Quantitation of 6 Nitrosamines in Losartan API by LC-MS/MS system as per the proposed USP General Chapter <1469>

Significance of the Topic

The presence of nitrosamine impurities in pharmaceutical products has emerged as a critical safety concern due to their potent mutagenic and carcinogenic properties. Regulatory agencies worldwide, including the US FDA and USP, have established stringent intake limits for nitrosamines in drug substances to protect patient health. Developing robust analytical methods capable of detecting multiple nitrosamine species at trace levels is essential for ensuring the safety and compliance of active pharmaceutical ingredients (APIs) such as Losartan.

Objectives and Overview of the Study

This application study aimed to implement and validate an LC-MS/MS method for simultaneous quantification of six nitrosamines (NDMA, NMBA, NDEA, NEIPA, NDIPA, NDBA) in Losartan API in accordance with the proposed USP General Chapter <1469> (Procedure 3). The method performance was assessed on the Shimadzu LCMS-8045 triple quadrupole system coupled with a Nexera™ XS UHPLC platform.

Methodology and Instrumentation

The analytical workflow comprised sample preparation, chromatographic separation, and tandem mass spectrometric detection:

• Sample Preparation: 80 mg Losartan API was spiked with internal standards, vortexed, centrifuged, and filtered.
• Chromatography: Raptor™ ARC-18 column (150 × 3.0 mm, 2.7 µm) with gradient elution using 0.1 % formic acid in water (A) and methanol (B) at 0.5 mL/min.
• Detection: APCI interface in positive MRM mode monitored two transitions per analyte with deuterated analogues as internal standards.
• Calibration: Nine-point curves ranging from low-ppb to tens of ppb concentrations, weighted linear regression (1/C).

Main Results and Discussion

Calibration curves for all six nitrosamines showed excellent linearity (R² > 0.997). Limits of quantitation (LOQs) ranged from 0.66 to 15.7 ppb, corresponding to signal-to-noise ratios above 10 and relative standard deviations (n=6) below 25 %. Recovery experiments at LOQ levels in spiked Losartan API yielded 70–130 % recovery, meeting USP criteria. Representative chromatograms demonstrated clear separation of nitrosamines with minimal interference from the API matrix.

Benefits and Practical Applications

This validated LC-MS/MS method offers:

• High sensitivity and specificity for six regulated nitrosamines.
• Compliance with proposed USP <1469> requirements for Losartan API.
• Efficient sample throughput using a fast UHPLC gradient and rapid polarity switching.
• Reliable quantitation supporting routine QA/QC and regulatory submissions.

Future Trends and Possibilities of Use

Advances in high-resolution MS and automated sample handling will further enhance nitrosamine screening capabilities. Future developments may include:

• Extension to additional drug classes and metabolites.
• Integration with online sample cleanup (e.g., SPE-LC-MS/MS).
• Adoption of high-throughput platforms for batch screening.
• Application of data-driven approaches for predictive impurity profiling.

Conclusion

A robust LC-MS/MS method on the Shimadzu Nexera™ XS-LCMS-8045 system has been demonstrated for reliable quantification of six nitrosamines in Losartan API. The approach satisfies stringent regulatory limits with high sensitivity, reproducibility, and throughput, making it well suited for pharmaceutical quality control.