Polymyxin B Sulfate and Trimethoprim Ophthalmic Solution

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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Polymyxin B Sulfate and Trimethoprim Ophthalmic Solution is a sterile, isotonic, aqueous solution of Polymyxin B Sulfate and Trimethoprim Sulfate or of Polymyxin B Sulfate and Trimethoprim that has been solubilized with Sulfuric Acid. It contains not less than 90.0 percent and not more than 130.0 percent of the labeled amount of polymyxin B and the equivalent of not less than 90.0 percent and not more than 110.0 percent of the labeled amount of trimethoprim (C₁₄H₁₈N₄O₃). It contains one or more preservatives.

Packaging and storage—Preserve in tight, light-resistant containers, and store at controlled room temperature.

Labeling—Label it to indicate that it is to be stored at 15° to 25°, protected from light.

USP Reference standards 〈11〉—

USP Polymyxin B Sulfate RS

USP Trimethoprim RS

Identification—

A: It meets the requirements for polymyxin B under Thin-Layer Chromatographic Identication Test 〈201BNP〉.

B: The retention time of the trimethoprim peak in the chromatogram of the Assay preparation corresponds to that in the chromatogram of the Standard preparation, as obtained in the Assay for trimethoprim.

Sterility Tests 〈71〉 —It meets the requirements when tested as directed for Membrane Filtration under Test for Sterility of the Product to be Examined.

pH 〈791〉: between 4.0 and 6.2.

Assay for polymyxin B—Proceed as directed for polymyxin B under Antibiotics—Microbial Assays 〈81〉, using an accurately measured volume of Ophthalmic Solution, diluted quantitatively and stepwise with Buffer B.6, to obtain a Test Dilution having a concentration of polymyxin B assumed to be equal to the median dose level of the Standard.

Assay for trimethoprim—

Diluent—Prepare a mixture of 0.01 N hydrochloric acid and acetonitrile (870:130).

Mobile phase—Dissolve 1.65 g of ethanesulfonic acid in 1000 mL of a mixture of water and acetonitrile (870:130). Adjust with 10 N sodium hydroxide or 0.1 N hydrochloric acid to a pH of 3.5. Pass this solution through a filter having a 0.5-µm or finer porosity, and degas. Make adjustments if necessary (see System Suitability under Chromatography 〈621〉).

Standard preparation—Dissolve an accurately weighed quantity of USP Trimethoprim RS in Diluent to obtain a solution having a known concentration of about 0.04 mg per mL.

Assay preparation—Transfer an accurately measured volume of Ophthalmic Solution, equivalent to about 1 mg of trimethoprim, to a 25-mL volumetric ask, dilute with Diluent to volume, and mix.

Chromatographic system (see Chromatography 〈621〉)—The liquid chromatograph is equipped with a 254-nm detector and a 3.9-mm × 30-cm column that contains packing L11. The flow rate is about 1.5 mL per minute. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the tailing factor is not more than 1.5, when calculated at 10% height of the peak; and the relative standard deviation for replicate injections is not more than 2.0%.

Procedure—Separately inject equal volumes (about 10 µL) of the Standard preparation and the Assay preparation into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the quantity, in mg, of trimethoprim (C₁₄H₁₈N₄O₃) in each mL of the Ophthalmic Solution taken by the formula:

25(C/V)(r_U /r_S )

in which C is the concentration, in mg per mL, of USP Trimethoprim RS in the Standard preparation; V is the volume, in mL, of Ophthalmic Solution taken to prepare the Assay preparation; and r_U and r_S are the trimethoprim peak area responses obtained from the Assay preparation and the Standard preparation, respectively.