Warfarin Sodium 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

Warfarin Sodium Tablets contain NLT 95.0% and NMT 105.0% of the labeled amount of warfarin sodium (C₁₉H₁₅NaO₄).

2 IDENTIFICATION

A. The retention time of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay.

B. SPECTROSCOPIC IDENTIFICATION TESTS (197), Infrared Spectroscopy: 197K

Sample: Triturate a quantity of finely powdered Tablets, equivalent to about 200 mg of warfarin sodium, with 50 mL of water, centrifuge, and filter the supernatant. Extract with 50 mL of ether, transfer the aqueous layer to a second separator, and discard the ether. Adjust with hydrochloric acid to a pH of less than 3, using short-range pH indicator paper, and extract with 50 mL of chloroform. Transfer the chloroform layer to another separator, extract with 50 mL of sodium hydroxide solution (1 in 250), and discard the chloroform. Transfer the aqueous layer to a beaker, and adjust with hydrochloric acid to a pH of less than 3 (using the pH indicator paper) to precipitate the warfarin. Stir the mixture, and allow the precipitate to coagulate. Filter, and wash the precipitate with four, 5-mL portions of water. If the precipitate is not white or practically white, dissolve it in a minimum volume of sodium hydroxide solution (1 in 250), dilute with water to 50 mL, and repeat the foregoing procedure, beginning with "Extract with 50 mL of ether." Dry the warfarin under vacuum over phosphorus pentoxide for 4 h.

3 ASSAY

PROCEDURE

Buffer: Transfer 1.36 g of monobasic potassium phosphate to a 200-mL volumetric flask, and dissolve in 50 mL of water. Add 39.1 mL of 0.2 N sodium hydroxide, and dilute with water to volume. Adjust with sodium hydroxide or phosphoric acid to a pH of 7.4 ± 0.1.

Diluent: Acetonitrile and Buffer (15:85)

Mobile phase: Methanol, glacial acetic acid, and water (68:1:32)

Standard stock solution: 0.31 mg/mL of USP Warfarin RS prepared as follows. Transfer USP Warfarin RS to a suitable volumetric flask, and dissolve in 0.1 N sodium hydroxide equivalent to 39% of the final volume. Add 0.2 M monobasic potassium phosphate, equivalent to 25% of the final volume, and dilute with water to volume.

Standard solution: Transfer 15.0 mL of Standard stock solution into a 50-mL volumetric flask, and dilute with Diluent to volume.

Sample solution: Nominally 0.1 mg/mL of warfarin sodium in Diluent prepared as follows. Weigh, and finely powder NLT 20 Tablets. Transfer an accurately weighed portion of the powder, equivalent to about 5 mg of warfarin sodium, to a 50-mL volumetric flask, and add about 30 mL of Diluent. Sonicate for 10 min, and then shake by mechanical means for 60 min. Dilute with Diluent to volume, and filter.

Chromatographic system

(See Chromatography (621), System Suitability.)

Mode: LC

Detector: UV 280 nm

Column: 4.6-mm x 25-cm; packing L7

Flow rate: 1.4 mL/min

Injection volume: 20 µL

System suitability

Sample: Standard solution

Suitability requirements

Relative standard deviation: NMT 1.0%

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of the labeled amount of warfarin sodium (C₁₉H₁₅NaO₄) in the portion of Tablets taken:

                         Result = (r_U/r_S) × (C_S/C_U) x (M_r1/M_r2) × 100

r_U = peak response of warfarin from the Sample solution

r_S = peak response of warfarin from the Standard solution 

C_S = concentration of USP Warfarin RS in the Standard solution (mg/mL) 

C_U = nominal concentration of warfarin sodium in the Sample solution (mg/mL) 

M_r1 = molecular weight of warfarin sodium, 330.31

M_r2 = molecular weight of warfarin, 308.33 

Acceptance criteria: 95.0%-105.0%

4 PERFORMANCE TESTS

4.1 DISSOLUTION (711)

Medium: Water; 900 mL.

Apparatus 2: 50 rpm

Time: 30 min

Mobile phase: Prepare as directed in the Assay.

Standard solution: USP Warfarin RS in water, having a known concentration of 0.0008L mg/mL, L being the labeled amount, in mg, of warfarin sodium in the Tablets. [NOTE-Use a small amount of 0.1 N sodium hydroxide to aid in dissolution.]

Sample solution: Pass a portion of the solution under test through a suitable filter.

Chromatographic system: Proceed as directed in the Assay except the Injection volume.

Injection volume: 40 µL

System suitability: Proceed as directed in the Assay.

