Determination of Nitrosamine Impurities Using the Agilent 6475 Triple Quadrupole LC/MS System

Significance of the Topic

Nitrosamine impurities are trace by-products in the manufacturing of pharmaceutical drugs, classified as potential genotoxic and probable carcinogens. Regulatory agencies mandate sensitive detection in final drug products to ensure patient safety and compliance.

Objectives and Study Overview

This study assesses the performance of the Agilent 6475 triple quadrupole LC/MS system coupled with the 1290 Infinity II Bio LC and APCI source for quantifying eight nitrosamine compounds at ultra-low levels. The targeted analytes include NDMA, NDEA, NMOR, NMEA, NPYR, NPIP, NDPA, and NDBA.

Methodology and Instrumentation

• Chromatography: Agilent 1290 Infinity II Bio LC with Poroshell 120 EC-C18 column (3×150 mm, 2.7 µm), gradient elution with water and methanol containing 0.1% formic acid, flow rate 0.5 mL/min, column temperature 40 °C.
• Mass Spectrometry: Agilent 6475 triple quadrupole MS with APCI in positive mode; MRM acquisition with optimized dwell times, fragmentor voltages, and collision energies for each analyte.
• Sample Preparation: Standards spiked in solvent (methanol:water) and losartan potassium extract; API extracted by sonication, centrifugation, and dilution; calibration from 0.0125 to 10 ng/mL in solvent and 0.05 to 1 ng/mL in drug matrix.

Main Results and Discussion

• Chromatographic separation achieved sharp peak shapes for all eight nitrosamines.
• Lower limit of quantitation (LLOQ) ≤ 0.025 ng/mL with S/N >10, RSD <20%, and accuracy within 80–120%.
• Calibration curves demonstrated linearity (R² > 0.99) across the tested range.
• Precision and accuracy met regulatory criteria at both solvent and API matrix levels, including 50–100 ppt in losartan extract.
• Sensitivity levels were well below acceptable nitrosamine thresholds set by regulatory guidelines.

Benefits and Practical Applications

The validated method offers robust, sensitive, and reproducible quantification of nitrosamine impurities, supporting quality control in pharmaceutical production. It can be adapted to various angiotensin receptor blocker (ARB) products with minimal adjustments.

Future Trends and Applications

Advancements may include integration with high-resolution MS, automated sample preparation, expansion to broader impurity panels, and alignment with evolving regulatory standards. The approach can be extended to other drug matrices and higher-throughput workflows.

Conclusion

The Agilent 6475 LC/TQ system combined with the 1290 Infinity II Bio LC and APCI source provides a sensitive and reliable platform for nitrosamine impurity analysis at trace levels, ensuring compliance and safeguarding drug safety.

References

1. U.S. Food and Drug Administration. FDA Updates and Press Announcements on Angiotensin II Receptor Blocker Recalls; March 2023.
2. Center for Drug Evaluation and Research, U.S. FDA. Control of Nitrosamine Impurities in Human Drugs: Guidance for Industry; February 2021.
3. European Medicines Agency, Committee for Medicinal Products for Human Use. Nitrosamine Impurities in Human Medicinal Products; June 2020.