Preparation Of Hypoglycemic Nanocapsules Loaded With Gallic Acid-Antarctic Krill Peptide Copolymer Based On Microjet Technology

In recent years,natural active ingredients extracted from nature as anti-diabetes functional food have attracted more and more attention.The preparation of two or more active ingredients into hypoglycemic copolymers is beneficial to achieve the complementary hypoglycemic mechanism and enhance the hypoglycemic effect.In vivo delivery of nanoparticles has always been a research hotspot at home and abroad.By selecting appropriate carriers and combining with effective embedding technology,the targeted and controlled release delivery of target components in the form of nanocapsules can be achieved,which provides great possibilities for the efficient utilization of drugs.In this paper,the DPP-IV inhibitory peptide isolated from the enzymatic hydrolysis products of Antarctic Krill in our previous research group was used as the original material.Gallic Acid was introduced to form the Gallic Acid-Antarctic Krill peptide(GA-AKP)copolymer with hypoglycemic effect.Based on the preparation of microcapsules by traditional emulsion solvent evaporation method,the structure and characteristics of the emulsion were further optimized by means of high-pressure microfluidization,then the hypoglycemic nanocapsules loaded with copolymer were obtained.The microstructure of the nanocapsules,the distribution and release characteristics of the copolymer in the capsule were investigated.(1)The optimum reaction solvent,temperature and p H of the GA-AKP polymer were investigated by ultraviolet spectrophotometry.The polymer was analyzed by means of particle size analyzer,scanning electron microscope and Fourier infrared transform(FITR).At the same time,the thermal stability and inhibitory activity of the copolymer on three glucose-related enzymes were investigated under different gallic acid dosage.The results showed that the optimum conditions for GA-AKP were as follows: pH7.0 phosphate buffer solution at 20℃.The average particle size of the copolymer particles was 191±8 nm,and the polydispersity index was 0.208.GA was used as the crosslinking agent to make the peptides polymerize and fold,forming a three-dimensional complex structure.The FTIR showed that the reaction was driven by covalent and also non-covalent forces.During the reaction,C-N chemical bond was formed,and the random curl of AKP increased from 38.81% to 45.05%.The introduction of 0.5 mg/m L GA increase the thermal denaturation temperature of Antarctic krill peptide to 88.2℃,at the same time the Antarctic krill peptide ofα-amylase,α-glycosidase enzymes,DPP-Ⅳinhibition rate raised 61.0,39.5 and 4.6%respectively.(2)The technological parameters of GA-AKP copolymer nano-emulsion prepared by high pressure microfluidization were studied.The microstructures,particle size,PDI,potential,rheological viscosity and centrifugal stability and raw material embedding rate of nano-emulsion were used as evaluation indexes.The optimal process parameters were determined by single factor experiment: oil phase PLGA concentration of 20 mg/m L,microfluidization pressure of 150 Pa,and cycle times of 3times.At this condition,the particle size of the nano-emulsion was 285.4 nm,the PDI was 0.13,and the static stability,static flow stability and centrifugal stability were showed well.The embedding rates of GA and AKP were 71.5 and 85.2%,respectively,as well as the PLGA loading capacity were 14.3 and 170.4 mg/g,respectively.(3)Nano-capsules containing GA-AKP copolymer were prepared after spin steaming,centrifugation and freeze-drying.The characterization,release and stability of nano-capsules were investigated.Scanning electron microscopy and laser confocal microscopy were used to observe the microscopic morphology and the distribution of raw materials inside nano-capsules.FITR was used to analyze the reaction groups.The release curve of the nano-capsules was measured in the simulated gastrointestinal digestive fluid and the release equation was fitted.In general,the high pressure microjet brings the microstructure of well-structured and uniform particle size to the nano-capsules,and the particle size of the capsules is concentrated in the range of300-400 nm.The capsule w/o structure was complete,and the water-soluble core material was completely wrapped in the oil phase and distributed evenly.The capsule structure can effectively reduce the damage of raw materials caused by low p H in gastric fluid,so that the release of raw materials in simulated gastric fluid is very low,and the release of raw materials in intestinal fluid is slow.The best fit with Higuchi drug release equation is consistent with the characteristics of sustained release preparation.In addition,the double-layer embedding reduces the material exchange between raw materials and the external environment and improves the hygroscopic and thermal stability of raw materials.The activity experiment showed that the activity retention rate of raw materials after embedding was better than that before embedding,and-20℃was the most favorable for the activity retention.