Clioquinol 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 

Clioquinol Ointment contains NLT 90.0% and NMT 110.0% of the labeled amount of clioquinol (C₉H₅ClINO) in a suitable ointment base. 

2 IDENTIFICATION 

A. 

Standard solution: Prepare as directed for the Standard solution in the Assay, except use 1.0 mL of pyridine instead of the Internal standard solution. 

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

B. 

Sample solution: Place nominally 25 mg of clioquinol from Ointment in a 100-mL volumetric flask, add 75 mL of dilute hydrochloric acid (1 in 4), and heat on a steam bath to melt the Ointment, shaking vigorously to extract the clioquinol. Cool under running water, and add dilute hydrochloric acid (1 in 4) to volume. Filter through paper, and dilute 3 mL of the filtrate with dilute hydrochloric acid (1 in 4) to 100 mL. Acceptance criteria: The UV absorption spectrum of the Sample solution exhibits maxima and minima at the same wavelengths as that of a similar solution of USP Clioquinol RS, concomitantly measured. 

3 ASSAY 

Procedure 

Internal standard solution: 2 mg/mL of pyrene in pyridine 

Standard stock solution: 3 mg/mL of USP Clioquinol RS in a mixture of pyridine and n-hexane (4:1) 

Standard solution: Transfer 1.0 mL of the Standard stock solution to a screw-capped glass vial fitted with a septum, add 1.0 mL of bis(trimethylsilyl)acetamide and 1.0 mL of Internal standard solution, and attach the cap. Heat in a water bath at 50° for 15 min, and then cool to ambient temperature. 

Sample solution: Transfer nominally 150 mg of clioquinol from Ointment to a 125-mL separator. Add 75 mL of n-hexane, then add 15 mL of dimethylformamide, and mix for 1 min. Allow the layers to separate, and transfer the lower layer to a 50-mL volumetric flask. Repeat the extraction with separate 15- and 10-mL portions of dimethylformamide, and transfer the lower layers to the 50-mL volumetric flask. Dilute with dimethylformamide to volume. Transfer 1.0 mL of this solution to a screw-capped glass vial fitted with a septum, and evaporate at 60° under a stream of nitrogen to dryness. Add 1.0 mL of a mixture of pyridine and n-hexane (4:1) to the residue, add 1.0 mL of bis(trimethylsilyl)acetamide and 1.0 mL of Internal standard solution, and attach the cap. Heat in a water bath at 50° for 15 min, then cool to ambient temperature. 

Chromatographic system 

(See Chromatography 〈621〉, System Suitability.) 

Mode: GC 

Detector: Flame ionization 

Column: 1.83-m × 2-mm glass; packed with 3% liquid phase G3 on 80- to 100-mesh support S1AB 

Temperatures 

Column: The initial temperature is 200° for a conditioning period of NLT 16 h (not connected to the detector) and is then reduced to 165°. Injection port: 170° 

Detector: 250° 

Carrier gas: Helium 

Flow rate: 30 mL/min for helium. Hydrogen and air are introduced into the detector at rates of 25 and 500 mL/min, respectively. Injection volume: 1 µL 

System suitability 

Sample: Standard solution 

[Note—The relative retention times for clioquinol and pyrene are 0.6 and 1.0, respectively.] 

Suitability requirements 

Resolution: NLT 3 between the clioquinol and the internal standard peaks 

Analysis 

Samples: Standard solution and Sample solution 

Calculate the percentage of the labeled amount of clioquinol (C₉H₅ClINO) in the portion of Ointment taken: 

Result = (R_U/R_S) × (C_S/C_U) × 100 

r_U = peak response ratio of clioquinol to the internal standard from the Sample solution

r_S = peak response ratio of clioquinol to the internal standard from the Standard solution

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

C_U = nominal concentrationof clioquinol in the Sample solution (mg/mL)

Acceptance criteria: 90.0%–110.0% 

4 ADDITIONAL REQUIREMENTS 

Packaging and Storage: Preserve in collapsible tubes or tight, light-resistant containers. 

USP Reference Standards 〈11〉 

USP Clioquinol RS