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of the labeled amount of warfarin sodium (C₁₉H₁₅NaO₄) dissolved:

                         Result = (r_U/r_S) × C_S × V × (M_r1/M_r2) × (1/L) x 100

r_U = peak response of warfarin from the Sample solution

r_S = peak response of warfarin from the Standard solution

C_S = concentration of USP Warfarin RS in the Standard solution (mg/mL) 

V = volume of Medium, 900 mL 

M_r1 = molecular weight of warfarin sodium, 330.31

M_r2 = molecular weight of warfarin, 308.33

L = label claim (mg/Tablet)

Tolerances: NLT 80% (Q) of the labeled amount of warfarin sodium (C₁₉H₁₅NaO₄) is dissolved.

4.2 UNIFORMITY OF DOSAGE UNITS (905)

Meet the requirements

5 IMPURITIES

ORGANIC IMPURITIES

Diluent: Methanol and water (25:75). Alternatively, a mixture of acetonitrile and water (25:75) can be used.

Mobile phase: Methanol, tetrahydrofuran, glacial acetic acid, and water (57:1:1:42)

Standard stock solution: 0.126 mg/mL of USP Warfarin RS prepared as follows. Transfer USP Warfarin RS to a suitable volumetric flask, and add 0.1 N sodium hydroxide and methanol equivalent to 2% and 25% of the final volume, respectively. Sonicate, if necessary, to dissolve. Dilute with water to volume.

Standard solution (for 1-mg Tablets): 1.0 µg/mL of USP Warfarin RS in Diluent from Standard stock solution

Standard solution (for other strengths): 2.0 µg/mL of USP Warfarin RS in Diluent from Standard stock solution

System suitability stock solution: 0.4 mg/mL of USP Warfarin Related Compound A RS in methanol

System suitability solution: 2.0 µg/mL of each of USP Warfarin RS and USP Warfarin Related Compound A RS in Diluent from Standard stock solution and System suitability stock solution

Sample solution (for 1-mg Tablets): Nominally 0.4 mg/mL of warfarin sodium in Diluent prepared as follows. Transfer 10 Tablets into a suitable volumetric flask, dissolve Tablets in Diluent by sonicating for 10 min and shaking for 60 min. Dilute with Diluent to volume. Pass through a suitable filter, and discard the first few mL.

Sample solution (for other strengths): Nominally 0.8 mg/mL of warfarin sodium in Diluent prepared as follows. Transfer 8-10 Tablets into a suitable volumetric flask, dissolve Tablets in Diluent by sonicating for 10 min and shaking for 60 min. Dilute with Diluent to volume. Pass through a suitable filter, and discard the first few mL.

Chromatographic system

(See Chromatography (621), System Suitability.)

Mode: LC

Detector: UV 280 nm

Column: 4.6-mm x 25-cm; packing L11

Column temperature: 35°

Flow rate: 1.5 mL/min

Injection volume: 200 µL

Run time: NLT 4 times the retention time of warfarin

System suitability

Samples: Standard solution and System suitability solution

[NOTE-The relative retention times for warfarin and warfarin related compound A are about 1.0 and 1.4, respectively.]

Suitability requirements

Resolution: NLT 2 between warfarin and warfarin related compound A peaks, System suitability solution

Relative standard deviation: NMT 5.0%, Standard solution

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of Alice's ketone in the portion of Tablets taken:

                         Result = (r_U/r_S) × (C_S/C_U) x (M_r1/M_r2) × (1/F) x 100

r_U = peak response of warfarin related compound A from the Sample solution

r_S = peak response of warfarin from the Standard solution

C_S  = concentration of USP Warfarin RS in the Standard solution (mg/mL)

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

M_r1 = molecular weight of warfarin sodium, 330.31

M_r2 = molecular weight of warfarin, 308.33 

F = relative response factor for Alice's ketone, 0.9

[NOTE-Alice's ketone is a sodium salt of warfarin related compound A, 3-(o-hydroxyphenyl)-5-phenyl-2-cyclohexen-1-one sodium salt.]

Acceptance criteria: NMT 0.5%

6 ADDITIONAL REQUIREMENTS

PACKAGING AND STORAGE: Preserve in tight, light-resistant containers. Store at controlled room temperature.

Change to read:

USP REFERENCE STANDARDS (11)

USP Warfarin RS

USP. Warfarin Related Compound ARS

3-(^▲2_{▲(ERR 1-Feb-2022)} -Hydroxyphenyl)-5-phenyl-2-cyclohexen-1-one.

C₁₈H₁₆O₂             ^▲264.32_{▲(ERR 1-Feb-2022)}