Inhibition Of α-glucosidase By Rosa Davurica Pall. Flavonoids And The Study Of Its Nanoparticles

Rosa davurica pall.is rich in polyphenols,flavonoids,saponins,and other chemical components,and also has good anti-fatigue,antioxidant,anti-tumor,and other physiological activities.α-glucosidase is a key enzyme involved in the breakdown of glucose in the intestine,and inhibition of this enzyme may be a key strategy to control hyperglycemia.α-glucosidase inhibitors can control blood glucose levels in the body,and natural plant flavonoids show inhibitor drug potential.The aim of this paper is to study the optimization of the extraction process of Rosa davurica pall.flavonoids,the inhibition kinetics and inhibition mechanism study of Rosa davurica pall.flavonoids and Hypericin,and the physicochemical properties and biological activity study of Hypericin nanoparticles,which provide a basis for further development and utilization of natural products,and provide a theoretical basis for the development and application of inhibitors and functional ingredients of food.The specific results of the study are as follows:(1)The linear regression equation of the rutin standard was:y=0.259x-0.0411,R~2=0.9991,with good linearity when the flavonoid content was determined by the aluminum nitrate color development method.The optimum process conditions were obtained by ultrasonic-assisted enzyme extraction method for the extraction of flavonoids from Rosa davurica pall.The theoretically predicted value of flavonoid extraction rate was 29.381 mg/g when single-factor and response surface experiments were conducted:p H 5.127,enzymatic digestion time 123.886 min,enzymatic digestion temperature 48.99℃,and ultrasonic time 29.548 min.The modified conditions were verified:p H 5.1 The extractivity of flavonoids was 28.841 mg/g with the modified conditions:p H 5.1,124 min of enzymatic digestion time,49°C and 29.5 min of sonication time,which was basically consistent with the theoretical prediction.After purification by adsorption with macroporous resin,the purity of flavonoids obtained was 75.78±0.928 mg/g,which was 2.63 times higher than that before purification.(2)The IC50 values of total flavonoids of Rosa davurica pall.against DPPH and ABTS radicals were 64.111 and 21.179μg/ml,respectively,and the IC50 values of positive control vitamin C were 3.438 and 7.282μg/ml,respectively,which proved the antioxidant property of total flavonoids.Enzyme kinetic experiments showed that total flavonoids bound toα-glucosidase in a reversible manner,inhibitingα-glucosidase in a non-competitive manner with an IC50 value of 112.5±4.63μg/ml.the value of the Mee’s constant(Km)was 2.90±0.12 mg/ml.Fluorescence spectroscopy analysis showed that total flavonoids quenched the intrinsic fluorescence ofα-glucosidase,and it was inferred from the n value approximately equal to 1 that total The total flavonoids had a single binding site forα-glucosidase.Simultaneous fluorescence and three-dimensional fluorescence analysis showed that total flavonoids were able to induce secondary structure changes ofα-glucosidase and alter the microenvironment ofα-glucosidase mainly through the interaction with Tyr and Trp.(3)The IC50 values of DPPH and ABTS radical scavenging by hypericin were9.845 and 4.376μg/ml,respectively.hypericin inhibitedα-glucosidase with an IC50value of 77.28±2.68μg/ml and acarbose with an IC50 value of 38.44±1.26μg/ml.hypericin is a non-competitive inhibitor,and binding to the enzyme is reversible.According to Lineweaver-Burk plots,Km and Ki values were 4.873±0.34 mg/m L and46.95μg/m L,respectively.combined with fluorescence quenching and thermodynamic analysis,auriculosides exhibited potential enzyme inhibition by inducing a change in the secondary structure conformation of the enzyme,and an n value equal to 1 indicated that auriculosides had a single binding site forα-glucosidase.The value of Ksv decreased from 16.85 mg/m L to 12.71 mg/m L when the temperature was increased from 298 to 310 K,indicating that the process of fluorescence quenching was dominated by a static quenching mechanism.ΔG was negative and the values ofΔH andΔS were positive,which indicated that both were bound by intermolecular forces dominated by hydrophobic interactions.Simultaneous fluorescence and molecular simulation analysis showed that auriculosides modify the microenvironment ofα-glucosidase by interacting with Tyr and Trp and are able to interact with amino acids near the active center,further identifying auriculosides as a high-affinity binding site onα-glucosidase.(4)The particle size of whey protein/konjac glucomannan nanoparticles was227±8.14 nm with a PDI of(0.263±0.014)and a potential of-32.77±0.3 m V.After loading with hypericin,the particle size increased to 299±7.8 nm and the potential changed to-30.73±0.6 m V,with an encapsulation rate of 91.61%.Scanning electron microscopy observed a significant change in the appearance of the complex with larger hydrophobic cavities.FTIR analysis indicated that the flavonoids were encapsulated by nanoparticles and the stability of the loaded nanoparticles was maintained mainly by hydrogen bonding,hydrophobic and electrostatic interactions.Combined with X-ray diffraction and DSC results,it was inferred that auriculosides were encapsulated by nanoparticles and belonged to the amorphous state of binding.After digestion by simulated gastrointestinal fluid,the release rate of the auriculoside complex was 11.32%in gastric fluid and 90.09%in intestinal fluid.The scavenging ability of aurantioside complex on DPPH and ABTS radicals increased with gastrointestinal digestion,and the percentage inhibition ofα-glucosidase was 89.17%when the simulated digestion was over.