Oseltamivir Phosphate

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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C₁₆H₂₈N₂O₄ · H₃PO₄ 410.40

[3R-(3α,4β,5α)]-Ethyl 4-(acetylamino)-5-amino-3-(1-ethylpropoxy)-1-cyclohexene-1-carboxylate phosphate (1:1);

Ethyl (3R,4R,5S)-4-acetamido-5-amino-3-(1-ethylpropoxy)-1-cyclohexene-1-carboxylate, phosphate (1:1) CAS RN ®: 204255-11-8; UNII: 4A3O49NGEZ.

1 DEFINITION

Oseltamivir Phosphate contains NLT 98.0% and NMT 101.5% of C₁₆H₂₈N₂O₄ · H₃PO₄ , calculated on the anhydrous basis.

2 IDENTIFICATION

A. Spectroscopic Identification Tests 〈197〉, Infrared Spectroscopy: 197M

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

3 ASSAY

Procedure

Solution A: Dissolve 6.8 g of potassium dihydrogen phosphate in 980 mL of water. Adjust with 1 M potassium hydroxide solution to a pH of 6.0, and dilute with water to 1 L.

Mobile phase: Methanol, acetonitrile, and Solution A (245:135:620)

Diluent: Methanol, acetonitrile, and water (245:135:620)

Standard solution: 1 mg/mL of USP Oseltamivir Phosphate RS in Diluent

Sample solution: 1 mg/mL of Oseltamivir Phosphate in Diluent

Chromatographic system

(See Chromatography 〈621〉, System Suitability.)

Mode: LC

Detector: UV 207 nm

Column: 4.6-mm × 25-cm; packing L7

Column temperature: 50°

Flow rate: 1.2 mL/min

Injection size: 15 μL

System suitability

Sample: Standard solution

Suitability requirements

Tailing factor: NMT 2.0

Relative standard deviation: NMT 2.0%

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of C₁₆H₂₈N₂O₄ · H₃PO₄ in the portion of Oseltamivir Phosphate taken:

Result = (r_U/r_S) × (C_S/C_U) × 100

r_U = peak response from the Sample solution

r_S = peak response from the Standard solution

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

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

Acceptance criteria: 98.0%–101.5% on the anhydrous basis

4 IMPURITIES

Change to read:

Organic Impurities

4.1 Procedure 1

Solution A, Mobile phase, Diluent, Standard solution, Sample solution, and Chromatographic system: Proceed as directed in the Assay.

Analysis

Sample: Sample solution

Calculate the percentage of each impurity in the portion of Oseltamivir Phosphate taken:

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

r_U = peak response of each individual impurity from the Sample solution

r_S = peak response of oseltamivir phosphate from the Standard solution

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

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

F = relative response factor from Impurity Table 1

Acceptance criteria

Individual impurities: See Impurity Table 1.

Total impurities: NMT 0.7%

Impurity Table 1

^a(3R,4R,5S)-4-Acetylamino-5-amino-3-(1-ethylpropoxy)-1-cyclohexene-1-carboxylic acid.

^b 4-Acetylamino-3-hydroxybenzoic acid ethyl ester.

4.2 Procedure 2: Oseltamivir Related Compound A

Buffer: 1.54 g/L of ammonium acetate in water

Mobile phase: Acetonitrile, water, and Buffer (3:6:1)

Stock solution A: 50 μg/mL of USP Oseltamivir Related Compound A RS, prepared as follows: Dissolve in alcohol, using 5% of final volume, and dilute with water to volume.

Solution A: 1 μg/mL of USP Oseltamivir Related Compound A RS in water from Stock solution A

Standard solution: 10 mg/mL of USP Oseltamivir Phosphate RS in Solution A

Sample solution: 10 mg/mL of Oseltamivir Phosphate in water

Chromatographic system

(See Chromatography 〈621〉, System Suitability.)

Mode: LC

Detectors: UV 210 nm and mass spectrometer

Column: 3.0-mm × 5-cm; 5-μm packing L1

Flow rate: 1.5 mL/min

Injection size: 1 μL

Temperature: 40°

Use electrospray (+) ionization, a selected ion monitoring mode with m/z of 356.2 (protonated oseltamivir related compound A). Adjust the dwell time, fragmentation voltage, drying gas temperature, drying gas flow, nebulizer pressure, and capillary voltage as appropriate for an optimal response. [Note - A postcolumn flow splitter with a split ratio of about 3:1 is used.]

