Uterine Estrogen Receptor (ER): Mestranol binds to rat uterine ER with a Ki value of 0.8 nM, showing lower affinity than ethinyl estradiol (Ki=0.3 nM) [1]
- Hepatic Microsomal Mixed-Function Oxidase: Mestranol inhibits this enzyme (involved in steroid metabolism) with an IC50 of 5 μM in rat liver microsomes [1]

Mestranol, a synthetic estrogen with modest potency, has demonstrated much greater stability in hepatoma cell culture as compared to 17β-Estradiol [3]. Over a period of six days, mestranol (10 μM) can boost the proliferation of ERpositive MCF-7 WS8 cells up to 250% over control levels. Tamoxifen can partially reverse this growth stimulation. Mestranol (10 μM; 6 days) on Hep G2 hepatoma cells, however, reduces the development of Hep 3B cells by 40% in comparison to control cells. It is possible to stop cell proliferation using mestranol alone or in combination with tamoxifen. Additionally, tamoxifen and cotreatment have a cumulative effect on growth inhibition[2].

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1. ER Binding Activity ([1]):
Incubation of rat uterine ER preparations with Mestranol (0.1–10 nM) for 2 hours at 4°C competed with [³H]-estradiol for ER binding in a concentration-dependent manner. At 0.8 nM, Mestranol displaced 50% of bound [³H]-estradiol (Ki=0.8 nM). This binding was specific: no displacement was observed with progesterone or testosterone (100 nM) [1]
2. Inhibition of Hepatic Microsomal Enzyme ([1]):
Treatment of rat liver microsomes with Mestranol (1–20 μM) for 30 minutes inhibited mixed-function oxidase activity (measured via ethoxycoumarin O-deethylation). The IC50 was 5 μM: 5 μM Mestranol reduced enzyme activity by 50%, while 20 μM reduced it by 80% [1]

Mestranol (0.2 mg/kg) results in an increased percentage of liver occupied by AHF expressing glutathione S-transferase (PGST) in rats. Mestranol administration increases the focal hepatocyte labeling index compared with the control with a trend at the lower dose and a significant difference at the higher dose. Mestranol (0.02 mg/kg and 0.2 mg/kg diet) results in a decrease in the non-focal labeling index compared with that observed for the corresponding concentration of Mestranol alone. Mestranol significantly increases the non-focal hepatic labeling index in uninitiated rats compared with that observed in rats administered the basal diet only. Mestranol (50 mg/100 g b.wt.) has a significant reduction in the apparent number of alpha-2-adrenoceptors in the frontal cortex and nucleus tractus solitarius (NTS) of rat, while apparent numbers of both alpha-1 and alpha-2-adrenoceptors are depressed in the locus coeruleus.

1. Uterine Estrogen Receptor Binding Assay ([1]):
1. ER Preparation: Uteri were excised from ovariectomized rats, homogenized in Tris-HCl buffer (pH 7.4) containing EDTA and dithiothreitol, then centrifuged (100,000×g, 60 minutes) to obtain cytosolic ER fraction.
2. Reaction System: A 200 μL system contained 50 μg cytosolic ER, 0.5 nM [³H]-estradiol, and Mestranol (0.1–10 nM).
3. Incubation & Separation: Incubated at 4°C for 2 hours; unbound [³H]-estradiol was removed by adding dextran-coated charcoal (1% charcoal, 0.1% dextran) and centrifuging (3000×g, 10 minutes).
4. Detection & Calculation: Radioactivity of the supernatant was measured via liquid scintillation counter; Ki value was calculated using the Cheng-Prusoff equation [1]
2. Hepatic Microsomal Oxidase Assay ([1]):
1. Microsome Preparation: Livers were excised from male rats, homogenized in sucrose buffer, and centrifuged (9000×g, 20 minutes) followed by 100,000×g (60 minutes) to isolate microsomes.
2. Reaction System: A 500 μL system contained 100 μg microsomal protein, 1 mM NADPH (cofactor), 0.1 mM ethoxycoumarin (substrate), and Mestranol (1–20 μM).
3. Incubation & Termination: Incubated at 37°C for 30 minutes; reaction stopped by adding 1 mL ice-cold trichloroacetic acid.
4. Detection & Calculation: Fluorescence of the product (7-hydroxycoumarin) was measured (excitation 360 nm, emission 460 nm); IC50 was derived from dose-response curves [1]

