| Scutellaria baicalensis Georgi(S. baicalensis) is the dried roots of labiatae Scutellaria baicalensis Georgi. It is widely distributed in nature, mainly produced in Hebei, Shanxi, Liaoning and other places in China. S. baicalensis has been widely used in the treatment of cancer, infectious diseases, inflammation and so on. The main active constituents of S. baicalensis are flavonoids, such as baicalin, wogonoside, baicalein, wogonin and so on, they had been investigated and were found to have the properties of anti-hepatitis B virus, exert anti-inflammatory, antioxidant, anti-allergic, anti-tumor and anxiolytic. However, the traditional methods to extract flavonoids from S. baicalensis are mostly decocting, refluxing and ultrasonic method, which lead to high organic solvent loss, time consuming and low extraction efficiency. Therefore, it has been a focus to pursue the method for extracting of the flavonoids efficiently, quickly and less pollution.Ionic liquids, which are composed of bulky organic cations and inorganic or organic anions, are liquid near room temperature. They have showed a great promising prospect in many areas on account of their unique physical and chemical properties:negligible vapor pressure, wide liquid range, tunable viscosity, good thermal stability, miscibility with organic solvents and water, as well as the ability of efficiently transferring and absorbing microwave energy. Due to the good extractability and solubility for various organic compounds, ionic liquids have been used as the extraction solvent successfully in the extraction and analysis of different types of natural products at present. As a typical kind of ILs, imidazolium-based ionic liquids consist of various N, N’-dialkylimidazolium cations, and a wide range of halide-based anions. Due to the easy preparation and the easy regulation of the structure, imidazolium-based ILs have been one of the most promising ILs. According to our surveys of the literatures, there were not any reports about the extraction of flavonoids from S. baicalensis using imidazolium-based ionic liquids yet.In the present work, a series of 1,3-dialkylimidazolium-type ILs were assessed for extraction efficiency, and 1-octyl-3-methylimidazolium bromide([C8mim]Br) was selected as the optimal solvent. The single factor experiment and orthogonal test were used to optimize the ionic-liquids microwave-assisted (ILMAE) procedures systematically, according to the determination of the four flavonoids in the extracts. Then, the proposed ILMAE approach was compared with the pharmacopeia method and exhibited good results. The preliminary mechanism of 1-octyl-3-methylimidazolium bromide based microwave-assisted extraction([C8mim]Br MAE) of baicalin was discussed according to the solubility in different agents and the cell structure change of S. baicalensis before and after extraction, as well as the inhibition of [C8mim]Br against hydrolase of baicalin. At the same time, a stable and reliable HPLC analysis method had been established, in order to determine the four flavonoids in S. baicalensis simultaneously.1. Studied on the extraction technology of S. baicalensis:A series of 1,3-dialkylimidazolium-type ionic liquids with different kinds of cations and anions were studied and various influential parameters of the ILMAE procedures were optimized with orthogonal test systematically, according to the determination of baicalin, wogonoside, baicalein, wogonin in the extracts. Then, the proposed ILMAE approach was compared with the pharmacopoeia method. The results showed that the amount of the four flavonoids in the extract have significant changes with different extraction conditions. Considering the effect of various factors, the optimum ILMAE conditions were found to be:[C8min]Br concentration of 1.0 M, irradiation power of 400 W, temperature of 80 ℃, irradiation time of 90 s, solid-liquid ratio of 1:6(g/mL), and one extraction cycle. Compared with the pharmacopoeia method, the optimum method provided a little higher extraction yield and less time.2. Preliminary exploration of the mechanism of 1-octyl-3-methylimidazolium bromide based microwave-assisted extraction of baicalin:The microstructures of S. baicalensis powders were observed before and after extraction with the help of a scanning electron microscope (SEM) in order to explore the extraction mechanism, and the activity of the crude enzyme solution from S. baicalensis was determined through the hydrolysis of baicalin. The results showed that the proposed approach provided 0.79% higher extraction yield and less time (from 3 h to 5 min) than the traditional pharmacopoeia extraction method. The solubility of baicalin in [C8mim]Br solution was better than in 70% ethanol and distilled water. The pictures of scanning electron microscopy of the S. baicalensis samples cell before and after extraction by [C8mim]Br showed that [C8mim]Br has a strong ability of breaking the cell. Moreover, [C8mim]Br solution also could inhibit the activity of the baicalin hydrolase in S. baicalensis.3. The quantitative determination of flavonoids in S. baicalensis:A stable and reliable HPLC analysis method had been established, in order to determine baicalin, wogonoside, baicalein and wogonin in S. baicalensis simultaneously. To evaluate the method, linearity, precision, repeatability, stability and recovery were investigated under the optimized conditions. The results showed that the method was simple and credible, and can be used for the determination of the content of effective components in S. baicalensis. |
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