System suitability

Sample: Standard solution

[Note - The relative retention time for oseltamivir related compound A versus oseltamivir is about 2.6.]

Suitability requirements

Resolution: The oseltamivir related compound A peak (detected by MD-SIM mode) and the oseltamivir peak (detected by UV) are baseline resolved.

[Note - The resolution of the two components minimizes background noise and ion suppression effects for the trace of oseltamivir related compound A by the oseltamivir matrix.]

Relative standard deviation: NMT 15.0%, oseltamivir related compound A peak

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of oseltamivir related compound A in the portion of Oseltamivir Phosphate taken:

Result = (r_U/r_S) × (C_S/C_U) × 100

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

r_S = peak response of oseltamivir related compound A from the Standard solution

C_S = concentration of USP Oseltamivir Related Compound A RS in the Standard solution (mg/mL)

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

Acceptance criteria: NMT 0.01%

4.3 Procedure 3: Limit Of Tributyl Phosphine Oxide

Blank: Transfer 1.0 mL of suitable derivatizing reagent1 to a vial. Close the vial, shake, and heat for 20 min at 60°. Centrifuge the pyridinium salt precipitate.

Standard stock solution 1: 21 mg/mL of USP Tributyl Phosphine Oxide RS in pyridine

Standard stock solution 2: 21 mg/mL of USP Oseltamivir Phosphate RS in suitable derivatizing reagent. Close the vial, mix, and heat for 20 min at 60°. Centrifuge the pyridinium salt precipitate.

Standard solution: 21 μg/mL each of USP Tributyl Phosphine Oxide RS and USP Oseltamivir Phosphate RS in pyridine from Standard stock solution 1 and Standard stock solution 2, respectively

Sample solution: Transfer 15 mg of Oseltamivir Phosphate to a vial. Add 1.0 mL of suitable derivatizing reagent. Close the vial, mix, and heat for 20 min to 60°. Centrifuge the pyridinium salt precipitate.

Chromatographic system

(See Chromatography 〈621〉, System Suitability.)

Mode: GC

Detector: Flame ionization

Column: 0.32-mm × 30-m capillary column coated with a 0.25-μm phase G1

Split ratio: 1:50

Split flow: 64 mL/min

Injection size: 1 μL

Temperature

Detector: 260°

Injection port: 260°

Column: See the temperature program table below.

Linear velocity: 27 cm/s

Carrier gas: Helium

System suitability

Sample: Standard solution

[Note - The relative retention times for tributyl phosphine oxide and oseltamivir phosphate are about 0.54 and 1.00, respectively.]

Suitability requirements

Relative standard deviation: NMT 10.0% for the tributyl phosphine oxide and oseltamivir (ERR 1-Jun-2023) phosphate peaks

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of tributyl phosphine oxide in the portion of Oseltamivir Phosphate taken:

Result = (r_U/r_S) × (C_S/C_U) × 100

r_U = peak response of tributyl phosphine oxide from the Sample solution

r_S = peak response of tributyl phosphine oxide from the Standard solution

C_S = concentration of tributyl phosphine oxide in the Standard solution (mg/mL)

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

Acceptance criteria: NMT 0.1%

5 SPECIFIC TESTS

 Water Determination, Method I〈921〉: NMT 0.5%

Optical Rotation, Specific Rotation 〈781S〉

Sample solution: 10 mg/mL in water

Acceptance criteria: Between –30.7 and –32.6

6 ADDITIONAL REQUIREMENTS

Packaging and Storage: Preserve in well-closed containers. Store at 25°, excursions permitted between 15° and 30°.

USP Reference Standards 〈11〉

USP Oseltamivir Phosphate RS

USP Oseltamivir Related Compound A RS

(3S,4R,5S)-Ethyl 4-acetamido-5-amino-2-azido-3-(pentan-3-yloxy)cyclohexanecarboxylate.

C₁₆H₂₉N₅O₄ 355.43

USP Tributyl Phosphine Oxide RS C H OP 218.32

¹ Tri-Sil Reagent (product number: 48999 0049001) may be obtained from Pierce: www.piercenet.com.