Norwogonin   中文名称: 去甲汉黄芩素

- Enterovirus 71 (EV71) (IC50 = 31.83 μg/ml) [1]
- Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) [2]

去甲汉黄素（0.4、2、10 和 50 μg/mL；48 小时）抑制 EV71 在 Vero 细胞中的复制，并防止 EV71 感染引起的细胞毒性[1]。
去甲汉黄素（50 μg/mL；48 小时）显着降低病毒VP2蛋白表达[1]。
48小时，去甲汉黄素(50 μg/mL)显着降低EV71 NCR基因的表达[1]。

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抑制EV71复制：Norwogonin在0.4、2、10和50 μg/ml浓度下处理48小时，可抑制Vero细胞中EV71的复制，阻止EV71感染诱导的细胞毒性。在50 μg/ml浓度下处理48小时，能显著降低病毒VP2蛋白的表达，并强烈减少EV71非编码区（NCR）基因的表达[1]

结果：黄芩和白勺药对（HBHP）显著降低DSS诱导的UC小鼠的DAI评分，缩短结肠长度。给予HBHP能够有效地减轻粘膜溃疡和上皮破坏。此外，HBHP治疗显著降低了结肠组织中TNF-α、IL-6和IL-1β的表达（p<0.05或p<0.01）。获得35个与UC相关的生物活性化合物和290个HBHP靶点。其中，在药物-化合物靶向网络中鉴定出3个度值较高的关键活性化合物（黄芩苷、穿心莲素和norwogonin）和21个枢纽基因（STAT3、JAK2、SRC、AKT1、PIK3CA和VEGFA等）。KEGG富集分析表明，HBHP的机制主要涉及JAK-STAT途径。JAK/STAT信号的异常激活被认为与UC的发病机制有关。值得注意的是，WB和IHC显示HBHP显著下调p-JAK2（p<0.05）和p-STAT3（p<0.05或p<0.01）的蛋白表达水平。JAK2和STAT3可能是HBHP作用的核心靶点；分子对接也支持了这种可能性。[2]
结论：HBHP可减轻DSS诱导的UC，减轻组织炎症，其机制可能主要通过抑制JAK2/STAT3信号通路来实现。同时，我们的工作表明，网络药理学与实验验证相结合是研究中药作用机制的有力手段。[2]

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缓解溃疡性结肠炎：含有Norwogonin的黄芩-白芍药对可缓解葡聚糖硫酸钠（DSS）诱导的小鼠溃疡性结肠炎。它能降低疾病活动指数（DAI）评分，减轻结肠缩短程度，缓解黏膜溃疡和上皮损伤，还能降低结肠组织中肿瘤坏死因子-α（TNF-α）、白细胞介素-6（IL-6）和白细胞介素-1β（IL-1β）的表达。其作用机制可能与抑制JAK2/STAT3信号通路有关，可显著下调p-JAK2和p-STAT3的蛋白表达水平[2]

在本研究中，使用细胞病变效应（CPE）还原法评估了黄芩中的去骨原素、oroxylinA和mosloflavone的抗EV71活性，结果表明这三种化合物都具有较强的抗EV七十一活性，并减少了可见CPE的形成。在病毒感染的初始阶段，Norwogonin、oroxylinA和mosloflavone也抑制了病毒的复制，它们抑制了病毒VP2蛋白的表达，从而抑制了病毒衣壳蛋白的合成。然而，与其他药物相比，利巴韦林的疗效相对较弱。因此，这些发现提供了重要的信息，将有助于利用norwogonin、oroxylinA和mosloflavone治疗EV71[1]。

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抗病毒活性测定[1]
抗病毒活性的测定是用SRB方法进行的，该方法评估细胞病变效应（CPE）的减少，这是Choi等人先前报道的。（2009a）。观察了norwogonin、oroxylina和mosloflavone对EV71诱导的CPE的影响。简言之，将Vero细胞以每孔2×104个细胞的浓度接种到96孔培养板上。第二天，取出培养基并用PBS洗涤，加入0.09mL含有TCID50（50%组织培养感染剂量）病毒原液的EV71稀释病毒悬浮液，以在感染后2天内产生适当的细胞病变作用，并加入0.01mL去甲精素、oroxylinA、mosloflavone和利巴韦林（分别为0.4、2、10和50μg/mL）。在37°C的5%CO2中孵育2天后，在32×10放大倍数的显微镜下观察细胞的形态，并记录图像。

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添加测定时间[1]
根据先前描述的程序（Chiang et al.，2003），对norwogonin、oroxylinA和mosloflavone的添加时间效应进行了轻微修改。将Vero细胞以每孔2×104个细胞的密度接种到96孔培养板上，并孵育1天。用磷酸盐缓冲盐水（PBS）洗涤后，在EV71感染前（−1小时）、感染期间（0小时）或感染后（1、2、4、6和8小时），向细胞中加入50μg/mL的去甲精、oroxylinA和mosloflavone。2天后，使用SRB测定法检测抗病毒活性，并使用利巴韦林作为阳性对照。

