mERα (IC50 = 2.3 nM); hERα (IC50 =28 nM); HBx

苯甲酸雌二醇是天然雌激素雌二醇的合成酯，更准确地说是其 3-苯甲酰酯。胚胎发生和青春期期间女性基因型的发育受到雌激素的调节。绝经前卵巢分泌的主要雌激素是雌二醇。为了诱导家畜发情，苯甲酸雌二醇（一种在 C-3 位具有苄酯的雌二醇类似物）经常与孕激素联合使用。该化合物的 IC50 值在 22 至 28 nM 之间，可与人类、小鼠和鸡的雌激素受体 α (ERα) 结合。与雌二醇相比，这表明结合亲和力降低了 6-10 倍。[1]

在 Ovx 女性模型中，苯甲酸雌二醇（20-100 μg/kg，皮下注射，每天一次，持续 4-5 周）可增强学习和记忆行为 [1]。苯甲酸雌二醇（皮下注射，单剂量，0.015–15000 μg/kg）

HBx在cccDNA表观遗传修饰中起着重要作用，调节乙型肝炎病毒（HBV）的生命周期以及肝细胞增殖和癌变。通过使用睡美人转座子系统，我们构建了一个四环素诱导的表达HBx的稳定细胞系SBHX21。使用基于NanoLuc的HiBiT检测系统可以快速检测到带有HiBiT标签的HBx。通过使用SBHX21细胞筛选药物库，我们鉴定出苯甲酸雌二醇是一种新型的抗HBx药物。苯甲酸雌二醇还以剂量依赖的方式显著降低了HBeAg、HBsAg、HBV pgRNA和HBV DNA的产生，表明苯甲酸雌醇可能是一种抗HBV药物。对接模型结果显示，苯甲酸雌二醇在TRP87和TRP107处与HBx结合。总的来说，我们的结果表明苯甲酸雌二醇抑制了HBx蛋白和HBV的转录和复制，这可能是一种新的抗HBV分子化合物，用于研究HBV感染的新治疗策略。[3]

细胞毒性试验[3]
通过细胞计数试剂盒-8（CCK8）测定评估苯甲酸雌二醇和17β-雌二醇的细胞毒性作用。HepG2细胞在96孔板中以每孔1×104个细胞的速度培养，每组三个重复，浓度梯度为1µM、50µM、100µM、150µM、200µM、250µM、300µM、350µM、400µM、450µM、500µM、550µM、600µM、650µM和700µM。所有治疗组的DMSO标准化为1%。24小时后，用在新鲜DMEM中稀释的10%CCK-8代替培养基。将CCK-8溶液和培养基（无细胞）加入空白对照中。记录吸光度（OD值）。使用GraphPad Prism计算细胞存活率曲线和IC50。

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蛋白质印迹[3]
为了分析HiBiT-HBx的表达，在12孔板中用Dox（1µg/mL）诱导SBHX21细胞48小时。然后，在没有Dox的情况下，用不同浓度的苯甲酸雌二醇和17β-雌二醇处理细胞24小时。我们使用0.5%DMSO DMEM作为对照组。处理后，用100µL RIPA缓冲液裂解细胞。细胞提取物在十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS-PAGE）上分离，并在硝化纤维（NC）印迹膜上印迹。使用乙型肝炎病毒X抗原的小鼠单克隆抗体分析蛋白质表达，并用Odyssey CLX系统进行可视化。

Animal/Disease Models: Ovx mouse [1]
Doses: 20, 100 or 200 μg/kg
Route of Administration: subcutaneous injection
Experimental Results: Significant It reverses Ovx-induced decrease in uterine weight. There was no significant effect on the locomotor activity of Ovx mice. Effectively reverses Ovx-induced changes in synaptic cleft enlargement and PSD thickness reduction. Replacement with EB (100 μg/kg) for 4 weeks increased the number and density of synaptic vesicles.

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There is increasing evidence that estrogen is involved in CNS activity, particularly memory. Several studies have suggested that estrogen improves memory by altering neuronal plasticity, including increased hippocampus CA1 dendritic spine density and enhanced long-term potentiation (LTP). In the present study, we investigated the effects of estrogen on the ultrastructural modifications in cerebral frontal cortex and hippocampus of female ovariectomized mice. One week after ovariectomy (Ovx), ICR female mice received daily injection of estradiol benzoate (EB, 20, 100, 200 microg/kg, s.c.) for 4-5 weeks. Spatial memory was then tested in the water maze, and the overall locomotor activity was monitored in open field. Synaptic morphologic parameters were examined using a graph analyzer. The results from open field did not show any alterations in locomotor activity following Ovx and EB replacement. Both the latency to find the platform and the distance to reach the platform were significantly reduced in Ovx mice by EB at 20 or 100 microg/kg when compared to vehicle treated Ovx mice. The results from synaptic ultrastructural measurement and analysis did not show any differences in hemispheric or hippocampal volumes, the numeric synaptic density, the length of active zones, or the curvature of synaptic interface among Sham, Ovx, and Ovx plus EB replacement mice. However, EB replacement effectively normalized the changes induced by Ovx, reducing the width of the synaptic cleft, enlarging the thickness of postsynaptic density (PSD), and increasing the number of synaptic vesicles in the presynapse in both cerebral cortex Fr1 and hippocampus CA1 areas. These results suggest that the beneficial effects of EB on improving memory behavior of Ovx female mice are associated with the changes of some subtle structural parameters of synapses, including the width of PSD and synaptic cleft rather than some basic and permanent structure in frontal cortex and hippocampus regions.[1]

