Neomycin and Polymyxin B Sulfates, Bacitracin, and Hydrocortisone Acetate Ophthalmic Ointment

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

Neomycin and Polymyxin B Sulfates, Bacitracin, and Hydrocortisone Acetate Ophthalmic Ointment contains the equivalent of NLT 90.0% and NMT 140.0% of the labeled amounts of neomycin, polymyxin B, and bacitracin, and NLT 90.0% and NMT 110.0% of the labeled amount of hydrocortisone acetate in a suitable ointment base.

2 IDENTIFICATION

A. Thin-Layer Chromatographic Identification Test 〈201BNP〉: Meets the requirements

B. The retention time of the hydrocortisone acetate peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay for Hydrocortisone Acetate.

3 ASSAY

3.1 Neomycin

(See Antibiotics-Microbial Assays 〈81〉.)

Sample solution: Shake a portion of Ophthalmic Ointment in a separator with 50 mL of ether. Extract with four 20-mL portions of Buffer B.3. Combine the aqueous extracts, and dilute with Buffer B.3 to a suitable volume.

Analysis: Proceed as directed in the chapter. Dilute the Sample solution with Buffer B.3 to obtain a Test Dilution having a neomycin concentration that is nominally equivalent to the median level of the standard.

Acceptance criteria: 90.0%–140.0%

3.2 Polymyxin B

(See Antibiotics-Microbial Assays 〈81〉.)

Sample solution: Shake a portion of Ophthalmic Ointment with 50 mL of ether in a separator. Extract with four 25-mL portions of Buffer B.6. Combine the aqueous extracts, and dilute with Buffer B.6 to a suitable volume.

Analysis: Proceed as directed in the chapter. Dilute the Sample solution with Buffer B.6 to obtain a Test Dilution having a concentration that is nominally equivalent to the median level of the standard (10 polymyxin B units/mL). Add to each Test Dilution of the standard a quantity of USP Neomycin Sulfate RS, dissolved in Buffer B.6, to obtain the same concentration of neomycin as in the Test Dilution of the sample.

Acceptance criteria: 90.0%–140.0%

3.3 Bacitracin

(See Antibiotics-Microbial Assays 〈81〉.)

Sample solution: Shake a portion of Ophthalmic Ointment with 50 mL of ether in a separator. Extract with four 20-mL portions of Buffer B.1. Combine the buffer extracts, and dilute with Buffer B.1 to a suitable volume.

Analysis: Proceed as directed in the chapter. Add sufficient 0.01 N hydrochloric acid to a portion of the Sample solution so that the amount of hydrochloric acid in the Test Dilution is the same as in the median level of the standard. Dilute with Buffer B.1 to obtain a Test Dilution having a Bacitracin concentration that is nominally equivalent to the median level of the standard.

Acceptance criteria: 90.0%–140.0%

3.4 Hydrocortisone Acetate

Mobile phase: Butyl chloride, water-saturated butyl chloride, tetrahydrofuran, methanol, and glacial acetic acid (475:475:70:35:30)

Standard solution: 0.10 mg/mL of USP Hydrocortisone Acetate RS in water-saturated chloroform

Sample solution: Nominally 0.10 mg/mL of hydrocortisone acetate from Ophthalmic Ointment prepared as follows. Transfer a portion of Ophthalmic Ointment containing nominally 2.5 mg of hydrocortisone acetate to a closable container. Add 25.0 mL of water-saturated chloroform and about 10 glass beads. Securely close the container, and shake vigorously for approximately 15 min. Centrifuge, and use the clear, lower chloroform layer.

Chromatographic system

• (See Chromatography 〈621〉, System Suitability.)
• Mode: LC
• Detector: UV 254 nm
• Column: 3.9-mm × 30-cm; 10-µm packing L3

System suitability

• Sample: Standard solution
• Suitability requirements
• Relative standard deviation: NMT 2.0%

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of the labeled amount of hydrocortisone acetate (C₂₃H₃₂O₆) in the portion of Ophthalmic Ointment taken:

Result = (rᵤ/rₛ) × (Cₛ/Cᵤ) × 100

rᵤ = peak response from the Sample solution

rₛ = peak response from the Standard solution

Cₛ = concentration of USP Hydrocortisone Acetate RS in the Standard solution (mg/mL)

Cᵤ = nominal concentration of hydrocortisone acetate in the Sample solution (mg/mL)

Acceptance criteria: 90.0%–110.0%

4 SPECIFIC TESTS

Sterility Tests 〈71〉: Meets the requirements

Other Requirements: It meets the requirements for Particulate and Foreign Matter and Container Contents in Ophthalmic Products—Quality Tests 〈771〉, Drug Product Quality, Universal Tests, Particulate and Foreign Matter and Container Contents.

5 ADDITIONAL REQUIREMENTS

Packaging and Storage: Preserve in collapsible ophthalmic ointment tubes.

USP Reference Standards 〈11〉

USP Bacitracin Zinc RS

USP Hydrocortisone Acetate RS

USP Neomycin Sulfate RS

USP Polymyxin B Sulfate RS