Pyridostigmine Bromide

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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C₉H₁₃BrN₂O₂ 261.12 

Pyridinium, 3-[[(dimethylamino)carbonyl]oxy]-1-methyl-, bromide; 

3-Hydroxy-1-methylpyridinium bromide dimethylcarbamate CAS RN®: 101-26-8; UNII: KVI301NA53. 

1 DEFINITION 

Pyridostigmine Bromide contains NLT 98.0% and NMT 102.0% of pyridostigmine bromide (C₉H₁₃BrN₂O₂), calculated on the dried basis.  

2 IDENTIFICATION 

Change to read: 

A. Spectroscopic Identification Tests 〈197〉, Infrared Spectroscopy: 197A or 197K

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

C. Identification Tests—General 〈191〉, Chemical Identification Tests, Bromide 

Sample solution: 20 mg/mL of Pyridostigmine Bromide 

Acceptance criteria: Meets the requirements 

3 ASSAY 

Procedure 

Buffer: 4.3 g/L of sodium dodecyl sulfate in water. Adjust with phosphoric acid to a pH of 2.0. 

Mobile phase: Acetonitrile and Buffer (32:68) 

System suitability solution: 2 µg/mL each of USP Pyridostigmine Bromide RS, USP Pyridostigmine Related Compound A RS, and USP Pyridostigmine Related Compound B RS in Mobile phase 

Standard solution: 0.3 mg/mL of USP Pyridostigmine Bromide RS in Mobile phase 

Sample solution: 0.3 mg/mL of Pyridostigmine Bromide in Mobile phase 

Chromatographic system 

(See Chromatography 〈621〉, System Suitability.) 

Mode: LC 

Detector: UV 220 nm. For Identification B, use a diode array detector in the range of 200–400 nm. 

Column: 4.6-mm × 15-cm; 5-µm packing L1 

Flow rate: 1 mL/min 

Injection volume: 20 µL 

Run time: NLT 2.6 times the retention time of pyridostigmine 

System suitability 

Samples: System suitability solution and Standard solution 

Suitability requirements 

Resolution: NLT 1.5 between pyridostigmine related compound A and pyridostigmine, System suitability solution Tailing factor: NMT 2.0, Standard solution 

Relative standard deviation: NMT 0.73%, Standard solution 

Analysis 

Samples: Standard solution and Sample solution 

Calculate the percentage of pyridostigmine bromide (C₉H₁₃BrN₂O₂) in the portion of Pyridostigmine Bromide taken:  

r_U = peak response from the Sample solution 

r_S = peak response from Standard solution  

C_S = concentration of USP Pyridostigmine Bromide RS in Standard solution (mg/mL) 

C_U = concentration of Pyridostigmine Bromide in the Sample solution (mg/mL)

Acceptance criteria: 98.0%–102.0% on the dried basis 

4 IMPURITIES 

Residue on Ignition 〈281〉: NMT 0.1% 

Organic Impurities 

Mobile phase and Chromatographic system: Proceed as directed in the Assay. 

Sensitivity solution: 0.2 µg/mL of USP Pyridostigmine Bromide RS in Mobile phase 

Standard solution: 0.6 µg/mL each of USP Pyridostigmine Bromide RS, USP Pyridostigmine Related Compound A RS, and USP Pyridostigmine Related Compound B RS in Mobile phase 

Sample solution: 1000 µg/mL of Pyridostigmine Bromide in Mobile phase 

System suitability 

Samples: Sensitivity solution and Standard solution 

Suitability requirements 

Resolution: NLT 1.5 between pyridostigmine related compound A and pyridostigmine, Standard solution 

Relative standard deviation: NMT 5.0% for all the three peaks, Standard solution 

Signal-to-noise ratio: NLT 10, Sensitivity solution 

Analysis 

Samples: Standard solution and Sample solution 

Calculate the percentage of pyridostigmine related compound A and pyridostigmine related compound B in the portion of Pyridostigmine Bromide taken: 

Result = (r_U/r_S) × (C_S/C_U) × 100 

r_U = peak response of pyridostigmine related compound B or any individual unspecified degradation product from the Sample solution 

r_S = peak response of pyridostigmine related compound A or pyridostigmine related compound B from the Standard solution

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

C_U = concentration of Pyridostigmine Bromide in the Sample solution (mg/mL) 

Calculate the percentage of each unspecified impurity in the portion of Pyridostigmine Bromide taken: 

r_U = peak response of each impurity from the Sample solution

r_S = peak response of pyridostigmine from the Standard solution

C_S = concentration of USP Pyridostigmine Bromide RS in Standard solution (mg/mL) 

C_U = concentration of Pyridostigmine Bromide in the Sample solution (mg/mL) 

Acceptance criteria: See Table 1. The reporting threshold is 0.02%. 

Table 1 

5 SPECIFIC TESTS 

Loss on Drying 〈731〉 

Analysis: Dry at 100° for 4 h in a suitable vacuum drying tube, using phosphorus pentoxide as the desiccant. 

Acceptance criteria: NMT 2.0% 

6 ADDITIONAL REQUIREMENTS 

Packaging and Storage: Preserve in tight containers. 

USP Reference Standards 〈11〉 

USP Pyridostigmine Bromide RS 

USP Pyridostigmine Related Compound A RS Pyridin-3-yl dimethylcarbamate. 

C₈H₁₀N₂O₂ 166.18 

USP Pyridostigmine Related Compound B RS 3-Hydroxy-1-methylpyridin-1-ium bromide. C6H8BrNO 190.04