Columbianadin, a natural coumarin from, is known to have various biological activities including anti-inflammatory and anti-cancer effects.

Apoptosis^[1]

Columbianadin (CBN) effectively suppresses the growth of colon cancer cells. Low concentration (up to 25 μM) of Columbianadin induces apoptosis, and high concentration (50 μM) of Columbianadin induces necroptosis. The induction of apoptosis by Columbianadin is correlated with the modulation of caspase-9, caspase-3, Bax, Bcl-2, Bim and Bid, and the induction of necroptosis is related with RIP-3, and caspase-8. In addition, Columbianadin induces the accumulation of ROS and imbalance in the intracellular antioxidant enzymes such as SOD-1, SOD-2, catalase and GPx-1. Columbianadin shows the most effective growth inhibitory activity against human colorectal cancer cells. Accordingly, further study is performed using HCT116 cells to give the detailed growth-inhibitory mechanism of action mediated by Columbianadin. The cells treated with various concentrations of Columbianadin (0-100 μM) exhibit a dose- and time-dependent growth inhibition with an IC₅₀ value of 47.2 and 32.4 μM after 48 and 72 h incubation, respectively. Treatment of various concentrations (12.5, 25, and 50 μM) of Columbianadin for 48 h in HCT116 cells decreases the number of cells and increases the floating cells. Apparent morphological changes with round-shape and dying cells are also observed at 25 and 50 μM Columbianadin -treated cells^[1].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

The analysis method is successfully applied to a tissue distribution study of Columbianadin (CBN) and Columbianetin (CBT) after intravenous administration of Columbianadin to rats. The results of this study indicated that Columbianadin can be detected in all of the selected tissues after i.v. administration. Columbianadin is distributed to rat tissues rapidly and can be metabolized to CBT in most detected tissues. Of the detected tissues, heart had the highest uptake of Columbianadin, which suggests that heart might be one of the main target tissues of Columbianadin ^[2].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

328.36

C₁₉H₂₀O₅

5058-13-9

二氢欧山芹醇当归酸酯；二氢欧山芹当归酸酯

Room temperature in continental US; may vary elsewhere.

Powder	-20°C	3 years
	4°C	2 years
In solvent	-80°C	6 months
	-20°C	1 month

In Vitro: 

DMSO : 33.33 mg/mL (101.50 mM; Need ultrasonic)

*

请根据产品在不同溶剂中的溶解度选择合适的溶剂配制储备液；一旦配成溶液，请分装保存，避免反复冻融造成的产品失效。
储备液的保存方式和期限：-80°C, 6 months; -20°C, 1 month。-80°C 储存时，请在 6 个月内使用，-20°C 储存时，请在 1 个月内使用。

In Vivo:

请根据您的实验动物和给药方式选择适当的溶解方案。以下溶解方案都请先按照 In Vitro 方式配制澄清的储备液，再依次添加助溶剂：

——为保证实验结果的可靠性，澄清的储备液可以根据储存条件，适当保存；体内实验的工作液，建议您现用现配，当天使用； 以下溶剂前显示的百
分比是指该溶剂在您配制终溶液中的体积占比；如在配制过程中出现沉淀、析出现象，可以通过加热和/或超声的方式助溶

• 1.

  请依序添加每种溶剂： 10% DMSO    90% corn oil

  Solubility: ≥ 3.5 mg/mL (10.66 mM); Clear solution

  此方案可获得 ≥ 3.5 mg/mL (10.66 mM，饱和度未知) 的澄清溶液，此方案不适用于实验周期在半个月以上的实验。

  以 1 mL 工作液为例，取 100 μL 35.0 mg/mL 的澄清 DMSO 储备液加到 900 μL玉米油中，混合均匀。

• [1]. Kang JI, et al. Columbianadin Inhibits Cell Proliferation by Inducing Apoptosis and Necroptosis in HCT116 Colon Cancer Cells. Biomol Ther (Seoul). 2016 May 1;24(3):320-7.

  [2]. Zhang YB, et al. Tissue distribution study of columbianadin and its active metabolite columbianetin in rats. Biomed Chromatogr. 2016 Feb;30(2):256-62.

The effect of Columbianadin on the cell proliferation is evaluated by SRB cellular protein-staining method. The cells are seeded in 96-well plates with various concentrations of Columbianadin and incubated at 37°C in a humidified incubator with 5% CO₂ for 48 and 72 h. The cells are fixed with 10% trichloroacetic acid (TCA) solution for 30 min at 4°C, washed 5 times with tap water, and dried in the air. The cells are stained with 0.4% SRB in 1% acetic acid solution for 1 h at room temperature. After washing out the unbound dye and drying, the stained cells are dissolved in 10 mM Tris buffer (pH 10.0), and the absorbance is measured at 515 nm. Cell viability is calculated by comparing to the absorbance of the vehicle-treated control group. The IC₅₀ values are calculated by non-linear regression analysis using the Table Curve 2D v5.01 software^[1].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

Rats^[2]
For tissue distribution study, 20 male Sprague-Dawley rats (200±10 g) are divided into four groups (n=5 per group), and Columbianadin is i.v. administered to overnight fasted rats at a single dose of 20 mg/kg. The rats in the four groups are sacrificed by decapitation at 10, 30, 60 and 120 min. Then the tissues of liver, lung, kidney, spleen, heart, stomach, intestine, brain, testis and muscle are removed and washed. Each tissue sample is weighted and stored at -20 °C until analysis.

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

• [1]. Kang JI, et al. Columbianadin Inhibits Cell Proliferation by Inducing Apoptosis and Necroptosis in HCT116 Colon Cancer Cells. Biomol Ther (Seoul). 2016 May 1;24(3):320-7.

  [2]. Zhang YB, et al. Tissue distribution study of columbianadin and its active metabolite columbianetin in rats. Biomed Chromatogr. 2016 Feb;30(2):256-62.