The Extraction, Isolation, Purification And Content Determination Of The Active Components From Three Chinese Traditional Drugs

The paper studied the extraction, isolation, identification and content determination of RMC and QA from Rubia cordifolia L., and established the method of content determination of rutin and amentoflavone from Forsythia suspense and Selaginella tamariscina(Beauv.)Spring respectively.Studies on RMC: material ratio and the weight of dry rubia material (V/W) and time have been investigated for their influences on the extraction rate (%) of RMC. Oputimum combinations of extraction conditions were obtained using the L9(34) orthogonal analysis technique. The result showed that the material ratio had the most evident influence. It was found that an extraction rate of 0.8 % could be achieved at petroleum. A material ratio of 20:1 and an extraction time of 2 h and 2 times were efficacy.All kinds of silica gel column chromatography including nomal pressure and decompression and Sephadex LH-20 chromatography were used to isolate RMC in a sequence order. MS, 1H NMR and 13C-NMR were used to identify the structure of RMC. RP-HPLC (reversed-phase high-performance liquid chromatography) method was used to determine the content of RMC. Seven samples from seven regions including Luanchuan, Sichuan, Hubei, Guangxi, Guizhou, Henan (baishikang) and Shanxi. HPLC analysis conditions were the Purospher star column (250 mm×4.6 mm, 5μm, Merk) at 25℃using methanol-water-Tetrafuran (90:9.3:0.7) as mobile phase with a flow rate of 1.0 ml/min and detection wavelength at 250 nm. There was good linear relationship between the peak area and the sample content injected at the ranges of 0.200～2.00μg (r=1) for RMC. The result showed that the content of RMC for different samples were 1.23 % (Luanchuan), 0.77% (Sichuan), 0.75% (Hubei), 0.60 % (Guangxi), 0.37 % (Guizhou) and 0.32 % ( Henan baishikang). The sample from Shanxi had a shoulder peak and couldn't be quantified.Studies on QA: the roots of R. cordifolia were extracted using acetone-water (7:3, V/V). After evaporation of the solution, the residue was suspended in water and extracted with petroleum, ethyl acetate and n-butanol. The n-butanol extract was isolated for QA.All kinds of silica gel column chromatography including nomal pressure and decompression and Sephadex LH-20 chromatography were used to isolate QA in a sequence order. MS, 1H NMR and 13C-NMR were used to identify the structure of RMC. RP-HPLC (reversed-phase high-performance liquid chromatography) method was used to determine the content of QA. Six samples from six regions including Luanchuan, Sichuan, Hubei, Guangxi, Guizhou and Shanxi. HPLC analysis conditions were the Purospher star column (250 mm×4.6 mm, 5μm, Merk) at 25℃using methanol-water-tetrafuran (65:34.7:0.3) as mobile phase with a flow rate of 1.0 ml/min and detection wavelength at 276 nm. There was good linear relationship between the peak area and the sample content injected at the ranges of 0.020～0.160μg (r=0.9998) for QA. The result showed that the content of QA for different samples were 0.42 % (Luanchuan), 0.053 % (Shanxi). QA in other samples from other regions couldn't be seperated well.Content determination of rutin: RP-HPLC (reversed-phase high-performance liquid chromatography) method was used to determine the content of rutin extracted from F. suspensa. HPLC conditions were the Purospher star column (250 mm×4.6 mm, 5μm, Merk) at 25℃using methanol-2% glacial acetic acid (42:58) as mobile phase with a flow rate of 0.8 ml/min and detection wavelength at 359 nm. There was good linear relationship between the peak area and the sample content injected at the ranges of 0.68～6.8μg (r=1) for rutin.Content determination of amentoflavone: RP-HPLC (reversed-phase high-performance liquid chromatography) method was used to determine the content of amentoflavone from S. tamariscina(Beauv.)Spring and S. pulvinate(Hook.Et Grev.)Maxim. HPLC conditions were the Purospher star column (250 mm×4.6 mm, 5μm, Merk) at 25℃using methanol- 0.1% phosphoric acid (65:35) as mobile phase with a flow rate of 0.8 ml/min and detection wavelength at 337 nm. There was good linear relationship between the peak area and the sample content injected at the ranges of 0.038～0.342μg (r=0.9999) for amentoflavone.