Highly Sensitive Quantification of Nitrosamines in Polymer-based Therapeutics - A Case Study of Sevelamer Hydrochloride 800mg Tablets

Importance of the Topic

Recent recalls of drug products due to nitrosamine impurities highlight the critical need for highly sensitive methods to detect trace levels of these genotoxic contaminants in pharmaceuticals.

Study Objectives and Overview

The study aims to develop and validate a high‐throughput LC‐MS/MS method capable of simultaneous quantification of six nitrosamines (NDMA, NDEA, NEIPA, NDIPA, NDPA, NDBA) in Sevelamer hydrochloride 800 mg tablets at sub‐ppb levels.

Methodology

• Sample preparation: Tablet crushing, weighing equivalent to 100 mg API, extraction with 95:5 water:methanol, sonication (30 min), centrifugation and filtration.
• Chromatography: Agilent 1290 Infinity II UHPLC system, Poroshell C18 column (4.6×150 mm, 2.7 μm), 0.5 mL/min flow, gradient elution with 0.1% formic acid in water and methanol over 21 min.
• Mass spectrometry: Agilent 6470 triple quadrupole LC/MS with APCI in positive mode, optimized MRM transitions for each nitrosamine.

Použitá instrumentace

• 1290 Infinity II pump (G7120A), multisampler (G7167B), multicolumn thermostat (G7116B), variable wavelength detector (G7114B)
• 6470 triple quadrupole LC/MS (G6470B) with APCI interface and optimized source parameters (Gas Temp 300 °C, Gas Flow 7 L/min, Nebulizer 25 psi, Capillary 4000 V, APCI heater 350 °C)

Main Results and Discussion

• Achieved baseline separation of six nitrosamines from polymer and excipient peaks in Sevelamer HCl matrix.
• Limits of quantification: 0.1 ng/mL for NDMA (0.005 ppm) and 0.04 ng/mL for other nitrosamines (0.002 ppm).
• Method reproducibility: %RSD ≤5.1 for MRM peak area at 0.6 ng/mL (0.03 ppm), demonstrating high precision.
• Recovery ranged from 80% to 106% at low-level spikes, within ±20% acceptance criteria.

Benefits and Practical Applications

• Meets stringent regulatory requirements (US FDA) for nitrosamine control in drug products.
• High sensitivity and selectivity allow reliable routine batch testing of Sevelamer tablets.
• Applicable to quality control in pharmaceutical manufacturing to ensure patient safety.

Future Trends and Potential Uses

• Extension of the method to other polymeric and small-molecule APIs vulnerable to nitrosamine formation.
• Integration with high-resolution MS for non-targeted impurity screening and broader contaminant profiling.
• Automation of sample preparation and data processing to increase laboratory throughput.

Conclusion

The developed UHPLC-MS/MS assay provides a robust, sensitive, and reproducible platform for the quantitative determination of six nitrosamine impurities in Sevelamer hydrochloride tablets, fulfilling regulatory guidelines and enabling comprehensive quality control.

Reference

1. FDA guidance document: LC-HRMS Method for Determination of Six Nitrosamine Impurities in ARB Drugs.
2. FDA guidance document: Control of Nitrosamine Impurities in Human Drugs.
3. Agilent Technologies application note 5994-1383EN, 2019: Determination of Nitrosamine Impurities Using Ultivo Triple Quadrupole LC/MS.
4. Agilent Technologies application note 5994-2286EN, 2020: Simultaneous Determination of Eight Nitrosamine Impurities in Metformin Using Agilent 6470 TQ LC/MS.