| Hypericum perforatum L.(St. John’s wort) is a traditional Chinese herbal medicine. It has significant roles in diseases treatments, such as depression, cancer, viral infection, bacterial infection, inflammation and so on. Melatonin as one of its main active components is one of the strongest endogenous free radical scavengers ever found, and has a very important physiological role in the body. It can improve people’s sleep, improve the body’s anti-oxidation ability, anti-aging ability, and improve the immune system. Establishing a stable and accurate determination method of melatonin has guiding significance for the study of the role of melatonin in plants. Studies have shown that melatonin has a role in reducing blood glucose, which may be related to its strong free radical scavenging ability. There is a link between diabetes and oxidative stress. Antioxidant therapy can alleviate oxidative stress in the body, thus playing some role in the prevention or delay of diabetes and its complications. Therefore, the choice of an effective anti-oxidant drug and the study of its role on the prevention and treatment of diabetes and its complications are of great significance. The purpose of this study is to establish a stable, accurate detection method of melatonin, and determine the content of melatonin in St. John’s wort. And further study the hypoglycemic effect of H. Perforatum, explore whether the H. Perforatum extract and melatonin can reduce the level of oxidative stress in the diabetic body. The main contents of the thesis are as follows.1. A stable and accurate determination method of melatonin was established. Use Phenomenex C18column(5μm,4.6mm×250mm), mobile phases of0.2%(v/v) glacial acetic acid aqueous solution, acetonitrile and methanol at30℃, a flow rate of1.0mL/min,223nm to determine.The Repeatability, precision and sample stability of this method was proved to be good, and there was a good linear in the range of0.1μg-1.5μg. The recovery was99.6%and RSD was1.86%. Use this method to determine the content of melatonin in the wild H. Perforatum and H. Perforatum extract.2.4-6week-old male Kunming mice were randomly divided into normal control group (NC), diabetic model group (DM), positive control group (PC), Hypericum perforatum extract group(HYE) and melatonin group(MT), Hypericum perforatum group and melatonin group, depending on the concentration were divided into low (L), medium (M), high (H) groups, each group including10mice. After adaptive feeding for4days, in addition to the NC group, other groups were made diabetic model by intraperitoneal injection of the alloxan solution150mg/kg-bw0.2mL, the NC group was injected an equal volume of ultra-pure water. In the course of the experiment, all mice were fed and drank water free, were not given other auxiliary hypoglycemic drugs. The HYE groups and MT groups were administered by gavage the corresponding concentrations of Hypericum perforatum extract and melatonin respectively, the PC group was given an equal volume of drug solution, NC group and DM group were given an equal volume of ultra-pure water by gavage, once a day,0.2mL per time for3weeks. In the beginning and the end of the experiment, the mice were tested in blood glucose and weighed. After the end of the experiment, the mice were killed to measure the weight of stress indicating organs (liver, kidney) for each group and antioxidant enzymes (SOD, GSH-PX, CAT) activity, malondialdehyde (MDA) content, glutathione (GSH) levels and nitric oxide (NO) content, nitric oxide synthase (NOS) activity in the liver, kidney respectively. The data was analyzed by SPSS software.Results:1. The extraction and purification method of melatonin is good, and the HPLC detection method is precision and stable, and Hypericum perforatum presents a higher content of melatonin;2. Hypericum perforatum extract has a significant hypoglycemic effect, and a role in mitigation for diabetes-induced weight loss in mice;3. Hypericum perforatum extract can alleviate liver and kidney’s hypertrophy and pathological changes in diabetic mice;4. Hypericum perforatum extract can improve the activity of antioxidant enzymes(SOD, CAT) and GHS levels in the organization of diabetic mice, and reduce the oxide content(MDA, NO), Inhibit the activity of NOS. The results lay the foundation for the further study of function and content variation of melatonin in the plants, provide research data for the pharmacological effects of antioxidant stress of H. Perforatum and antioxidant therapy of diabetes. |
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