Research On Determination And Antioxidant Activity Of Tannins From The Roots Of Agrimonia Pilosa Ledeb

Agrimony roots are the roots of a perennial herb Agrimony(Rosaceae). In our country, Agrimony is rich in resources, widely distributed, which is widely used for the treatment of clinical gynecology, respiratory system, urinary system and digestive system diseases. But the utilization rate isn’t very high. Because tannin compounds have anti-tumor, anti-oxidation, enhance immunity and other pharmacological effects, there’re more researches on tannins in Agrimony in recent years. But the study on tannins of Agrimony roots is barely reported. So the study on tannins of Agrimony roots and the effect on anti-oxidant for the full development and utilization of Agrimony resources has important significance.First, the method to extract, separate and authenticate the tannins from Agrimony roots is followed.70% acetone and 8 times the amount as solvent, treat roots with ultrasonic in power 160W and temperature 25℃ for 1 hour, then discard residue and recovered acetone after filtration with vacuum filtration. The diethyl ether, ethyl acetate, water saturation of n-butyl alcohol in the proportion of 1:1 is used to extract with recovered acetone 3 times and be recovered, then get what we need.Casein and HPLC are used to determine the content of tannins in Agrimony roots. The result of Casein method shows that the content of tannins in the acetone layer is 61.36%, in ether layer is 2.01%, in the ethyl acetate layer is 63.25%, in n-butyl alcohol layer is 28.17%. The result of HPLC method shows that the content of gallic acid in the acetone extract is 57.14%, in ether layer is 1.84%, in ethyl acetate layer is 58.72% and in n-butyl alcohol layer is 22.59%.Then, through the two aspects of experiments in vivo and in vitro study on antioxidant activity of Agrimonia roots. In vitro experiments, Vc or gallic acid is used as control group, to explore the effects of tannins in Agrimony roots in eliminating superoxide anion free radical and DPPH free radical, determination of liposome peroxide activity and the inhibition of oil peroxide activity. The results show that in the superoxide anion radical scavenging experiment, scavenging ability of tannins in Agrimony roots and Vc are enhanced with increasing concentration among the effective concentration range of 0.1-0.9mg/mL.Vc scavenging activity on superoxide anion is slightly higher than tannins in Agrimony roots. In the DPPH radical scavenging experiment, scavenging ability of tannins in Agrimony roots and Vc are enhanced with increasing concentration among the effective concentration range of 1-9μg/mL. Vc scavenging activity on superoxide anion is lower than tannins in Agrimony roots. In the experiment of exploring the inhibition of liposome peroxide activity, tannins extracted from Agrimony roots has strong inhibition effect on lipid peroxidation, the inhibition is higher than the same concentration of vitamin C, and with the mass concentration enhanced increased. In the experiment of exploring the inhibition of oil peroxide activity, oil peroxide value (POV) and malondialdehyde (MDA)absorbance value of the indexes results show the effect of inhibition of tannins extracted from Agrimony roots on lipid oxidation is significant.Tannins in Agrimony roots are researched on the influence on D-galactose in male mice in vivo study. Except the blank group, the remaining mice are intraperitoneal injection of D-galactose for 4 weeks.Blank group is injected with physiological saline. Establishment of subacute aging model mice. After modeling, use Vc as the positive control group. Agrimony roots tannins extract low、middle and high dose group gastric perfusion 100、200 and 300mg/(kg·d). D-galactose model group and blank group receive normal saline.4 weeks later, do the research on the eye blood measurement in serum of mice SOD and MDA content. Results indicate that the effect of these three doses of tannins extracted from Agrimony roots in turn is the middle dose group,the low dose groupand the high dose group. And the effect of the middle dose group is better than the effect of the positive control group. Therefore,with appropriate dose extractive of tannins from Agrimony roots can increase the vitality of SOD and reduce the vitality of MDA, which means that the tannins in Agrimony roots have good ability on treating subacute aging model mice of D- galactose of antioxygenation.In conclusion, the research on the extraction, separation, determination of Agrimony roots tannins and its antioxidant activity, not only can be as the basis for the study of Chinese medicine tannins on pharmacology research, but also can bring new ideas on looking for non-toxic natural antioxidants and fully developing Agrimonia pilosa resources.