Estradiol and Norethindrone Acetate Tablets

This article is compiled based on the United States Pharmacopeia (USP) – 2025 Edition

Issued and maintained by the United States Pharmacopeial Convention (USP)

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1 DEFINITION

Estradiol and Norethindrone Acetate Tablets contain NLT 90.0% and NMT 110.0% of the labeled amount of estradiol (C₁₈H₂₄O₂) and NLT 90.0% and NMT 110.0% of the labeled amount of norethindrone acetate (C₂₂H₂₈O₃).

2 IDENTIFICATION

A. The UV spectra of the major peaks of the Sample solution correspond to those of the Standard solution, as obtained in the Assay.

8. The retention time of the major peaks of the Sample solution correspond to those of the Standard solution, as obtained in the Assay.

3 ASSAY

PROCEDURE

Mobile phase: Acetonitrile and water (55:45)

Diluent: Dehydrated alcohol and water (50:50)

Estrone standard stock solution: 0.12 mg/mL of USP Estrone RS in dehydrated alcohol

Estradiol standard stock solution: 0.25 mg/mL of USP Estradiol RS in dehydrated alcohol

Norethindrone acetate standard stock solution: 0.15 mg/mL of USP Norethindrone Acetate RS in dehydrated alcohol

System suitability solution: Combine 800 µL of the Estradiol standard stock solution, 600 µL of the Norethindrone acetate standard stock solution, 200 µl of the Estrone standard stock solution, and 10.0 mL of Diluent.

Standard solution: 20 µg/ml of USP Estradiol RS from the Estradiol standard stock solution and 10 µg/mL of USP Norethindrone Acetate RS from the Norethindrone acetate standard stock solution in Diluent

Sample solution: Nominally 20 µg/ml, of estradiol and 10 µg/ml, of norethindrone acetate from Tablets (NLT 12) in Diluent

Chromatographic system

(See Chromatography (621) System Suitability)

Mode: LC

Detector: UV dual wavelength (254 nm/280 nm) or equivalent. For Identification A, use a diode array detector in the range of 190-400 nm.

[No-The absorption of estradiol at 280 nm and norethindrone acetate at 254 nm can be included in a single run by altering the wavelength.]

Column: 4.6-mm x 15-cm, 4-μm packing L1

Flow rate: 1 mL/min

Injection volume: 50 µL

Run time: NLT 4 times the retention time of estradiol

System suitability

Samples: System suitability solution and Standard solution

Suitability requirements

Resolution: NLT 1.8 between estradiol and estrone, System suitability solution

Relative standard deviation: NMT 3%, Standard solution

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of the labeled amount of estradiol (C₁₈H₂₄O₂) in the portion of Tablets taken":

Result = (r_U/(r_S) × (C_S /C_U) x 100

r_U = peak response at 280 nm of estradiol from the Sample solution

r_S = peak response at 280 nm of estradiol from the Standard solution

C_S = concentration of USP Estradiol RS in the Standard solution (µg/mL)

C_U = nominal concentration of estradiol in the Sample solution (µg/mL)

Calculate the percentage of labeled amount of norethindrone acetate (C₂₂H₂₈O₃) in the portion of Tablets taken:

Result = (r_U/(r_S) × (C_S /C_U) x 100

r_U = peak response at 254 mm of norethindrone acetate from the Sample solution

r_S = peak response at 254 nm of norethindrone acetate from the Standard solution

C_S = concentration of USP harethindrone Acetate RS in the Standard solution (μg/mL)

C_U = nominal concentration of norethindrone acetate in the Sample solution (μg/mL)

Acceptance criteria

Estradiol: 90.0%–110.0%

Norethindrone acetate: 90.0%–110.0%

4 PERFORMANCE TESTS

Change to read:

Test 1

Medium: 0.3% dum laurd nictats 500 ml.

Apparatus 2: 50 грm

Times: 30 man for Tablets labeled to contain 1 mg of estradiol and 0.5 mg of nemethodrone acetate, and 50 man for Tablets labeled to contain 0.5 mg of estradiol and 1.1 mg of northindrone acetate

Mobile phase: Acetontile and water (55:45)

Standard stock solution A: 20 µg/ml of P Extradid AS in alohal or in a mixture of alcohol and water

Standard stock solution B: 10 µg/m of USP Norethindrone Acetate fi in alcohol or in a mixture of alcohol and water

Standard solution: Olute suitabile quantities of Standard rock solution A and Standard stock solution in Medium or a misure of and water to obtain a final concentration of both analytes similar to the expected concentration of the Sample solution

Sample colution: Pass a portion of the solution through a filter of 0.45-m pore size

Chromatographic system

(See Chromatography (621), System Suitability.)

