ER/Estrogen receptor. Estradiol enanthate is a long-acting estrogen ester used in combination with dihydroxyprogesterone acetophenide (DHPA) as a once-monthly injectable contraceptive. No specific binding affinity or potency data (e.g., IC₅₀, EC₅₀) were reported in this study. [1]
Estradiol enanthate targets the estrogen receptor (ER), specifically ERα and ERβ, as a prodrug of 17β-estradiol. Upon administration, the enanthate ester is cleaved to release the active hormone 17β-estradiol, which binds to and activates estrogen receptors. This activation modulates the transcription of estrogen-responsive genes involved in reproductive function, bone metabolism, cardiovascular health, and various other physiological processes. The compound also acts as a hormonal contraceptive when combined with a progestogen. Its primary mechanism of action is through estrogen receptor-mediated signaling pathways.

In vitro, estradiol enanthate itself is relatively inactive as it requires enzymatic cleavage to release the active 17β-estradiol. The released estradiol binds to estrogen receptors with high affinity and activates estrogen-dependent gene transcription in ER-positive cell lines. Estradiol stimulates cell proliferation in ER-positive breast cancer cell lines such as MCF-7. It also modulates the expression of various estrogen-responsive genes involved in cell cycle regulation, apoptosis, and differentiation. The compound's in vitro activity is typically assessed using reporter gene assays, proliferation assays, and gene expression analysis in appropriate cell culture models.

Contraceptive Efficacy: In a prospective, open, randomized study of 365 adolescents (aged 14–19 years) using a monthly injectable contraceptive containing estradiol enanthate 10 mg and dihydroxyprogesterone acetophenide 150 mg, a total of 5 pregnancies occurred over 12 cycles of use: 2 in Group 1 (injection every 30 ± 3 days) and 3 in Group 2 (injection on days 7–10 of the menstrual cycle). The difference was not statistically significant (p = 0.48). Life table analysis showed cumulative pregnancy rates of 1.4% and 1.7% in Groups 1 and 2, respectively. [1]
Bleeding Pattern: The mean duration of bleeding episodes was 5.8 ± 1.7 days in Group 1 and 5.5 ± 1.2 days in Group 2 (p = 0.187). The mean cycle length was 28.7 ± 4.1 days in Group 1 and 26.6 ± 3.8 days in Group 2 (p < 0.001). Spotting episodes ranged from 2.0 ± 1.0 to 3.5 ± 6.0 days in Group 1 and 2.3 ± 1.2 to 3.6 ± 2.3 days in Group 2 over 90-day segments, with no significant differences between groups. [1]
Treatment Completion: Of 365 enrolled subjects, 266 (73%) completed the 12-month treatment: 130/186 (70%) in Group 1 and 136/179 (76%) in Group 2 (p = 0.21). Mean time in the study was 7.64 ± 0.5 months in Group 1 and 7.55 ± 0.4 months in Group 2. [1]
Introduction: This study compared two regimens of a monthly injectable contraceptive containing dihydroxyprogesterone acetophenide 150 mg and estradiol enanthate 10 mg (Perlutan) over 12 cycles of use.Methods: Three hundred sixty-five adolescents were randomized into two groups. The patients in Group 1 received an initial injection of Perlutan on the 1st-5th day of their menstrual cycle and subsequent injections every 30 +/- 3 days, whereas those in Group 2 followed the traditional schedule of administration in which the first injection is administered between Days 7 and 10 of their menstrual cycle and subsequent injections 7-10 days after Day 1 of withdrawal bleeding. This schedule may result in an irregularity in the timing of injections.Results: No significant difference was found between the two groups regarding tolerability or pregnancy (two in Group 1 and three in Group 2).Conclusion: Monthly administration limits the annual number of injections to a maximum of 12, thus frequently reducing the total annual dose while maintaining efficacy and tolerability similar to those obtained with the traditional regimen.[1]

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In vivo, estradiol enanthate functions as a long-acting prodrug of 17β-estradiol. When administered via intramuscular injection in oil, a depot effect occurs, providing sustained release of estradiol over an extended period. A single 10 mg intramuscular dose has been found to result in significantly elevated levels of estradiol for at least 31 days. The duration of effect is considerably longer than that of other commonly used estradiol esters such as estradiol valerate and estradiol cypionate. The combination with dihydroxyprogesterone acetophenide provides a once-monthly injectable contraceptive.

