Cloxacillin Sodium

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

Cloxacillin Sodium contains the equivalent of NLT 825 µg/mg of cloxacillin (C₁₉H₁₈CIN₃O₅S).

2 IDENTIFICATION

Change to read:

A. SPECTROSCOPIC IDENTIFICATION TESTS (197), Infrared Spectroscopy: 197KA (CN 1-MAY-2020)

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

C. IDENTIFICATION TESTS-GENERAL, Sodium (191): Meets the requirements

3 ASSAY

3.1 PROCEDURE

Protect solutions containing cloxacillin from light.

Solution A: 1.18 g/L of sodium 1-hexanesulfonate monohydrate and 0.8 mL/L of ammonium hydroxide in water, adjusted with phosphoric acid to a pH of 2.9-3.1

Solution B: Acetonitrile

Mobile phase: See Table 1.

Table 1

Return to the original conditions and re-equilibrate the system.

Diluent: Acetonitrile and water (50:50)

System suitability stock solution: 0.1 mg/mL of USP Cloxacillin Related Compound D RS in Diluent. Sonicate as needed to dissolve.

System suitability solution: 0.001 mg/mL of USP Cloxacillin Related Compound D RS from System suitability stock solution and 0.1 mg/mL of

USP Cloxacillin Sodium RS in Diluent. Store this solution at 4°.

Standard solution: 0.1 mg/mL of USP Cloxacillin Sodium RS in Diluent. Sonicate as needed to dissolve. Store this solution at 4°.

Sample solution: 0.1 mg/mL of Cloxacillin Sodium in Diluent. Sonicate as needed to dissolve. Store this solution at 4°.

Chromatographic system

(See Chromatography (621), System Suitability.)

Mode: LC

Detector: UV 225 nm

Column: 4.6-mm × 25-cm; 5-µm packing L1

Temperatures

Column: 40°

Autosampler: 4°

Flow rate: 1.5 mL/min

Injection volume: 10 µL

System suitability

Samples: System suitability solution and Standard solution

[NOTE-The relative retention times for cloxacillin and cloxacillin related compound D are about 1.0 and 1.1, respectively.]

Suitability requirements

Resolution: NLT 1.5 between cloxacillin and cloxacillin related compound D, System suitability solution

Tailing factor: 0.8-1.5, Standard solution

Relative standard deviation: NMT 0.73%, Standard solution

Analysis

Samples: Standard solution and Sample solution

Calculate the quantity, in µg/mg, of cloxacillin (C19H18CINOS) in the portion of Cloxacillin Sodium taken:

Result = (r_u/r_s) × (C_s/C_u) × P

r_u = peak response from the Sample solution

r_s= peak response from the Standard solution s

C_s = concentration of USP Cloxacillin Sodium RS in the Standard solution (mg/mL)

C_u = concentration of Cloxacillin Sodium in the Sample solution (mg/mL)

P = potency of cloxacillin in USP Cloxacillin Sodium RS (µg/mg)

Acceptance criteria: NLT 825 µg/mg

4 IMPURITIES

4.1 ORGANIC IMPURITIES

Protect solutions containing cloxacillin from light.

Solution A, Solution B, Diluent, and Chromatographic system: Proceed as directed in the Assay.

Mobile phase: See Table 2.

Table 2

Return to the original conditions and re-equilibrate the system.

System suitability stock solution: 0.1 mg/mL of USP Cloxacillin Related Compound D RS in Diluent

System suitability solution: 0.01 mg/mL of USP Cloxacillin Related Compound D RS from System suitability stock solution and 1 mg/mL of USP Cloxacillin Sodium RS in Diluent. Store this solution at 4°.

Standard solution: 0.01 mg/mL of USP Cloxacillin Sodium RS in Diluent. Sonicate as needed to dissolve. Store this solution at 4°.

Sample solution: 1 mg/mL of Cloxacillin Sodium in Diluent. Sonicate as needed to dissolve. Store this solution at 4°.

