Study On Purification Of Reveratrol From Peanut Root, Preparation And Antitumor Activity Of Reveratrol Nanoliposomes

Resveratrol, a natural polyphenolic compound, has been reported to have a number of biological andpharmacological activities including anticancer, anti-microbial, anti-inflammatory, neuro-andliver-protective, which caused a great deal of attention in recent years. However, the poor watersolubility and stability of RSV results in low bioavailability and absorptivity. Nanoliposomes, aseffective drug delivery vehicles, have the bioactives including promoting stability, enabling precisiontargeting and controlled-releasing for the encapsulated material. The application of nanoliposomes inencapsulating and deliverying reaveratrol could improve its bioavailability and controlled-releasingeffect, which could extend its application in food, cosmetic and pharmaceutical industries. Theextraction and purification for resveratrol from peanut root were studied, the preparation of resveratrolnanoliposomes by high pressure microfluidization was optimized, and the characterization andanti-tumor activity of reaveratrol nanoliposomes were evaluated in this research. The main conclusionswere summarized as follows:The total content of resveratrol in peanut root were165.64±0.38μg/g, determined by repeatedlysolvent extraction. After solvent extraction for three times, the extraction rate was89.02±0.28%. Theoptimum separation conditions were studied by static and dynamic adsorption and desorptionexperiments, which were as follows: polyamide as macroporous resin, sample concentration26.58μg/mL, eluent80%ethanol, desorption flow rate2BV/h. Based on the thin-layer chromatography andsilica gel column chromatography experiments, it was found that the crude extracts could be purified onthe conditions of eluent chloroform/methanol=40:1(v/v), flow rate30mL/min, silica gel:sample=1:1(m/m). After recrystallization by methanol/chloroform=3:1for three times, the purity ofresveratrol was97.01±0.45%, determined by HPLC.On the basis of single factor experiments, the technology of preparing resveratrol nanoliposomes byhigh pressure microfluidization method was optimized by quadratic regression rotation designexperiment. The results showed that the optimum preparation conditions were as follows: the mass ratioof lectine to vitamin E10:1, lecithin to resveratrol11.6:1, lecithin to cholesterol10.5:1,microfluidization pressure18366PSI and cycles3times. Under these conditions, the encapsulationefficiency of reveratrol nanoliposomes was as high as87.74±1.01%. Finally, the nanoliposomes werelyophilized with5%(m/v) cryoprotectant, sucrose and mannitol (1:1m/m) with homogeneous color,good reconstitution properties and87.67±0.76%encapsulation effiency.The characterization of resveratrol nanoliposomes including physicochemical properties, such asmicrostructure, particle size, Zeta potential and pH, and stability at different temperature, pH andstorage conditions, was evaluated. The appearance was well-distributed suspension with lacte color. Themicrostructure observed by transmission electron microscope was that resveratrol encapsulated orinserted in the lipid bilayer of nanoliposomes with spherical and elliptic shape. The average particle sizewas (78.31±1.37) nm and the Zeta potential was-55.5mV. It was stable at pH7-8, unstable at acidicconditions. The leakage rate increased with the temperature rise. The resveratrol nanoliposomes were stable at4℃for50d, with the leakage rate as low as2.78±0.26%.The controlled-releasing and antitumor effect of resveratrol nanoliposomes were evaluated bysimulated release experiment and cell culture test. Dynamic dialysis experiment showed that reseratrolnanoliposomes released for26h,13times of free resveratrol, which indicated significant delayedreleasing effect. The releasing process was fitting for Nieberhull equation (1-Q)1/2=-0.033t+0.954(r2=0.997), indicating that RSV was released by diffusing and skeleton dissolution. The cell experimentsin vitro demonstrated that the inhibition ratio of resveratrol nanoliposomes to hepatoma carcinoma cellHepG2and gastric carcinoma cell AGS were77.36%and72.64%respectively, higher than resveratrolgroup and resveratrol with blank nanoliposomes group, indicating a good antitumor effect.