In vitro enzyme/receptor binding assays for estradiol enanthate typically involve hydrolysis of the ester to release 17β-estradiol, followed by competitive binding experiments using estrogen receptors (ERα and ERβ) and radiolabeled estradiol as tracer. The binding affinity of the released estradiol to estrogen receptors is assessed, with IC₅₀ or Kd values determined. Assays are conducted in buffered solutions at physiological pH with appropriate receptor preparations. The compound's prodrug nature means that enzymatic activity (esterase activity) may also be evaluated in these assays. The deuterated form serves as a standard for analytical quantification.

In vitro cell-based assays for estradiol enanthate typically involve treatment of ER-positive breast cancer cell lines such as MCF-7 with the compound, allowing cellular esterases to convert it to active estradiol. Cells are treated for 24-72 hours, followed by assessment of cell proliferation using MTT, CCK-8, or BrdU incorporation assays. Estrogen-dependent gene transcription is evaluated using reporter gene assays with ERE-luciferase constructs or by qPCR analysis of estrogen-responsive genes. Standard cell culture conditions (37°C, 5% CO₂) with charcoal-stripped serum are employed to minimize background estrogenic activity. Dose-response curves are generated to determine EC₅₀ values.

Study Design: This was a prospective, open, randomized clinical study conducted at six research centers in Brazil. A total of 365 female adolescents aged 14–19 years were enrolled and randomized into two groups. [1]

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Dosing Regimen: Estradiol enanthate (10 mg) was administered intramuscularly as part of a combination injectable contraceptive (Perlutan®) containing dihydroxyprogesterone acetophenide 150 mg. Group 1 received the first injection between days 1–5 of the menstrual cycle, with subsequent injections every 30 ± 3 days. Group 2 received the first injection between days 7–10 of the menstrual cycle, with subsequent injections scheduled 7–10 days after the first day of withdrawal bleeding. All injections were given deeply intramuscularly. [1]
Duration: The study duration was 12 cycles (approximately 12 months) of contraceptive use. [1]
Exclusion Criteria: Patients were excluded if they had used any hormonal contraception in the preceding 3–8 months, had known neoplastic disease, serious hepatopathy, kidney insufficiency, hypersensitivity to any component, were pregnant or lactating, amenorrheic, or had contraindications to hormonal contraception. [1]
Monthly injectable steroid contraceptives which contained the long-acting progestogen dihydroxyprogesterone acetofenide plus a shorter-acting estrogen (usually estradiol enanthate) were used by women in two of the countries (Chile and Mexico) from which data were collected. In preliminary analyses of data from Chile (1979-1983), a strong association was observed between use of these products and invasive cervical cancer. Therefore, three additional data sets from these two countries were analyzed in further detail for this report. Analyses of additional data from Chile on invasive cervical cancer (1983-1985) and cervical carcinoma in situ (1979-1986) and of data on invasive cervical cancer from Mexico (1979-1986) failed to confirm the initially observed association. The original finding was probably due to chance, but a causal interpretation cannot be confidently ruled out, and additional studies are warranted.[2]

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In vivo animal studies with estradiol enanthate typically involve intramuscular administration of the compound in oil to rodent models to evaluate its estrogenic effects and pharmacokinetic profile. Studies may investigate the compound's duration of action, with blood samples collected at various time points to measure estradiol levels. Efficacy studies may evaluate the compound's effects on reproductive function, bone density, or other estrogen-sensitive endpoints. The combination with progestogens may be studied in contraceptive efficacy models. All procedures must comply with institutional animal care and use guidelines.

Estradiol enanthate has a very long duration of effect due to its depot formulation. A single 10 mg intramuscular dose results in significantly elevated estradiol levels for at least 31 days. The duration is considerably longer than that of estradiol valerate and estradiol cypionate. The compound has a molecular weight of 384.55 g/mol. It is soluble in chloroform and methanol but has limited water solubility. The compound is typically stored under refrigerated conditions. When administered in oil via intramuscular injection, the ester is gradually released from the depot and hydrolyzed to active estradiol.

