Preparation and characterization of restructured vesicles of grape-derived multicomponents for active peptides loading / 中草药

ABSTRACT

Objective To reconstruct the functional vesicles by using the components isolated from grapes, and investigate its properties of the loading of peptides. Methods Grape polyphenols (GP) were extracted by ethanol, and the extraction technology of GP was optimized by single factor and orthogonal experiments. GP was purified by macroporous resin adsorption and the purification technology. The preparation of GP and melittin (Mel) complexes (GPMC) were determined by orthogonal tests, and Sucrose density gradient centrifugation was used to extract grape-derived vesicles (GDVs) for loading complex GPMC to get GPMC-GDVs. The stability of GPMC-GDVs in PBS, DMEM, and 10% FBS were investigated. The cytotoxicity of GPMC and GPMC-GDVs on SMMC-7721 or HepG 2 cells was investigated by MTT method. Results The extraction process of GP was as follows the extraction solvent was 60% ethanol solution, the heating temperature was 50 ℃, the solid-liquid ratio was 110, the extraction time was 50 min and extract 2 times. The purification conditions of GP were as follows 4 mg/mL GP crude sample volume was 7 times of bed volume (BV), the washing dosage was 5 BV, and the elution volume of 60% ethanol was 5 BV. The preparation of GPMC-GDVs was as follows 0.4 mg/mL GDVs was slowly dripped into GPMC solution with equal volume of Mel containing 2 mg/mL and incubated at room temperature for 30 min. As-prepared GPMC-GDVs had good stability in PBS, DMEM, and 10% FBS. The results of MTT method showed that GPMC-GDVs had a better tumor inhibitive effect. Conclusion By extracting the components of grape GP and GDVs and reorganizing the structures, the vesicles can be prepared for the loading of active melittin, which has a good application prospect in the field of delivery and anti-tumor effect of polypeptide drugs.