Isoorientin is a potent inhibitor of COX-2 with an IC₅₀ value of 39 μM.

Isoorientin is a Selective Inhibitor of Cyclooxygenase-2 (COX-2) from the Tubers of Pueraria tuberosa^[1]. PANC-1 and PATU-8988 cells are grown for 24 hours in the presence of Isoorientin (0, 20, 40, 80, and 160 μM), and a CCK8 solution is added. The cell viability decreases significantly at the concentrations of 20, 40, 80, and 160 μM. After the cells are cultured with Isoorientin (0, 20, 40, 80, and 160 μM for PANC-1; 0, 20, 40, 80, 160, and 320 μM for PATU-8988) for 24 hours, the expression of p-AMPK and AMPK is assessed by Western blotting. After the Isoorientin treatment, the p-AMPK expression is increased. Then, in the shRNA group, the concentration of 80 μM is used to detect the effects of Isoorientin. The expression levels of AMPK and p-AMPK are much lower in the shRNA group than in the wild-type PC cells (WT) and the group that is transfected with a negative control lentivirus (NC)^[2].

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

Animals treated with Isoorientin at 10 mg/kg and 20 mg/kg body weight have a statistically significant reduction in paw edema, with a mean peak thickness of 1.19±0.05 mm and 1.08±0.04 mm, respectively. This indicated that Isoorientin significantly attenuates paw edema compared with the control group^[3].

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

448.38

C₂₁H₂₀O₁₁

4261-42-1

异荭草素；异荭草苷

Room temperature in continental US; may vary elsewhere.

DMSO : ≥ 100 mg/mL (223.03 mM)

* “≥” means soluble, but saturation unknown.

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。

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 ;; 40% PEG300 ;; 5% Tween-80 ;; 45% saline

  Solubility: ≥ 2.5 mg/mL (5.58 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (5.58 mM，饱和度未知) 的澄清溶液。

  以 1 mL 工作液为例，取 100 μL 25.0 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀；向上述体系中加入50 μL Tween-80，混合均匀；然后继续加入 450 μL生理盐水定容至 1 mL。

  将 0.9 g 氯化钠，完全溶解于 100 mL ddH₂O 中，得到澄清透明的生理盐水溶液

• 2.

  请依序添加每种溶剂：;10% DMSO ;; 90% (20% SBE-β-CD in saline)

  Solubility: ≥ 2.5 mg/mL (5.58 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (5.58 mM，饱和度未知) 的澄清溶液。

  以 1 mL 工作液为例，取 100 μL 25.0 mg/mL 的澄清 DMSO 储备液加到 900 μL 20% 的 SBE-β-CD 生理盐水水溶液中，混合均匀。

  将 2 g 磺丁基醚 β-环糊精加入 5 mL 生理盐水中，再用生理盐水定容至 10 mL，完全溶解，澄清透明

• [1]. Sumalatha M, et al. Isoorientin, a Selective Inhibitor of Cyclooxygenase-2 (COX-2) from the Tubers of Pueraria tuberosa. Nat Prod Commun. 2015 Oct;10(10):1703-4.

  [2]. Ye T, et al. Isoorientin induces apoptosis, decreases invasiveness, and downregulates VEGF secretion by activating AMPK signaling in pancreatic cancer cells. Onco Targets Ther. 2016 Dec 12;9:7481-7492.

  [3]. Anilkumar K, et al. Evaluation of Anti-Inflammatory Properties of Isoorientin Isolated from Tubers of Pueraria tuberosa. Oxid Med Cell Longev. 2017;2017:5498054.

PANC-1 and PATU-8988 cells are plated onto 96-well plates. Each well contain ~5,000 cells and 200 μL of the medium with 10% FBS. When the cells of each well reach 70% confluency, the medium is changed, and FBS-free medium with different concentrations of Isoorientin is added. After 24 hours, the cells are washed with PBS once, the medium containing Isoorientin is discarded, and 100 μL of FBS-free medium with 10 μL of the Cell Counting Kit 8 (CCK8) reagent are added. The cells are incubated for another 1-2 hours at 37°C, and the absorbance of each well is detected using an ELISA reader at 490 nm. Cell viability is expressed as the fold change of absorbance^[2].

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

Mice^[3]
In the case of paw edema model the Isoorientin or Celecoxib is given intraperitoneally and Carrageenan is injected into the paw directly one hour later. In air pouch model all the treatments are given along with Carrageenan directly into the pouch cavity. Isoorientin is injected three hours earlier than injection of Carrageenan into the pouch cavity. Isoorientin and Celecoxib are administered into the mice air pouch. The stock solutions of Isoorientin (100 mg/mL) and Celecoxib (100 mg/mL) are prepared in DMSO and further dilutions are made at the time of treatments. Animals are divided, into 5 different groups as follows: control (DMSO treated); Carrageenan (0.5 mL of 1.5% (w/v) Carrageenan in saline) treated; Carrageenan+Celecoxib (20 mg/kg body weight) treated; Carrageenan+Isoorientin (10 mg/Kg body weight) treated; Carrageenan+Isoorientin (20 mg/Kg body weight) treated.

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

• [1]. Sumalatha M, et al. Isoorientin, a Selective Inhibitor of Cyclooxygenase-2 (COX-2) from the Tubers of Pueraria tuberosa. Nat Prod Commun. 2015 Oct;10(10):1703-4.

  [2]. Ye T, et al. Isoorientin induces apoptosis, decreases invasiveness, and downregulates VEGF secretion by activating AMPK signaling in pancreatic cancer cells. Onco Targets Ther. 2016 Dec 12;9:7481-7492.

  [3]. Anilkumar K, et al. Evaluation of Anti-Inflammatory Properties of Isoorientin Isolated from Tubers of Pueraria tuberosa. Oxid Med Cell Longev. 2017;2017:5498054.