| Cornus officinalis is a traditional Chinese medicine with a wide range of biological activities. In this study, the novel microwave system was first used to extract active constituents from Cornus officinalis. We systemically investigated several extraction parameters, including the concentration of ethanol, the ratio of material to solvent, the extraction time and system temperature on the effect of extraction of active constituents. Furthermore, the present study also evaluated the hypoglycemic activity of constituents extracted from Cornus officinalis. The main contents are as follows:1. In order to optimization the extraction condition for hypoglycemic active constituents, the single factor experiment and response surface experiment was designed. The three hypoglycemic active constituents-iridoid glycosides, polyphenol and pentacyclic triterpene acids-were used as the response value. The concentration of ethanol, the ratio of material to solvent, extraction temperature and extraction time were selected as the impact factors. The optimum extraction condition were found to bethe concentration of ethanol 30%;the solid/liquid ratio1:11; extraction temperature 90℃; extraction time 7.7 min. The maximum value of total iridoid glycosides, total polyphenols and total pentacyclic triterpene acids obtained by the validation test was 152.71±2.17mg/g, 51.89±1.52mg/g and 34.11±0.94mg/g, respectively. What`s more, to control the extraction temperature is more accurate and effective than to control the microwave power. In this study, the amount of ethanol and extraction time were much lower than the traditional microwave extraction method, which can ensure the stability of the extracts.2. The crude extracts were separated by macroporous resin after extracted by different polar solvent. The α-glucosidase inhibition test was designed to study the hypoglycemic activity of different extract components in vitro. The results showed that the n-butanol fraction(Fr. Bu), ethyl acetate fraction(Fr. Et) and water fraction had a high inhibitory activity to α-glucosidase. In the concentration of 1mg/mL of 30% of Fr. Bu(Fr. Bu-2) and 30% of Fr. Et(Fr. Et-2), the α- glucosidase inhibition ratio was 74.4% and 73.6%, respectively. It is much higher than the positive control group of 58%. Kinetic experiments showed that the Fr. Bu-2 inhibition was non competitive inhibition type. Competitive inhibition type for the Fr. Et-2. HPLC analysis showed that the main component of Fr. Bu-2 was loganin and morroniside. Ursolic acid and oleanolic acid was he main component of Fr. Bu-2.3. The HepG2 cell glucose intake experiments were designed to study hypoglycemic activity of different components from Cornus officinalis in vitro. The results showed that the Fr. Bu-2, Fr. Et-2, loganin, morroniside, ursolic acid and oleanolic acid significantly promoted the glucose intake of HepG2 cells with a dose dependent manner. In the concentration of 0.5mg/mL, the glucose intake of HepG2 cells for Fr. Bu-2, Fr. Et-2, loganin, morroniside, ursolic acid and oleanolic acid was about 2.76 times, 2.87 times, 2.97 times, 1.41 times,2.13 times, 2.37 times of the control group respectively and much better than the positive control group metformin hydrochloride.4. The HepG2 insulin resistance cell model was established to study the glucose intake of different components from Cornus officinalis. Results showed that the Fr. Bu-2, loganin, morroniside and oleanolic acid could promote glucose intake of model cells in high concentration. In the concentration of 0.5mg/mL, the glucose intake of model cells for Fr. Bu-2, loganin, morroniside, ursolic acid and oleanolic acid was about 1.88 times, 1.61 times, 1.09 times, 1.23 times, 1.27 times of the control group respectively. |
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