Mode: LC

Detector: UV 241 nm for norethindrone acetate and 280 nm for estradiol

Column: 4.6-mm × 15-cm; packing L1

Flow rate: 1 mL/min

Injection volume: 150 μL

System suitability

Sample: Standard solution

Sultability requirements

Talling factor: NMT 15

Relative standard deviationc NMT 2.01%

Analysis

Samples: Sturda solution and Samale solution

Calculate the percentages of the labeled amounts of estradiol (C₁₈H₂₄O₂) and norethindrone acetate (C₂₂H₂₈O₃) dissolved:

Result = (r_U/(r_S) × C_S x V x (1/L) x 100

r_U = peak response of estradiol or norethindrone acetate from the Sample solution

r_S = peak response of estradiol or norethindrone acetate from the Standard solunion

C_S = concentration of USP Estradiol RS of USP Norethindrone Acetate RS in the Standant solution (mg/ml)

V = volume of Medium, 500 mL

L = label claim of estradiol or norethindrone acetate (mg/Tablet)

Tolerances: For Tablets labeled to contein 1 mg of estradiol and 0.5 mg of norettondrone acetate: NLT 75% (0) of the labeled amounts of estradiol (C₁₈H₂₄O₂) and norethindrone acetate (C₂₂H₂₈O₃) ) are dissolved in 30 min. For Tablets labeled to contain 0.5 mg of extradial and 0,1 mg of monethindrone acetate: NLT 75% (Q) of the labeled amounts of estradiol (C₁₈H₂₄O₂) and norethindrone acetate (C₂₂H₂₈O₃) are dissolved in 50 min.

Test 2: If the product compiles with this best, the labeling indicates that it meets USP Dissolution Test 2

Medium: 0.7% 10dum laund sulfate: 500 ml, desented, if necessary

Apparatus 2:50 rpm

Timer 20 min

Mobile phase: Austuntile and wale: (55:45)

Standard stock solution A: 0.5 mg/mL of USP Estradiol RS in methanol. Sonicate as necessary.

Standard stock solution B: 0.5 mg/mL of USP Norethindrone Acetate RS in methanol. Sonicate as necessary.

Standard solution: (L /500) mg/mL of USP Estradiol RS and (L /500) mg/mL of USP Norethindrone Acetate RS prepared from Standard stock solution A and Standard stock solution B in Medium, where L is the label claim of estradiol and L is the label claim of norethindrone acetate, in mg/Tablet

Sample solution: Pass a portion of the solution through a filter of 0.45-sm pore sz

Chromatographic system

(See Chromatogrank (9211 xstem Sutta)

Mode: LC

Detector: LV-241 nm for corethindrone acetate and 210 m for estradiol

Column: 4.6mm 15cm, 5m packing L

Temperatures

Autosampler: 20

Column: 30

Flow rate: 1 mL/min

Injection volume: 100 μL

Run time: NLT 1.3 times the retention time of norettindrone acelate

Syutam suitability

Sample: Standard solutice

Suitability requirements

Tailing factor: NMT 15 for estradaul and nonrthindione scetate

Relative standard deviation: NMT 2.0% for extradiol and norethindrone acetate

Analysis

Samples: Standord solution and Sample solution

Camulate the percentages of the labeleid amounts of estradiol (C₁₈H₂₄O₂) and norethindrone acetate (C₂₂H₂₈O₃) dissolved:

Result = (r_U/(r_S) × C_S x V x (1/L) x 100

r_U = peak response of extradiol or norethindrone acetate from the Sample solution

r_S = peak response of estradiol or norethindrone acetate from the Standard solution

C_S = concentration of USP Estradiol RS of USP Norethindrone Acetate RS in the Standant solution (mg/ml)

V = volume of Medium, 500 mL

L = label claim of estradiol or norethindrone acetate (mg/Tablet)

Tolerances: NLT 80% (2) of the labeled amounts of estradiol (C₁₈H₂₄O₂) and norethindrone acetate (C₂₂H₂₈O₃) are dissolved in 20 min

Uniformity of Dosage Units (905): Meet the requirements

5 IMPURITIES

Organic Impurities

Solution A: Tetrahydrofuran and water (1:200)

Solution B: Acetonitrile, tetrahydrofuran, and water (160:1:40)

Mobile phase: See Table 1.

