Thimerosal

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

Thimerosal contains NLT 97.0% and NMT 102.0% of thimerosal, calculated on the dried basis.

2 IDENTIFICATION

Change to read:

A. Spectroscopic Identification Tests 〈197〉, Infrared Spectroscopy: 197K (CN 1-May-2020)

B.

Sample solution: 10 mg/mL

Analysis: To the Sample solution add a few drops of silver nitrate TS.

Acceptance criteria: A pale yellow precipitate is formed.

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

3 ASSAY

Procedure

Solution A: 0.05% Trifluoroacetic acid, prepared by adding 1.0 mL of trifluoroacetic acid to 2 L of water

Mobile phase: Methanol and Solution A (60:40)

Standard stock solution: 250 μg/mL of USP Thimerosal RS in water

Impurity stock solution: 250 μg/mL of USP Thimerosal Related Compound A RS in methanol and water (90:10)

System suitability solution: 25 μg/mL each of USP Thimerosal RS and USP Thimerosal Related Compound A RS from Standard stock solution and Impurity stock solution, respectively, in water

Standard solution: 25 μg/mL of USP Thimerosal RS in water from Standard stock solution

Sample solution: 25 μg/mL of Thimerosal in water

[Note—Prepare both the Standard solution and Sample solution at a concentration of NMT ±10% of the specified concentration.]

Chromatographic system

(See Chromatography 〈621〉, System Suitability.)

Mode: LC

Detector: UV 222 nm

Column: 2.1-mm × 10-cm; 2-μm packing L1

Autosampler temperature: 4°

Flow rate: 0.35 mL/min

Injection volume: 2.5 μL

System suitability

Samples: System suitability solution and Standard solution

[Note—The relative retention times of thimerosal and thimerosal related compound A are 1.0 and 1.3, respectively.]

Suitability requirements

Resolution: NLT 3.5 between the thimerosal and thimerosal related compound A peaks, System suitability solution

Tailing factor: NMT 1.5, Standard solution

Relative standard deviation: NMT 0.73%, Standard solution

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of thimerosal in the portion of Thimerosal taken:

Result = (r_U/r_S) × (C_S/C_U) × 100

r_U = peak response from the Sample solution

r_S = peak response from the Standard solution

C_S = concentration of USP Thimerosal RS in the Standard solution (μg/mL)

C_U = concentration of Thimerosal in the Sample solution (μg/mL)

Acceptance criteria: 97.0%–102.0% on the dried basis

4 IMPURITIES

Mercury Ions

Iodide reagent: 332 mg/mL of potassium iodide. Prepare fresh daily. Keep the stopper in the flask, and protect from light.

Standard solution: 95 μg/mL of mercuric chloride

Sample stock solution: 5 mg/mL of Thimerosal

Sample solution A: 1 mg/mL of Thimerosal from Sample stock solution

Sample solution B: 1 mg/mL of Thimerosal from Sample stock solution and 9.5 μg/mL of mercuric chloride from Standard solution

Instrumental conditions

Mode: UV

Analytical wavelength: Determine the wavelength of maximum absorbance for the tetraiodomercurate ion at about 323 nm using the solution prepared by mixing 1.0 mL of the Standard solution and 5.0 mL of the Iodide reagent, and diluting with water to 10.0 mL.

Cell: 1 cm

Blank: Water

Analysis

Protect all solutions from light before determining their absorbances.

Samples: Label five 10-mL volumetric flasks C, D, E, F, and R. Transfer 5.0 mL of Sample solution A to flasks C and D, 5.0 mL of Sample solution B to flasks E and F, and 5.0 mL of water to

ask R. Dilute flasks C and E with water to volume. Dilute flasks D, F, and R with Iodide reagent to volume.

Determine the absorbances of the solutions in flasks C, D, E, F, and R as A , A , A , A , and A , respectively.

Calculate the percentage of mercury ions in the portion of Thimerosal taken:

Result = (A_U/A_S) × (C_S/C_U) × (A_r/M_r) × 100

A_U = absorbance of the Sample solution obtained by: A_U = A_D − A_R − A_C

A_S = absorbance of the Standard solution obtained by: A_S = A_F − A_R − A_E − A_U

C_S = concentration of mercuric chloride in Sample solution B (mg/mL)

C_U = concentration of Thimerosal in Sample solution B (mg/mL)

A_r = atomic weight of mercury, 200.59

M_r = molecular weight of mercuric chloride, 271.50

Acceptance criteria: NMT 0.70%

Organic Impurities

Solution A: 0.05% Trifluoroacetic acid, prepared by adding 1.0 mL of trifluoroacetic acid to 2 L of water

Mobile phase: Methanol and Solution A (60:40)

Stock solution 1: 250 μg/mL of USP Thimerosal RS in water

Stock solution 2: 250 μg/mL of USP Thimerosal Related Compound A RS in methanol and water (90:10)

System suitability solution: 25 μg/mL each of USP Thimerosal RS and USP Thimerosal Related Compound A RS from Stock solution 1 and

Stock solution 2, respectively, in water

Standard solution: 0.25 μg/mL of USP Thimerosal Related Compound A RS from Stock solution 2 in water

Sample solution: 250 μg/mL of Thimerosal in water

Chromatographic system

(See Chromatography 〈621〉, System Suitability.)

Mode: LC

Detector: UV 222 nm

Column: 2.1-mm × 10-cm; 2-μm packing L1

Autosampler temperature: 4°

Flow rate: 0.35 mL/min

Injection volume: 2.5 μL

System suitability

Samples: System suitability solution and Standard solution

Suitability requirements

Resolution: NLT 3.5 between the thimerosal and thimerosal related compound A peaks, System suitability solution

Relative standard deviation: NMT 3%, Standard solution

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of each impurity in the portion of Thimerosal taken:

Result = (r_U/r_S) × (C_S/C_U) × 100

r_U = peak response of each impurity from the Sample solution

r_S = peak response of thimerosal related compound A from the Standard solution

C_S = concentration of USP Thimerosal Related Compound A RS in the Standard solution (μg/mL)

C_U = concentration of Thimerosal in the Sample solution (μg/mL)

Acceptance criteria: See Table 1. Disregard peaks less than 0.05%.

Table 1

^a 2-Sulfanylbenzoic acid.

5 SPECIFIC TESTS

Loss on Drying 〈731〉

Analysis: Dry to constant weight under vacuum over phosphorus pentoxide.

Acceptance criteria: NMT 0.5%

6 ADDITIONAL REQUIREMENTS

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

USP Reference Standards 〈11〉

USP Thimerosal RS

USP Thimerosal Related Compound A RS

2,2'-Disulfanediyldibenzoic acid.