinal fluid release rates of the curcumin in Examples 1, 2, 3 and Comparative Examples 1, 2, 3 were 68.9%, 71.1%, 73.6% and 26%, 27.8%, 31.36%, respectively, indicating that compared with the liposomes of Comparative Examples 1-3, the curcumin embedded in the liposomes of Examples 1-3 can be better released from the intestinal fluid, thereby being absorbed and utilized by the human body, and more than half of the curcumin embedded in the liposomes of Examples 1-3 can enter the body, among which the liposomes of the zein-soluble glycosylated protein loaded with curcumin in Example 3 have the best effect of entering the body, that is, the liposomes of the zein-soluble glycosylated protein loaded with curcumin prepared by the present invention can more effectively retain the biological activity of curcumin in a simulated digestive environment in vitro.

[0064] Example 7. Determination of the encapsulation efficiency, loading capacity and bioaccessibility of the corresponding dual-entrapped liposomes of curcumin-loaded zein nanoparticles.

[0065] 1. Determine the encapsulation efficiency and loading capacity of zein nanoparticles loaded with curcumin.

[0066] The nanoparticle complexes in Comparative Examples 1-3 were mixed with 80% ethanol solution at a volume ratio of 1:9 (v / v). The resulting mixture was rotated for 90 seconds, and the absorbance of the solution at a wavelength of 419 nm was recorded using a UV-visible spectrophotometer. The encapsulation efficiency and loading capacity of the nanoparticles were calculated using the following formula:

[0067]

[0068]

[0069] 2. Determine the bioaccessibility of liposomes.

[0070] The liposomes in Examples 1-3 and Comparative Examples 1-3 were mixed with simulated gastric fluid (SGF) in equal volumes, stirred at 100 r / min for 2 h at 37° C. to simulate the gastric digestion process, and then the content of curcumin in the final mixed micelles was measured by a visible spectrophotometer. The bioavailability of curcumin was calculated using the following formula:

[0071]

[0072] The experimental results are shown in Figure 8 The embedding efficiency of the three nanoparticles in Comparative Examples 1-3 was 69.33%, 75.49%, and 83.34%, and the loading capacity was 5.38%, 5.88%, and 6.51%. Under the same conditions, the embedding efficiency and loading capacity of the zein nanoparticles modified by enzymatic glycosylation were significantly higher than those of the zein nanoparticles modified by hydrolysis and cross-linking. Fig. 9 As shown, the bioaccessibility of the liposomes in Examples 1-3 and the nanoparticles in Comparative Examples 1-3 are 54.2%, 67.8%, 70.1%, 22.2%, 24.7% and 28.6%, respectively, indicating that the curcumin in the three liposomes and the three nanoparticles can be absorbed and utilized by the human body through the gastrointestinal tract, wherein the bioaccessibility of curcumin in the zein glycosylated protein liposomes in Example 3 is increased from 28.6% to 70.1%, indicating that the secondary encapsulation of curcumin by utilizing the good biocompatibility of liposome vesicles can significantly improve the bioaccessibility of curcumin compared with the single chitosan oligosaccharide-zein encapsulation, improve the survival rate of curcumin in the gastrointestinal fluid and the absorption and utilization of curcumin by the human body, and the encapsulation capacity is better than that of the nanoparticles, which further proves that the zein glycosylated protein liposomes prepared by the enzymatic glycosylation modification method can effectively improve the encapsulation efficiency and loading capacity.

[0073] In summary, zein-glycosylated proteoliposomes loaded with curcumin are the most ideal dual encapsulation materials. They not only have good particle morphology and stability, but their delivery system can also protect curcumin in gastric juice better than zein nanoparticle encapsulation system, and can be released to the maximum extent in intestinal juice and enter the body, which is beneficial to improve the bioavailability of curcumin.

Claims

1. A method for preparing zein-soluble glycosylated proteoliposomes loaded with curcumin, characterized in that: The method steps are as follows: S1: dissolving zein in water, adjusting the pH, adding Alcalase for enzymatic hydrolysis for 2 h, then stopping the hydrolysis and centrifuging, and freeze-drying the supernatant after centrifugation to obtain zein hydrolysate powder; S2: dissolving the zein hydrolysate powder in water to obtain zein hydrolysate, adding chitosan oligosaccharide and TGase enzyme, reacting for 6 hours at 37°C and pH 7.5, then inactivating the enzyme and dialyzing in deionized water for 2 days, freeze-drying, and obtaining glycosylated zein powder; S3: dissolving the glycosylated zein powder in water to obtain a zein glycosylated protein solution, dissolving curcumin in 70% ethanol, and injecting an equal volume of the curcumin solution into the zein glycosylated protein solution to form a complex; S4: Soybean lecithin and cholesterol were dissolved in anhydrous ethanol, rotary evaporated at 40°C for 15 min, and hydrated for 1 h by adding 45 mL of PBS solution, and then hydrated for 30 min by adding PBS solution containing the complex, and liposomes were prepared by treating with an ultrasonic probe.

2. The method according to claim 1, characterized in that The amount of Alcalase added is 1.2 g.

3. The method according to claim 1, characterized in that The mass ratio of the corn alcohol-soluble protein hydrolysate to chitosan oligosaccharide is 3:

1.

4. The method according to claim 1, characterized in that The TGase enzyme addition amount is 60U / g.

5. The method according to claim 1, characterized in that The mass volume ratio of the corn soluble protein hydrolysate powder dissolved in water is 3%.

6. The method according to claim 1, characterized in that The mass volume ratio of the glycosylated zein powder dissolved in water is 0.25%.

7. The method according to claim 1, characterized in that The curcumin is dissolved in 70% ethanol at a mass volume ratio of 0.3%.

8. The method according to claim 1, characterized in that The soybean lecithin is 120 mg, the cholesterol is 30 mg, and the anhydrous ethanol is 60 mL.

9. A zein-soluble glycosylated proteoliposome loaded with curcumin, characterized in that: The method is prepared by any one of claims 1 to 8.

10. Use of the zein-glycosylated proteoliposome loaded with curcumin according to claim 9 in the preparation of a drug for delivering curcumin.

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