Benzocaine 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

Benzocaine Ointment contains NLT 90.0% and NMT 110.0% of the labeled amount of benzocaine (C₉H₁₁NO₂) in a suitable ointment base.

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. The UV spectrum of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay.

3 ASSAY

3.1 PROCEDURE

Solution A: 0.1% Trifluoroacetic acid, prepared by adding 1.0 mL of trifluoroacetic acid to 1 L of water

Solution B: Acetonitrile

Mobile phase: See Table 1.

Table 1

Diluent: Solution A and Solution B (1:1)

Standard solution: 0.1 mg/mL of USP Benzocaine RS in Diluent. Sonication may be needed to aid in the dissolution.

Sample solution: Nominally 0.1 mg/mL of benzocaine in Diluent prepared as follows

Ointments having water-soluble bases: Transfer a portion of Ointment, equivalent to 10 mg of benzocaine, into a volumetric flask, and dissolve in Diluent.

Ointments having water-insoluble bases: Transfer a portion of Ointment, equivalent to 10 mg of benzocaine, into a volumetric flask, and dissolve in tetrahydrofuran, using about 2% of the final volume, then dilute with Diluent to volume. Pass through a suitable filter of 0.45-µm pore size, discarding the first 2-3 mL of the filtrate.

Chromatographic system

(See Chromatography (621), System Suitability.)

Mode: LC

Detector: UV 280 nm. For Identification test B, use a diode array detector in the range of 200-400 nm.

Column: 4.6-mm × 25-cm; 5-µm packing L7

Flow rate: 1.5 mL/min

Injection volume: 20 µL

System suitability

Sample: Standard solution

Suitability requirements

Tailing factor: NMT 1.5

Relative standard deviation: NMT 0.73%

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of the labeled amount of benzocaine (C₉H₁₁NO₂) in the portion of Ointment taken:

Result = (r_U/r_S) × (C_S/C_U) × 100

r_U = peak response of benzocaine from the Sample solution

r_S = peak response of benzocaine from the Standard solution

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

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

Acceptance criteria: 90.0%-110.0%

4 IMPURITIES

4.1 ORGANIC IMPURITIES

Solution A: 0.1% Trifluoroacetic acid, prepared by adding 1.0 mL of trifluoroacetic acid to 1 L of water

Solution B: Acetonitrile

Mobile phase: See Table 1 in the Assay.

Diluent: Solution A and Solution B (1:1)

System suitability solution: 0.2 mg/mL of USP Benzocaine RS and 0.01 mg/mL each of USP Aminobenzoic Acid RS and USP Ethyl 4-Nitrobenzoate RS in Diluent

Standard solution: 1 µg/mL each of USP Benzocaine RS, USP Aminobenzoic Acid RS, and USP Ethyl 4-Nitrobenzoate RS in Diluent

Sample solution: Nominally 1 mg/mL of benzocaine in Diluent prepared as follows

Ointments having water-soluble bases: Transfer a portion of Ointment, equivalent to 50 mg of benzocaine, into a volumetric flask, and dissolve in Diluent.

Ointments having water-insoluble bases: Transfer a portion of Ointment, equivalent to 50 mg of benzocaine, into a volumetric flask, and dissolve in about 10% of final volume of tetrahydrofuran, then dilute with Diluent to volume. Pass through a suitable filter of 0.45-µm pore size, discarding the first 2-3 mL of the filtrate.

Chromatographic system

(See Chromatography (621), System Suitability.)

Mode: LC

Detector: UV 280 nm

Column: 4.6-mm × 25-cm; 5-µm packing L7

Flow rate: 1.5 mL/min

Injection volume: 20 µL

System suitability

Samples: System suitability solution and Standard solution

Suitability requirements

Resolution: NLT 10 between aminobenzoic acid and benzocaine, and between benzocaine and ethyl 4-nitrobenzoate, System suitability solution

Relative standard deviation: NMT 2.0% for each peak corresponding to benzocaine, aminobenzoic acid, and ethyl 4-nitrobenzoate, Standard solution

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of aminobenzoic acid and ethyl 4-nitrobenzoate in the portion of Ointment taken:

Result = (r_U/r_S) × (C_S/C_U) × 100

r_U = peak response of aminobenzoic acid or ethyl 4-nitrobenzoate from the Sample solution

r_S = peak response of the corresponding Reference Standard from the Standard solution

C_S = concentration of USP Aminobenzoic Acid RS or USP Ethyl 4-Nitrobenzoate RS in the Standard solution (mg/mL)

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

Calculate the percentage of any other individual unspecified impurity in the portion of Ointment taken:

Result = (r_U/r_S) × (C_S/C_U) × 100

r_U = peak response for any other individual unspecified impurity from the Sample solution

r_S = peak response of benzocaine from the Standard solution

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

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

Acceptance criteria: See Table 2. Disregard peaks less than 0.05%.

Table 2

5 PERFORMANCE TESTS

MINIMUM FILL (755): Meets the requirements

6 SPECIFIC TESTS

MICROBIAL ENUMERATION TESTS (61) and TESTS FOR SPECIFIED MICROORGANISMS (62): It meets the requirements of the tests for absence of

Staphylococcus aureus and Pseudomonas aeruginosa.

7 ADDITIONAL REQUIREMENTS

PACKAGING AND STORAGE: Preserve in tight containers, protected from light, and store at room temperature at 15°-25°.

USP REFERENCE STANDARDS (11)

USP Aminobenzoic Acid RS

Benzoic acid, 4-amino.

C₇H₇NO₂  137.14

USP Benzocaine RS

USP Ethyl 4-Nitrobenzoate RS

Benzoic acid, 4-nitro-, ethyl ester.

C₉H₉NO₄  195.17