Adverse Events: The incidence of nonserious adverse events possibly related to the study drug was 26.9% in Group 1 and 20.7% in Group 2 (p = 0.22). Four serious adverse events were reported (influenza-like symptoms, pneumonia, cholecystectomy, and acute diarrhea), but none were considered by investigators to be related to the study medication. [1]
Weight Change: Mean body weight increased by 1.1 kg in Group 1 and 0.9 kg in Group 2 over the study duration (p = 0.931). The weight change was not significantly different between groups. [1]
Blood Pressure and Pulse Rate: No statistically significant changes in systolic blood pressure, diastolic blood pressure, or pulse rate were observed in either group during the study. [1]
Dysmenorrhea: There was no difference between the two groups regarding the occurrence of moderate to severe dysmenorrhea, reported by 24/175 patients in Group 1 and 23/175 in Group 2. [1]
Discontinuation Reasons: Discontinuation due to bleeding problems occurred in 0.7% of Group 1 and 2.0% of Group 2 (p not significant). Discontinuation due to personal reasons was 7.1% in Group 1 and 1.3% in Group 2 (p < 0.05). [1]

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Estradiol enanthate has been used clinically as a component of hormonal contraception since the mid-1960s. As an approved pharmaceutical ingredient, it has undergone comprehensive toxicological evaluation. The compound is not recommended for use during pregnancy. Common adverse effects are consistent with those expected from estrogen therapy and may include nausea, breast tenderness, headache, and increased risk of thromboembolic events. Patients should be monitored for potential side effects. The compound should not be used in individuals with known hypersensitivity to estrogens or with certain medical conditions such as history of thromboembolic disorders or estrogen-dependent cancers.

[1]. Comparison of two regimens of a monthly injectable contraceptive containing dihydroxyprogesterone acetophenide and estradiol enanthate. Contraception. 2006;73(3):249-252.

[2]. Monthly injectable steroid contraceptives and cervical carcinoma. Am J Epidemiol. 1989;130(2):237-247.

Estradiol heptaate is a steroidal ester.

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Estradiol enanthate (CAS#: 4956-37-0) has a molecular formula of C₂₅H₃₆O₃ and a molecular weight of 384.55 g/mol. Its IUPAC name is [(8R,9S,13S,14S,17S)-3-hydroxy-13-methyl-6,7,8,9,11,12,14,15,16,17-decahydrocyclopenta[a]phenanthren-17-yl] heptanoate. It is a synthetic, steroidal estrogen ester and a long-acting prodrug of 17β-estradiol. It is used with dihydroxyprogesterone acetophenide as a once-monthly injectable contraceptive. This combination has been marketed since the mid-1960s. Estradiol enanthate does not appear to be available as a standalone drug.

C25H36O3

384.55

384.266

4956-37-0

Estradiol;50-28-2;Estradiol (cypionate);313-06-4;Estradiol benzoate;50-50-0

21070

White to off-white solid powder

1.1±0.1 g/cm3

509.5±50.0 °C at 760 mmHg

94-96ºC

197.1±22.9 °C

0.0±1.4 mmHg at 25°C

1.560

7.68

1

3

7

28

546

5

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

RFWTZQAOOLFXAY-BZDYCCQFSA-N

InChI=1S/C25H36O3/c1-3-4-5-6-7-24(27)28-23-13-12-22-21-10-8-17-16-18(26)9-11-19(17)20(21)14-15-25(22,23)2/h9,11,16,20-23,26H,3-8,10,12-15H2,1-2H3/t20-,21-,22+,23+,25+/m1/s1

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

ESTRADIOL ENANTHATE; 4956-37-0; Oestradiol 17-heptanoate; Estradiol Enantate; Estradiol 17-heptanoate; Estradiol enanthate [USAN]; SQ 16,150; PAP315WZIA;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light.

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

DMSO: 125 mg/mL (325.06 mM)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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