System suitability

Samples: System suitability solution and Standard solution

Suitability requirements

Resolution: NLT 1.5 between cloxacillin related compound D and cloxacillin, System suitability solution

Tailing factor: 0.8-1.5, Standard solution

Relative standard deviation: NMT 2.5%, Standard solution

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of each impurity in the portion of Cloxacillin Sodium taken:

Result = (r_u/r_s) × (C_s/C_u) × Px (F₁/F₂) × 100

r_u = peak response of each impurity from the Sample solution

r_s = peak response from the Standard solution s

C_s = concentration of USP Cloxacillin Sodium RS in the Standard solution (mg/mL)

C_u = concentration of Cloxacillin Sodium in the Sample solution (mg/mL)

P = potency of cloxacillin in USP Cloxacillin Sodium RS (µg/mg)

F₁ = conversion factor, 0.001 mg/µg

F₂ = relative response factor (see Table 3)

Acceptance criteria: See Table 3. The reporting threshold is 0.05%.

Table 3

a 6-Aminopenicillanic acid; (2S,5R,6R)-6-Amino-3,3-dimethyl-7-oxo-4-thia-1-azabicyclo[3.2.0]heptane-2-carboxylic acid.

b (4S)-2-(Carboxy[3-(2-chlorophenyl)-5-methylisoxazole-4-carboxamido]methyl)-5,5-dimethylthiazolidine-4-carboxylic acid.

C (4S)-2-([3-(2-Chlorophenyl)-5-methylisoxazole-4-carboxamido]methyl)-5,5-dimethylthiazolidine-4-carboxylic acid.

d The system resolves two isomers. The limit is for the sum of the isomers.

e 3-(2-Chlorophenyl)-5-methylisoxazole-4-carboxylic acid.

f (2R,5R,6R)-6-[3-(2-Chlorophenyl)-5-methylisoxazole-4-carboxamido]-3,3-dimethyl-7-oxo-4-thia-1-azabicyclo[3.2.0]heptane-2-carbothioic S-acid.

g (2S,5R,6R)-6-((2S,5R,6R)-6-[3-(2-Chlorophenyl)-5-methylisoxazole-4-carboxamido]-3,3-dimethyl-7-oxo-4-thia-1-azabicyclo[3.2.0]heptane-2-carboxamido)-3,3-dimethyl-7-oxo-4-thia-1-azabicyclo[3.2.0]heptane-2-carboxylic acid.

h (2S,5R,6R)-6-((R)-2-[(2R,4S)-4-Carboxy-5,5-dimethylthiazolidin-2-yl]-2-[3-(2-chlorophenyl)-5-methylisoxazole-4-carboxamido]acetamido)-3,3-dimethyl-7-oxo-4-thia-1-azabicyclo[3.2.0]heptane-2-carboxylic acid.

4.2 DIMETHYLANILINE (223): Meets the requirements

5 SPECIFIC TESTS

5.1 CRYSTALLINITY (695): Meets the requirements

5.2 PH (791).

Sample solution: 10 mg/mL in water

Acceptance criteria: 4.5-7.5

5.3 STERILITY TESTS (71)

Meets the requirements where the label states that Cloxacillin Sodium is sterile. If the test for Direct Inoculation of the Culture Medium is used, perform the procedure as directed in the chapter with the following exceptions. Use Fluid Thioglycollate Medium containing Polysorbate 80 solution (1 in 200) and an amount of sterile penicillinase sufficient to inactivate the cloxacillin in each tube. Use Soybean-Casein Digest Medium containing polysorbate 80 solution (1 in 200) and an amount of sterile penicillinase sufficient to inactivate the cloxacillin in each tube. Shake the tubes once daily.

5.4 WATER DETERMINATION, Method (921): 3.0%-5.0%

6 ADDITIONAL REQUIREMENTS

6.1 PACKAGING AND STORAGE:

Preserve in tight containers, and store at a temperature not exceeding 25°.

6.2 LABELING

Where it is intended for use in preparing sterile dosage forms, the label states that it is sterile or must be subjected to further processing during the preparation of sterile dosage forms.

6.3 USP REFERENCE STANDARDS (11)

USP Cloxacillin Related Compound D. RS

3-(2-Chlorophenyl)-5-methylisoxazole-4-carboxylic acid.

C₁₁H₈CINO₃  237.64

USP Cloxacillin Sodium RS