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The current project was designed to determine the dose-response relationship of the prostate gland to estradiol exposure during the developmentally critical neonatal period in the rat. Male Sprague-Dawley (SD) rats were treated on Days 1, 3, and 5 of life by s.c. injections of a 7-log range of doses (0.015 microg/kg to 15.0 mg/kg) of beta-estradiol-3-benzoate (EB) in 25 microl of peanut oil (Arachis) as vehicle. In a separate block, neonatal Fisher 344 (F344) rats received 0.15, 15.0, or 1500.0 microg EB/kg. Rats were killed on Postnatal Day (PND) 35 or 90, and the prostates were microdissected, weighed, and frozen for immunohistochemistry. Preputial separation and hepatic testosterone hydroxlase activities were monitored and measured to determine the onset of puberty. On PND 35, there was an increase in prostate weights of SD rats treated with low doses of EB and a decrease in prostate weights of SD rats treated with high doses. The low-dose effect was entirely abolished by PND 90, and only high-dose suppression of organ sizes was found. The transient nature of the effect in low-dose animals suggests an advancement of puberty as the cause for increased reproductive organ weights on PND 35. F344 rats were more sensitive than SD rats to the suppressive effects of high doses of neonatal EB on PND 90. Despite this heightened responsiveness in the F344 rats, a low-dose estrogenic effect on adult prostate weights was not observed. Thus, in the rat model a sustained effect at low doses of natural estrogens is not present in the prostate glands.[2]

Metabolism / Metabolites
Exogenous estrogens are metabolized using the same mechanism as endogenous estrogens. Estrogens are partially metabolized by cytochrome P450.

Protein Binding
Estrogens circulate in the blood largely (>95%) bound to sex hormone binding globulin (SHBG) and to albumin.

[1]. Effects of estradiol benzoate on learning-memory behavior and synaptic structure in ovariectomized mice . Life sciences, 2006, 79(16): 1553-1560.

[2]. Neonatal low-and high-dose exposure to estradiol benzoate in the male rat: I. Effects on the prostate gland . Biology of reproduction, 2001, 65(5): 1496-1505.

[3]. Identification of Estradiol Benzoate as an Inhibitor of HBx Using Inducible Stably Transfected HepG2 Cells Expressing HiBiT Tagged HBx . Molecules, 2022, 27(15): 5000.

[4]. Investigation of Antiparasitic Activity of Two Marine Natural Products, Estradiol Benzoate, and Octyl Gallate, on Toxoplasma gondii In Vitro . Frontiers in Pharmacology, 2022, 13: 841941.

[5]. Effect of estradiol benzoate on reproductive organs and fertility in the male rat . European Journal of Obstetrics & Gynecology and Reproductive Biology, 1983, 15(3): 189-198.

[6]. Macromolecular prodrugs XI. Synthesis and characterization of polymer-estradiol conjugate. Int J Pharm. 2004 Nov 5;285(1-2):35-41.

[7]. Role of sex steroid hormones in bacterial-host interactions. Biomed Res Int. 2013;2013:928290.

[8]. Estradiol benzoate.

17beta-estradiol 3-benzoate is a benzoate ester resulting from the formal condensation of benzoic acid with the phenolic hydroxy group of 17beta-estradiol. It has a role as a xenoestrogen and an estrogen receptor agonist. It is a benzoate ester and a 17beta-hydroxy steroid. It is functionally related to a 17beta-estradiol.
Estradiol Benzoate is a pro-drug ester of [DB00783], a naturally occurring hormone that circulates endogenously within the human body. Estradiol is the most potent form of all mammalian estrogenic steroids and acts as the major female sex hormone. As a pro-drug of estradiol, estradiol benzoate therefore has the same downstream effects within the body through binding to the Estrogen Receptor (ER) including ERα and ERβ subtypes, which are located in various tissues including in the breasts, uterus, ovaries, skin, prostate, bone, fat, and brain. [DB00783] is commonly produced with an ester side-chain as endogenous estradiol has very low oral bioavailability on its own (2-10%). First-pass metabolism by the gut and the liver quickly degrades the estradiol molecule before it gets a chance to enter systemic circulation and exert its estrogenic effects. Esterification of estradiol aims to improves absorption and bioavailability after oral administration (such as with Estradiol valerate) or to sustain release from depot intramuscular injections (such as with Estradiol Cypionate) through improved lipophilicity. Following absorption, the esters are cleaved, resulting in the release of endogenous estradiol, or 17β-estradiol. Ester pro-drugs of estradiol are therefore considered to be bioidentical forms of estrogen. Estradiol benzoate is not currently available in Canada or the US.
Estradiol Benzoate is the synthetic benzoate ester of estradiol, a steroid sex hormone vital to the maintenance of fertility and secondary sexual characteristics in females. As the primary, most potent estrogen hormone produced by the ovaries, estradiol binds to and activates specific nuclear receptors. This agent exhibits mild anabolic and metabolic properties, and increases blood coagulability. (NCI04)
See also: Estradiol (has active moiety); Estradiol Benzoate; Progesterone (component of); Estradiol Benzoate; Testosterone Propionate (component of) ... View More ...

