Effects of Nodakenin, Columbianadin, and Umbelliferone Isolated from the Roots of Angelica decursiva on the Gene Expression and Production of MUC5AC Mucin from Human Airway Epithelial NCI-H292 Cells

Angelica decursiva has been utilised as remedy for controlling the airway inflammatory diseases in folk medicine. We investigated whether nodakenin, columbianadin, and umbelliferone isolated from the roots of Angelica decursiva inhibit the gene expression and production of MUC5AC mucin from human airway epithelial cells. Confluent NCI-H292 cells were pretreated with nodakenin, columbianadin or umbelliferone for 30 min and then stimulated with epidermal growth factor (EGF), phorbol 12-myristate 13-acetate (PMA) or tumor necrosis factor-α (TNF-α) for 24 h. The MUC5AC mucin gene expression was measured by reverse transcription - polymerase chain reaction (RT-PCR). Production of MUC5AC mucin protein was measured by enzyme-linked immunosorbent assay (ELISA). The results were as follows: (1) Nodakenin did not affect the expression of MUC5AC mucin gene induced by EGF, PMA or TNF-α. Columbianadin inhibited the expression of MUC5AC mucin gene induced by EGF or PMA. However, umbelliferone inhibited the expression of MUC5AC mucin gene induced by EGF, PMA or TNF-α; (2) Nodakenin also did not affect the production of MUC5AC mucin protein induced by EGF, PMA or TNF-α. Columbianadin inhibited the production of MUC5AC mucin protein induced by PMA. However, umbelliferone inhibited the production of MUC5AC mucin protein induced by EGF, PMA or TNF-α. These results suggest that, among the three compounds investigated, umbelliferone only inhibits the gene expression and production of MUC5AC mucin stimulated by various inducers, by directly acting on airway epithelial cells, and the results might explain the traditional use of Angelica decursiva as remedy for diverse inflammatory pulmonary diseases.

Simultaneous Determination of Coumarin Compounds in Yifu Pill by HPLC / 中国药房

OBJECTIVE:To establish a method for the simultaneous determination of coumarin compounds in Yifu pill. METH-ODS:HPLC was performed on the column of Inertsil ODS-3 with mobile phase of acetonitrile-water(49∶51,V/V) at a flow rate was 1.0 ml/min,the detection wavelength was 320 nm,column temperature was 30 ℃,injection volume was 20 μl. RESULTS:The linear range was 5.16-206.4μg/ml for osthole (r=0.999 9),37.40-1 495.89μg/ml for isoimperatorin(r=0.999 9) and 9.95-318.34 μg/ml for columbianadin(r=0.999 9);RSDs of precision,stability and reproducibility tests were lower than 3.0%;recov-eries were 96.05%-103.19%(RSD=2.18%,n=9) for osthole,96.90%-103.09%(RSD=2.07%,n=9) for isoimperatorin, 95.50%-103.57%(RSD=2.15%,n=9)for columbianadin. CONCLUSIONS:The method is simple with good precision,stability and reproducibility,and can be used for the simultaneous determination of coumarin compounds in Yifu pill.

Columbianadin Inhibits Cell Proliferation by Inducing Apoptosis and Necroptosis in HCT116 Colon Cancer Cells

Columbianadin (CBN), a natural coumarin from Angelica decursiva (Umbelliferae), is known to have various biological activities including anti-inflammatory and anti-cancer effects. In this study, the anti-proliferative mechanism of actions mediated by CBN was investigated in HCT-116 human colon cancer cells. CBN effectively suppressed the growth of colon cancer cells. Low concentration (up to 25 μM) of CBN induced apoptosis, and high concentration (50 μM) of CBN induced necroptosis. The induction of apoptosis by CBN was correlated with the modulation of caspase-9, caspase-3, Bax, Bcl-2, Bim and Bid, and the induction of necroptosis was related with RIP-3, and caspase-8. In addition, CBN induced the accumulation of ROS and imbalance in the intracellular antioxidant enzymes such as SOD-1, SOD-2, catalase and GPx-1. These findings demonstrate that CBN has the potential to be a candidate in the development of anti-cancer agent derived from natural products.

Comparative Study on the Main Medical Component Contents of Angelica pubescens in Famous Region Chongqing and Other Domestic Areas in China / 中国药房

OBJECTIVE:To compare the contents of osthole and columbianadin in Argelica. pubescens in Wushan county, Wuxi county in Chongqing and other domestic areas in china. METHODS:HPLC was performed on the column of Shiseido Cap-cell Pak C18 with mobile phase of acetonitrile-water(gradient elution) at flow rate of 1.0 ml/min,detection wavelength was 330 nm,column temperature was 30 ℃ and volume injection was 10 μl. RESULTS:The linear range was 0.015 1-0.301 6 mg/ml(r=0.999 9) for osthole and 0.016 7-0.334 4 mg/ml(r=0.999 9) for columbianadin;RSDs of precision,stability and reproducibility tests were lower than 2.0%;recoveries were 98.46%-106.11%(RSD=2.34%,n=9)and 98.24%-102.69%(RSD=1.29%,n=9), respectively. CONCLUSIONS:The method is simple and good reproducibility,and and can be used for the contents determination of osthole and columbianadin in A. pubescens. The contents of osthole and columbianadin in A. pubescens of Wushan county in Chongqing are obviously higher than Wuxi county and other domestic areas in China.

Separation and purification of coumarins from Heracleum millefolium by high-speed counter-current chromatography / 中草药

Objective: To separate and purify coumarins from Heracleum millefolium by high-speed counter-current chromatography (HSCCC). Methods: HPLC was used to analyze and optimize the solvent system and hexane-ethyl acetate-water-methanol (1.5:2.5:2:1.5) was chosen as the two-phase solvent system of HSCCC, in which the upper phase was used as the stationary phase, while the lower phase was used as the mobile phase with flow rate of 3 mL/min, the revolution speed was set at 850 r/min, and detected at 323 nm. Three compounds were obtained as crystals by vacuum concentration at 50℃, and determined through HPLC, then their structures were identified by IR, ESI-MS, 1H-NMR, and 13C-NMR. Results: Columbianetin acetate (6.3 mg), osthole (10.6 mg), and columbianadin (6.4 mg) were obtained from crude sample (300 mg) extracts and their purity were above 96.0%. Conclusion: HSCCC is a powerful technique for the rapid separation and purification of coumarins from crude extract of H. millefolium.

Determination of Osthole and Columbianadin in Wangbi Tablets by HPLC / 中国药师

Objective:To establish an HPLC method for the determination of osthole and columbianadin in Wangbi tablets. Meth-ods:The column was an Agilent XDB C18(150 mm ×4.6 mm, 5 μm). The mobile phase consisted of acetonitrile-water(49∶51)with gradient elution. The flow rate was 1. 0 ml·min-1 and the detection wavelength was at 330 nm. Results: The calibration curve was linear within the range of 0. 060-0. 597 μg(r=0. 999 9) for osthole and 0. 010-0. 102 μg(r=0. 999 9) for columbianadin. The aver-age recovery was 99. 12% and 98. 98%, respectively. Conclusion:The method is simple, practicable, accurate and rapid. It can be applied in the content determination of osthole and columbianadin in Wangbi tablets.