Analysis of Cyanocobalamin tablets, assay procedure 2 as per USP41-NF36 monograph, using Shimadzu Nexera X2 Fast LC system

Importance of the Topic

Accurate quantification of cyanocobalamin in tablet formulations is essential for ensuring dosage efficacy and patient safety. Because vitamin B12 deficiencies can lead to severe health issues, reliable analytical methods are crucial in both prescription and over-the-counter products.

Objectives and Study Overview

This work aims to demonstrate the fast and accurate analysis of vitamin B12 in cyanocobalamin tablets according to the USP41–NF36 Monograph 1114, Procedure 2. Using the Shimadzu Nexera X2 Fast LC system, the study evaluates system suitability and method performance under USP guidelines.

Methodology

The assay follows the USP monograph for sample and standard preparation, chromatographic conditions, and system suitability checks. Key steps include:

• Buffer preparation with hexane sulfonic acid sodium salt and phosphoric acid adjusted to pH 3.5.
• Standard solution at 1 µg/mL cyanocobalamin from USP reference standard.
• Tablet powder extraction by sonication, vigorous shaking, and filtration through a 0.22 µm membrane.
• UHPLC separation on a 2.1 × 100 mm L1 column (1.7 µm) at 35 °C with a buffer/acetonitrile gradient.
• Detection by UV at 361 nm with 15 µL injection volume and 0.5 mL/min flow rate.

Instrumentation Used

The analysis employed the Shimadzu Nexera X2 Fast LC system featuring:

• High-pressure capability up to 130 MPa for UHPLC performance.
• Micro-volume plungers for precise solvent delivery and low carryover.
• Column oven and autosampler modules for temperature control and automation.
• UV detector set at 361 nm.

Key Results and Discussion

The retention time for cyanocobalamin in both standard and sample solutions was 6.38 min. System suitability results satisfy USP criteria:

• %RSD of retention time: 0.09–0.19 (NMT 2.0%).
• %RSD of peak area: 0.26–0.30 (NMT 2.0%).
• Tailing factor: 1.43–1.51 (NMT 2.0).
• Theoretical plates: 80,833.

Overlay chromatograms confirmed excellent reproducibility and peak shape consistency.

Benefits and Practical Applications

This UHPLC method offers rapid throughput, high resolution, and reliable quantitation suitable for routine quality control in pharmaceutical laboratories. The low injection volume and minimal carryover improve analytical efficiency and reduce solvent consumption.

Future Trends and Applications

Advancements in UHPLC technology will enable even faster analyses, greener solvent use, and integration with multidimensional separations. Automation and high-throughput workflows are expected to expand applications in pharmacopeial and industrial settings.

Conclusion

The study demonstrates that the Shimadzu Nexera X2 Fast LC system effectively meets USP41 Procedure 2 requirements for cyanocobalamin tablet analysis. All system suitability parameters fell well within the monograph limits, underscoring the method’s robustness and suitability for quality control.

References

1. USP Monograph, Cyanocobalamin Tablets, USP 41–NF36, Monograph 1114, Procedure 2.
2. USP General Chapter 621, USP41–NF36, First Supplement.