Validation of an improved ion chromatography method for the limit of choline test in the USP Succinylcholine Chloride monograph

Importance of Topic

Succinylcholine chloride is an FDA-approved intravenous skeletal muscle relaxant used in short procedures such as intubation and convulsive therapy.
Monitoring residual choline in succinylcholine chloride is critical for patient safety and regulatory compliance.
Existing USP ion chromatography methods exhibit baseline drift and retention time loss after several hours due to strong binding of succinylcholine to the column.

Study Objectives and Overview

The goal was to develop and validate an improved ion chromatography method for the USP limit of choline test in succinylcholine chloride.
The study followed ICH Q2A/Q2B and USP <1225> and <621> guidelines for method validation.
An enhanced elution strategy was introduced to address baseline drift and retention issues observed in the original monograph method.

Methodology and Instrumentation

An ICS-5000+ HPIC system with a Dionex IonPac CS19 analytical column and CG19 guard column was used.
Suppressed conductivity detection employed a CERS 500 electrolytically regenerated suppressor in recycle mode.
Methanesulfonic acid (MSA) was generated electrolytically (EGC 500) and applied in a stepped gradient: 6 mM isocratic hold, ramp to 50 mM for wash, and return to 6 mM for re-equilibration.
The improved method adds an 18-minute 50 mM MSA wash per injection to elute succinylcholine around 26 minutes, yielding a total run time of 43 minutes.
Samples and standards were prepared in high-density polyethylene containers to avoid adsorption, with choline chloride calibration solutions from 0.2 to 50 µg/mL.

Main Results and Discussion

The original method showed baseline rise and choline retention loss after ~6–7 hours.
The improved method maintained stable choline retention (~10 minutes) and prevented baseline drift over extended sequences.
Calibration was linear (0.2–50 µg/mL, r2 = 0.9998) with LOD of 0.06 µg/mL and LOQ of 0.2 µg/mL.
System suitability met USP criteria: choline peak area RSD <1%, retention time RSD <0.1%, and resolution from potassium >10.
Sample precision over three days (n=3 per day) yielded overall choline content of 0.11% in succinylcholine chloride with interday RSD 3.4%.
Spike recovery for choline levels from 0.04% to 0.3% ranged from 104% to 118%, demonstrating accuracy.
Robustness tests with ±10% variations in flow rate, eluent concentration, and column temperature showed <1.1% change in measured choline and <4% change in peak parameters.

Method Benefits and Practical Applications

The validated method offers reliable quantification of trace choline in succinylcholine chloride for QC and regulatory compliance.
It overcomes long-run stability issues, reducing reanalysis and downtime in pharmaceutical laboratories.
The approach can be implemented on any IC system supporting electrolytic or chemical suppression.

Future Trends and Potential Uses

This elution-based wash strategy may be adapted for other strongly retained drug impurities or quaternary ammonium compounds.
Integration with mass spectrometric detection could further enhance selectivity and sensitivity.
Automation of gradient generation and data processing will streamline high-throughput analysis in industrial settings.

Conclusion

The improved ion chromatography method meets USP and ICH validation requirements, eliminating baseline drift and retention instability.
It provides precise, accurate, and robust measurement of choline in succinylcholine chloride and is recommended for USP monograph revision.

Reference

1. Hospira, Inc. Quelicin (Succinylcholine Chloride Injection, USP) Label, 2010.
2. Sandoz Inc. Anectine (Succinylcholine Chloride Injection, USP) Label, 2010.
3. The United States Pharmacopeia 40 National Formulary 35 (2018), Succinylcholine Chloride Monograph.
4. Pharmacopeia Forum 44(2) (2018), Proposed Revision for Limit of Choline Test.
5. ICH Q2A, Validation of Analytical Procedures: Definition and Terminology, 1995.
6. ICH Q2B, Validation of Analytical Procedures: Methodology, 1996.
7. USP <1225> Validation of Compendial Methods, 2017.
8. USP <621> Chromatography, 2017.