Diluent: Debated alculiol and water (5050)

System suitability solution: 240 µg/mL of USP Estradiol RS 60 µg/ml. of P Norethindrone Acetate 25 and 1 pg/ml of Lis Extrone RS in

Estradini standani stock solution: 250 µg/ml. of USP Extrudi RS in sohel

Norethindrone acetate standard stock solution: 150 pgint of USP Nethnanne Acetate RS in akachal

Standard solution: Combine 250 µl of the Estmatkal standard stock cokat and 100 µl of the Norethindrone acetate standard sick solution and dilute with 500ml of bent.

Sample solution: Nominally 240 pg/ml, of estradiol and 120 µg/ml of norethindrone acetate from Tablets (NLT 12) inuent

Chromatographic system

Bee Chromatoguany/621) System Sullability)

Mode: LC

Detector: UV 235 and 254 nm

Column: 3.9-mm × 30-cm; 4-μm packing L1

Flow rate: 0.8 mL/min

Injection volume: 100 μL

System suitability

Sample: Systen tullability solution

Nom The relative retention times for estradiol, estrone, and norethindrone acetate are about 1.0, 1.1, and 1.7, respectively]

Suitability requirements

Resolution: NLT 1.3 between estrone and estradiol, measured at 254 nm

Analysis

Samples: Standid sution and Sample solution

Calculate the piccentages of any estradiol related impurities in the portion of Tablets taken:

Result = (r_U/(r_S) × (C_S /C_U) x (1/F) x 100

r_U = peak response at 235 nm for each impunity from the Sample solution

r_S = peak response at 225 nm from the Shandand solution

C_S = concentration of the Standant solution (mg/ml)

C_U = concentration of the Sample solution (mg/mL)

F = relative response factor (see Table 2 or Table 3)

Calculate the percentages of any norethindrone acetate related impurities in the portion of Tablets taken:

Result = (r_U/(r_S) × (C_S /C_U) x (1/F) x 100

r_U = peak response at 254 nm for each impunity from the Sample solution

r_S = peak response at 254 nm from the Shandand solution

C_S = concentration of the Standant solution (mg/ml)

C_U = concentration of the Sample solution (mg/mL)

F = relative response factor (see Table 2 or Table 3)

Acceptance criteria: The Tablets meet the requirements given in Table 2 xor Table Soe Table 2 for Tablems labeled to contain 1 mg of ectradial and 05 mg of norethindrone asetate or Table 3 for Tablets labeled to contain 0.5 mg of estradiol and 1.1 mg of nornithindrone acetate

Table 2. For Tablets Labeled as Containing 1 mg of Estradiol and 0.5 mg of Norethindrone Acetate

^a 1,3,5(10)-Estratriene-3,6α,17β-triol.

^b 1,3,5(10)-Estratriene-3,6β,17β-triol.

^c 1,3,5(10)-Estratrien-3,17β-diol-6 one.

^d 1,3,5(10),6-Estratetraen-3,17β-diol.

^e 6β-Hydroxy-3-oxo-19-nor-17α-pregn-4-en-20-yn-17-yl acetate.

^f 17-Hydroxy-19-nor-17α-pregn-4-en-20-yn-3-one.

^g 3,6-Dioxo-19-nor-17α-pregn-4-en-20-yn-17-yl acetate

^h 17-Hydroxy-19-nor-17α-pregn-4,6-dien-20-yn-3-one acetate.

Table 3. For Tablets Labeled as Containing 0.5 mg of Estradiol and 0.1 mg of Norethindrone Acetate

^a 1,3,5(10)-Estratrien-3,6β,17β-triol.

^b 6β-Hydroxy-3-oxo-19-nor-17α-pregn-4-en-20-yn-17-yl acetate

^c 17-Hydroxy-19-nor-17α-pregn-4-en-20-yn-3-one.

^d 3,6-Dioxo-19-nor-17α-pregn-4-en-20-yn-17-yl acetate.

^e 17-Hydroxy-19-nor-17α-pregn-4,6-dien-20-yn-3-one acetate.

SPECIFIC TESTS

• Microbial Enumeration Tests (61) and Tests for Specified Microorganisms (62): The total aerobic microbial count is NMT 10³ cfu/g. The total combined molds and yeasts count is NMT 10² cfu/g. Meets the requirements of the tests for the absence of Salmonella species and Escherichia coli.

6 ADDITIONAL REQUIREMENTS

Packaging and Storage: Preserve in well-closed containers, and store at controlled room temperature.

Labeling: When more than one Dissolution test is given, the labeling states the Dissolution test used only if Test 1 is not used.