Absorption, Distribution and Excretion
Ethinylestradiol binds weakly to estrogen receptors. Its estrogenic effect is due to its rapid demethylation in the liver to form ethinylestradiol; however, demethylation is incomplete, thus requiring higher doses than ethinylestradiol to achieve a similar effect. Metabolites are excreted in urine at a rate of 10-27%; ethinylestradiol metabolites are excreted at a rate of 36-54%. When ethinylestradiol molecules are labeled with tritium at position 2 or 4, or with 14C, 14-45% of the radioactive material is released into body fluids. Metabolism/Metabolites Ethinylestradiol is rapidly absorbed and extensively metabolized to ethinylestradiol. Ethinylestradiol is rapidly and fully absorbed by the gastrointestinal tract, but undergoes first-pass metabolism in the intestinal wall. Compared to many other estrogens, its metabolism in the liver is slower. Ethinylestradiol is primarily excreted by the kidneys, with some also appearing in feces. In vivo, it is rapidly demethylated in the liver to ethinylestradiol, its active form. /Estrogen/
The 3-methyl ether of ethinylestradiol—mesethrin—is more lipophilic than ethinylestradiol and has a higher affinity for adipose tissue, as demonstrated in rat studies. Memesethrin itself does not bind significantly to estrogen receptors at their antifertility sites; its hormonal activity depends on conversion to ethinylestradiol. In mice, rabbits, and humans, approximately 35% of ethinylestradiol doses are converted to ethinylestradiol, compared to 61% in rats, 56% in rabbits, and 54% in humans. The demethylated portion is then metabolized according to species-specific ethinylestradiol metabolic pathways; for example, 2-hydroxylation occurs in rats, and D-homocyclocyclization occurs in rabbits and guinea pigs. Ethinylestradiol is also demethylated to ethinylestradiol in non-human primates.
The metabolism of ethinylestradiol in humans is closely related to the metabolism of ethinylestradiol itself. Ethinyl estradiol is converted to ethinyl estradiol via demethylation: After intravenous injection of 14C-ethinyl estradiol into human volunteers, approximately 50% of the dose is demethylated to ethinyl estradiol. The main compound found in plasma is ethinyl estradiol-3-sulfate. For more complete metabolite/metabolite data on mesralol (6 metabolites in total), please visit the HSDB record page. Known human metabolites of mesralol include ethinyl estradiol.

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1. Oral absorption: The oral bioavailability of meslaol in humans is approximately 40% (25% for ethinylestradiol); after oral administration of 50 μg, the peak plasma concentration (Cmax) of 80 pg/mL is reached 2 hours [2]
2. Metabolism: Meslaol is mainly metabolized in the liver to ethinylestradiol (the active metabolite) through demethylation; approximately 70% of the administered dose is converted to ethinylestradiol within 4 hours [2]
3. Plasma half-life: The elimination half-life of meslaol in humans is 12 hours (longer than the 6 hours of ethinylestradiol) [2]
4. Distribution and excretion: Meslaol is highly lipophilic and is mainly distributed in adipose tissue; approximately 60% of the metabolites are excreted in urine (in the form of glucuronide conjugates), and 40% are excreted in feces [2]
5. Plasma protein binding rate: In human plasma, more than 95% of estradiol is bound to albumin and sex hormone-binding globulin (SHBG) [2]

[1]. Affinity of ethynyl-estradiol and mestranol for the uterine estrogen receptor and for the microsomal mixed function oxidase of the liver. J Steroid Biochem. 1973 Mar;4(2):121-8.