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- Vero细胞实验：将Vero细胞以每孔1×10^4个细胞的密度接种于96孔板，孵育过夜。随后以0.1的感染复数（MOI）用EV71感染细胞，同时用不同浓度的Norwogonin（0.4、2、10和50 μg/ml）处理。孵育48小时后，通过MTT法检测细胞活力以评估对EV71复制的抑制作用；采用Western blot检测病毒VP2蛋白的表达，实时荧光定量PCR检测EV71 NCR基因的表达水平[1]
- 动物实验方案 - 溃疡性结肠炎小鼠实验：通过让小鼠饮用3% DSS溶液7天诱导溃疡性结肠炎。对小鼠给予含有Norwogonin的黄芩-白芍药对，文献中未提及具体给药途径和频率。之后，根据体重减轻、腹泻和便血情况每日计算DAI评分。实验结束后，测量结肠长度，收集结肠组织进行组织学检查和细胞因子表达水平检测，以及通过Western blot和免疫组织化学检测p-JAK2和p-STAT3蛋白的表达[2]

First, The UC model of mice induced by dextran sulfate sodium (DSS) was established. The mice were randomly divided into Control group, DSS group, SASP group (390 mg/kg), and HPHP group (1.95 g/kg), with 8 mice per group. Drugs were administrated via oral gavage for 7 days. Then, Disease activity index (DAI), length of the colon, histopathology, and changes in inflammatory cytokines in colonic tissues were analyzed to assess the effect of HBHP on UC. Besides, Network pharmacology was applied to identify the active compounds, core targets of HBHP in the treatment of UC, and the corresponding signaling pathways to explore the underlying mechanisms. Finally, Western blot (WB), immunohistochemistry (IHC) and molecular docking were performed to validate the results.[2]

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UC was induced in mice by administration of 3.5 % (w/v) DSS (MW 36–50 kDa) in drinking water for 7 days ad libitum as described previously. Mice were randomly divided into four groups (n = 8 each): Control group (normal drinking water + normal water orally once daily), DSS group (DSS drinking water + normal water orally once daily), SASP group (DSS drinking water + 390 mg/kg SASP orally once daily), and HPHP group (DSS drinking water + 1.95 g/kg HBHP orally once daily). The dosage of HBHP used here was calculated in proportion to human clinical doses (15 g/day per adult) using body surface area conversion.[2]
DSS solution was updated every day. The oral gavage administration was initiated simultaneously with DSS drinking water. Body weights, stool properties, and stool occult blood were monitored every day. At the end of the experiment, all mice were sacrificed and colonic specimens were collected. The lengths of the colons were measured and then immediately stored at −80 °C or fixed in 10 % paraformaldehyde for further procedures.[2]

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- Mouse experiment for ulcerative colitis: Mice are induced to have ulcerative colitis by drinking a 3% DSS solution for 7 days. The Huangqin-Baishao herb pair containing Norwogonin is administered to the mice, and the specific administration route and frequency are not mentioned in the literature. Then, the DAI score is calculated daily according to the criteria of weight loss, diarrhea, and hematochezia. After the experiment, the colon length is measured, and the colon tissues are collected for histological examination and detection of cytokine expression levels, as well as the detection of p-JAK2 and p-STAT3 protein expression by Western blot and immunohistochemistry [2]

[1]. Inhibitory Effects of Norwogonin, Oroxylin A, and Mosloflavone on Enterovirus 71. Biomol Ther (Seoul). 2016 Sep 1;24(5):552-8.

[2]. The effect and mechanism of Huangqin-Baishao herb pair in the treatment of dextran sulfate sodium-induced ulcerative colitis. Heliyon. 2023 Nov 30;9(12):e23082.

Norwogonin is a trihydroxyflavone with the hydroxy groups at positions C-5, -7 and -8. It has a role as an antioxidant and a metabolite.
Norwogonin has been reported in Scutellaria discolor, Scutellaria scandens, and other organisms with data available.

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Norwogonin is a flavonoid compound isolated from Scutellaria baicalensis Georgi. It has potential in treating EV71-related diseases due to its inhibitory effect on EV71 in vitro, and it is also one of the key active compounds in the Huangqin-Baishao herb pair, which plays an important role in alleviating ulcerative colitis by inhibiting the JAK2/STAT3 signaling pathway

C15H10O5

270.2369

270.053

4443-09-8

5281674

Typically exists as yellow to orange solids at room temperature

1.548g/cm3

528.4ºC at 760 mmHg

258 ºC (ethanol )

206ºC

1.732

Scutellaria baicalensis Georgi

2.576

90.9

3

5

1

20

413

0

O1C(C2C([H])=C([H])C([H])=C([H])C=2[H])=C([H])C(C2C(=C([H])C(=C(C1=2)O[H])O[H])O[H])=O

ZFKKRRMUPBBYRS-UHFFFAOYSA-N

InChI=1S/C15H10O5/c16-9-6-11(18)14(19)15-13(9)10(17)7-12(20-15)8-4-2-1-3-5-8/h1-7,16,18-19H

5,7,8-trihydroxy-2-phenylchromen-4-one

5,7,8-Trihydroxyflavone;5,7,8-Trihydroxy-2-phenyl-4H-chromen-4-one;FLAVONE, 5,7,8-TRIHYDROXY-; NSC-128304;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

注意： 本产品在运输和储存过程中需避光。

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~250 mg/mL (~925.10 mM)

配方 1 中的溶解度: ≥ 2.08 mg/mL (7.70 mM) (饱和度未知) in 10% DMSO + 40% PEG300 +5% Tween-80 + 45% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将100 μL 20.8 mg/mL澄清DMSO储备液加入400 μL PEG300中，混匀；然后向上述溶液中加入50 μL Tween-80+，混匀；加入450 μL生理盐水定容至1 mL。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
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10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
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