Study On The Antioxidative Effect Of Quercetin From Actinidia Arguta And The Antitumor Effect And Toxicity Of Doxorubicin Liposome Containing Quercetin And Vitamin C

Actinidia arguta Sied.et Zucc is a perennial deciduous vine mainly distributed in northeast Asia.The fruit of Actinidia arguta has good taste and high nutritional value which contains a lot of vitamin C and other nutrients.The whole plant of Actinidia arguta can be used as medicine,such as flavonoids,polysaccharides and other active components of antioxidative,anti-tumor and other bioactive effects.Following extraction and identification,it has been found that the main flavonoid in the fruits of Actinidia arguta are quercetin and quercetin glycoside compounds.Quercetin has a variety of pharmacological effects such as antioxidative and anti-tumor effects,but its low bioavailability limits its application.The nano-preparation can improve its bioavailability.Vitamin C is a natural antioxidant that has been shown in vitro to be toxic to tumor cells at high doses and has been found to play a role in the prevention and treatment of cancer in clinical studies.In this study,we extracted quercetin from the fruit of Actinidia arguta and evaluated its antioxidant capacity,and then we prepared quercetin-vitamin C-doxorubicin co-loaded lipsomes to evaluate the effects of quercetin and vitamin C on the safety and anti-tumor effect of doxorubicin.The main research conclusions of this paper were as follows:1.The quercetin in the fruit of Actinidia arguta was extracted by the joint use of compound pectinase hydrolysisby and ultrasonic assisted and the single factor experiments and response surface method were applied to determine the optimum process conditions of this extraction method on quercetin.The results showed that the optimum conditions were as follows:compound pectinase 16.0mg?30000U/g?,enzymolysis p H 4.5,enzymolysis temperature 45?,enzymolysis time 60min,60%ethanol as solvent,solid-liquid ratio of 1:4,ultrasonic power 200W,ultrasonic time 4min,the extraction temperature 60?and extraction time 30min.Under this condition extraction rate was 0.243mg/g?fresh fruit weight?.2.The antioxidative effect of quercetin from Actinidia arguta in vitro and in vivo was studied by compared with vitamin C.The total antioxidant capacity?T-AOC?and the capacuity of inhibitory activity of lipid peroxidation were used to detect and analyze the antioxidative effect of quercetin in vitro.The results were shown as follows:there was no significant difference in the total antioxidant capacity between quercetin and vitamin C at the same molar concentration in vitro,and the antioxidant capacity was positively correlated with the concentration within the experimental dose range;the IC₅₀of quercetin and vitamin C against lipid peroxidation was 0.79mg/m L and 1.41mg/m L,respectively.The anti-lipid peroxidation ability of quercetin was significantly higher than that of vitamin C,and quercetin and vitamin C could play a synergistic role in anti-lipid peroxidation.The antioxidative effect of quercetin in vivo was detected by liver injury model test in mice induced by CCl₄,and the results showed that the antioxidant capacity of quercetin was significantly better than that of vitamin C in vivo,and when they used together,they had a synergism effect on anti-oxidation.Quercetin and vitamin C could restore the damage induced by CCl₄on liver,kidney and blood lipids in mice.When combined with vitamin C,quercetin had a synergism effect on relieving the oxidative damage.3.In this study,the quercetin-vitamin C-doxorubicin co-loaded liposomes was prepared by the membrane dispersion method and doxorubicin loading into liposomes driven by vitamin C.Determined by single factor experiment,the optimum conditions to prepare the liposomes were concluded as follows:doxorubicin lipid ratio 1:5.5?m/m?,incubation p H 7.4,incubation time 15min.The quercetin-vitamin C-doxorubicin co-loaded liposomes were prepared under the conditions above,and the average particle size of the liposomes was128.83±20nm,the form of the liposomes was complete and spherical,and encapsulation efficiency of doxorubicin was 97.33±1.23%.4.The effect of co-loaded lipsome on the toxicity of doxorubicin was evaluated by measuring the survival time of mice after given doxorubicin in large dose and detecting the LD₅₀.The results of toxicity test in mice showed that the co-liposome could prolong the average survival time to 23.9 days and reduce the acute toxicity of doxorubicin.The results analysed by Karber's-method showed that the co-loaded liposomes increased the LD₅₀of doxorubicin to 41.34mg/kg body weight.5.The mouse model of liver cancer induced by H22 cells was used to evaluate the effects of co-loaded liposomes on improving the anti-tumor effect of doxorubicin.The tumor inhibition rates of high and low doses of co-loaded liposomes were 53.78%and 68.93%respectively,which were significantly higher than other groups,reflecting a synergistic effect on anti-tumor.The results of thymus index and spleen index showed that the co-loaded liposomes reduced the immunotoxicity of doxorubicin.The blood biochemistry index of mouse model of liver cancer demonstrated that the co-loaded liposomes had significant synergistic effect on the recovery of liver and kidney injury caused by doxorubicin.6.To study the effect of co-loaded lipsomes on the pharmacokinetics of doxorubicin,the concentration of doxorubicin in plasma and organs of mice were detected by HPLC,and the relationship curve between doxorubicin concentration and time was drawn.The concentration of doxorubicin in plasma decreased gradually over time,and it could not be detected in the plasma of free doxorubicin gruop at 2880min.The concenrrations of doxorubicin in the plasma of the doxorubicin lipsome and co-loaded lipsome gruops were 0.019±0.005?g/m Land0.028±0.007?g/m L respectively at the same time.This results above showed that the co-loaded liposmes could prolong the circulation time of doxorubicin in vivo.The results of concenrrations of doxorubicin in heart,lung,liver and spleen tissues showed that the co-loaded liposome could reduce the distribution of doxorubicin in various organ tissues,and reduce the damage to organs.