[2]. Pharmacokinetics of ethinyl estradiol and mestranol. Am J Obstet Gynecol. 1990 Dec;163(6 Pt 2):2114-9.

[3]. Tamoxifen inhibits hepatoma cell growth through an estrogen receptor independent mechanism. J Hepatol. 1995 Dec;23(6):712-9.

According to an independent committee of scientific and health experts, ethinylestradiol may be carcinogenic. Ethinylestradiol is a terminal alkyne compound with the structure (17α)-17-ethynylestradiol-1(10),2,4-triene, substituted with a methoxy group at the 3-position and a hydroxyl group at the 17-position. It is a prodrug and isoestrone. It is a 17β-hydroxy steroid, a terminal alkyne compound, and an aromatic ether. Its function is related to 17β-estradiol. It is the 3-methyl ether of ethinylestradiol. It must be demethylated to be biologically active. It is used as the estrogen component in many combined oral contraceptives. Ethinylestradiol is an estrogen. The mechanism of action of ethinylestradiol is as an estrogen receptor agonist. Ethinylestradiol has been reported to exist in Cunninghamella elegans, and there is relevant data. Ethinylestradiol is a semi-synthetic estrogen. It is metabolized in the liver to ethinylestradiol, the estrogen component in many combined oral contraceptives. (NCI04)
Ethinylestradiol is a 3-methyl ether. It must be demethylated to be biologically active. It is used as the estrogen component in many combined oral contraceptives.
Ethinylestradiol is a 3-methyl ether. It must be demethylated to be biologically active. It is used as the estrogen component in many combined oral contraceptives.
Drug Indications
Methylestradiol is one of the earliest oral contraceptives used.
Mechanism of Action
Methylestradiol is the 3-methyl ether of ethinylestradiol. Ethinylestradiol is a synthetic derivative of estradiol. Ethinylestradiol has oral biological activity and is the estrogen used in almost all modern combined oral contraceptive formulations. It binds to and activates the estrogen receptor. Methylestradiol is a biologically active prodrug of ethinylestradiol, which is demethylated to ethinylestradiol in the liver with a conversion efficiency of 70%. The estrogen diffuses to target cells and interacts with protein receptors. Target cells include the female reproductive tract, mammary glands, hypothalamus, and pituitary gland. Estrogen increases the synthesis of sex hormone-binding globulin (SHBG), thyroid-binding globulin (TBG), and other serum proteins in the liver, and inhibits the secretion of follicle-stimulating hormone (FSH) from the anterior pituitary. The combined use of estrogen and progestin suppresses the hypothalamic-pituitary system, reducing the secretion of gonadotropin-releasing hormone (GnRH). The mechanism of action of Norlini-1 is similar to other progestin/estrogen oral contraceptives, including ovulation inhibition, thickening of cervical mucus to form a barrier against sperm entry, and disreceptivity of the endometrium for embryo implantation. This activity is achieved through combined effects on one or more of the following structures: the hypothalamus, anterior pituitary, ovary, endometrium, and cervical mucus. Estrogen plays an important role in the female reproductive, skeletal, cardiovascular, and central nervous systems, primarily through the regulation of gene expression. When estrogen binds to the ligand-binding domain of the estrogen receptor, a biological response is initiated, leading to a conformational change that subsequently initiates gene transcription via a specific estrogen response element (ERE) of the target gene promoter. The activation or repression of the target gene is then mediated by two distinct transcriptional activation domains of the receptor (AF-1 and AF-2). Estrogen receptors can also mediate gene transcription using different response elements (e.g., AP-1) and other signaling pathways. Significant progress has been made in the molecular pharmacology of estrogen and its receptor in recent years, leading to the development of selective estrogen receptor modulators (e.g., clomiphene, raloxifene, tamoxifen, and toremifene). These drugs can bind to and activate estrogen receptors, but their action is tissue-specific and distinct from the mechanism of action of estrogen. The tissue-specific estrogen agonist or antagonist activity of these drugs appears to be related to structural differences in their estrogen receptor complexes (e.g., the AF-2 surface morphology of raloxifene differs from that of the estrogen (estradiol)-estrogens receptor complex). Furthermore, researchers have discovered a second estrogen receptor, and the presence of at least two estrogen receptors (ER-α and ER-β) may help explain the tissue-specific activity of selective modulators. Although the role of estrogen receptors in bone, cardiovascular tissue, and the central nervous system is still under investigation, emerging evidence suggests that the mechanisms of action of estrogen receptors in these tissues differ from those in reproductive tissues. /Estrogen Overview/
Intracellular sol-binding proteins of estrogen have been identified in estrogen-responsive tissues, including female reproductive organs, breasts, pituitary gland, and hypothalamus. The estrogen-binding protein complex (i.e., sol-binding protein and estrogen) is distributed into the cell nucleus, where it stimulates the synthesis of DNA, RNA, and proteins. The presence of these receptor proteins is the reason why women with metastatic breast cancer experience a remission response to estrogen therapy. /Estrogen Overview/
Estrogen generally has a beneficial effect on blood cholesterol and phospholipid concentrations. Estrogen lowers low-density lipoprotein cholesterol (LDL-C) concentrations and raises high-density lipoprotein cholesterol (HDL-C) concentrations in a dose-dependent manner. The decrease in LDL-C concentration associated with estrogen therapy appears to be due to increased LDL catabolism, while the increase in triglyceride concentration is caused by increased production of large, triglyceride-rich very low-density lipoprotein (VLDL); changes in serum HDL-C concentration appear to be primarily due to increased cholesterol and apolipoprotein A-1 content in HDL2-cholesterol and a slight increase in HDL3-cholesterol. /General Estrogen Statement/
For more complete data on the mechanisms of action of MESTRANOLs (7 in total), please visit the HSDB record page.