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Drug Indication
Estradiol benzoate is not currently available in any FDA or Health Canada approved products.

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Mechanism of Action
Estradiol enters target cells freely (e.g., female organs, breasts, hypothalamus, pituitary) and interacts with a target cell receptor. When the estrogen receptor has bound its ligand it can enter the nucleus of the target cell, and regulate gene transcription which leads to formation of messenger RNA. The mRNA interacts with ribosomes to produce specific proteins that express the effect of estradiol upon the target cell. Estrogens increase the hepatic synthesis of sex hormone binding globulin (SHBG), thyroid-binding globulin (TBG), and other serum proteins and suppress follicle-stimulating hormone (FSH) from the anterior pituitary.

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Pharmacodynamics
Estradiol, the principal intracellular human estrogen, is substantially more active than its metabolites, estrone and estriol, at the cellular level.

C25H28O3

376.49

376.203

C, 79.76; H, 7.50; O, 12.75

50-50-0

Alpha-Estradiol;57-91-0;Estradiol (Standard);50-28-2;Estradiol-d3;79037-37-9;Estradiol-d4;66789-03-5;Estradiol-d5;221093-45-4;Estradiol-13C2;82938-05-4;Estradiol (cypionate);313-06-4;Estradiol benzoate;50-50-0;Estradiol enanthate;4956-37-0;Estradiol hemihydrate;35380-71-3;Estradiol-d2;53866-33-4;Estradiol-13C6;Estradiol-d2-1;3188-46-3;rel-Estradiol-13C6; 979-32-8 (valerate); 113-38-2 (dipropionate); 57-63-6 (ethinyl); 172377-52-5 (sulfamate); 3571-53-7 (undecylate)

222757

White to off-white solid powder

1.2±0.1 g/cm3

531.2±50.0 °C at 760 mmHg

191-198 °C(lit.)

212.0±22.9 °C

0.0±1.5 mmHg at 25°C

1.604

6.24

46.53

1

3

3

28

582

5

O([H])[C@@]1([H])C([H])([H])C([H])([H])[C@@]2([H])[C@]3([H])C([H])([H])C([H])([H])C4C([H])=C(C([H])=C([H])C=4[C@@]3([H])C([H])([H])C([H])([H])[C@@]21C([H])([H])[H])OC(C1C([H])=C([H])C([H])=C([H])C=1[H])=O

UYIFTLBWAOGQBI-BZDYCCQFSA-N

InChI=1S/C25H28O3/c1-25-14-13-20-19-10-8-18(28-24(27)16-5-3-2-4-6-16)15-17(19)7-9-21(20)22(25)11-12-23(25)26/h2-6,8,10,15,20-23,26H,7,9,11-14H2,1H3/t20-,21-,22+,23+,25+/m1/s1

[(8R,9S,13S,14S,17S)-17-hydroxy-13-methyl-6,7,8,9,11,12,14,15,16,17-decahydrocyclopenta[a]phenanthren-3-yl] benzoate

Estradiol monobenzoate; Benzo-Gynoestryl; Ovasterol-B; Oestradiol benzoate; Oestradiol benzoate; Benovocylin; Estradiol 3-benzoate; Estradiol monobenzoate; Benzo-Gynoestryl; Estradiol Benzoate

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

DMSO: 19~100 mg/mL (265.61~50.5 mM)
H2O: ~0.1 mg/mL (~0.3 mM)

配方 1 中的溶解度: ≥ 2.5 mg/mL (6.64 mM) (饱和度未知) in 10% DMSO + 90% Corn Oil (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 25.0 mg/mL 澄清 DMSO 储备液加入900 μL 玉米油中，混合均匀。

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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));
2、取适量母液，按从左到右的顺序依次添加助溶剂，澄清后再加入下一助溶剂。以 下列配方为例说明 (注意此配方只用于说明，并不一定代表此产品 的实际溶解配方):
10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
3、溶剂前显示的百分比是指该溶剂在最终溶液/工作液中的体积所占比例;
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6、如不确定怎么将母液配置成体内动物实验的工作液，请查看说明书或联系我们;
7、 以上所有助溶剂都可在 Invivochem.cn网站购买。