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1. Drug Background ([1][2]):
Mestranol is a synthetic estrogen prodrug whose structure is related to ethinylestradiol (the difference being a methyl group at the 3 position). It is inactive in its parental form and needs to be demethylated in the liver to be converted into ethinylestradiol in order to exert its estrogenic effect [1][2]
2. Mechanism of action ([1][2]):
- After being converted into ethinylestradiol, it binds to the estrogen receptor (ER) (Ki=0.3 nM for ethinylestradiol), activating the transcription of estrogen-responsive genes in reproductive tissues [1]
- As a prodrug, estradiol is metabolized more slowly than ethinylestradiol and has a longer half-life, thus providing more sustained estrogenic activity [2]
3. Therapeutic use ([2]):
Ethinylestradiol has been used as a component of combined oral contraceptives (usually 50 micrograms per tablet) to inhibit ovulation through negative feedback on the hypothalamic-pituitary-gonadal axis [2]

C21H26O2

310.43

310.193

72-33-3

Mestranol-d2;Mestranol-d4

6291

White to off-white solid powder

1.2±0.1 g/cm3

442.3±45.0 °C at 760 mmHg

153-155 °C(lit.)

190.8±23.0 °C

0.0±1.1 mmHg at 25°C

1.591

5.17

1

2

2

23

519

5

C[C@]12CC[C@H]3[C@H]([C@@H]1CC[C@]2(C#C)O)CCC4=C3C=CC(=C4)OC

IMSSROKUHAOUJS-UHFFFAOYSA-N

InChI=1S/C21H26O2/c1-4-21(22)12-10-19-18-7-5-14-13-15(23-3)6-8-16(14)17(18)9-11-20(19,21)2/h1,6,8,13,17-19,22H,5,7,9-12H2,2-3H3

17-ethynyl-3-methoxy-13-methyl-7,8,9,11,12,13,14,15,16,17-decahydro-6H-cyclopenta[a]phenanthren-17-ol

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)

Solubility in Formulation 1: ≥ 2.08 mg/mL (6.70 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.08 mg/mL (6.70 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)