Improved quantitation of eight nitrosamine impurities in metformin drug products using a TSQ Fortis Plus mass spectrometer

Importance of the Topic

Trace nitrosamine impurities in pharmaceuticals have emerged as a critical safety concern due to their potent genotoxic and carcinogenic properties. Regulatory agencies worldwide now require strict control of these contaminants. Metformin, one of the most widely used diabetes treatments, must be tested for multiple nitrosamines at levels well below acceptable intake limits to ensure patient safety.

Objectives and Study Overview

This study demonstrates a single, fit-for-purpose LC-SRM-MS method for simultaneous quantitation of eight nitrosamines in metformin drug products. The workflow couples a Thermo Scientific Vanquish Core UHPLC system with a Hypersil GOLD C18 column to a TSQ Fortis Plus triple-quadrupole mass spectrometer, meeting FDA guidelines for sensitivity and selectivity.

Methodology and Instrumentation

Reference standards and isotopically labeled analogues were prepared at concentrations from 0.2 to 100 ng/mL. Metformin tablets were extracted and spiked following established protocols. Chromatography employed a 150 × 4.6 mm, 3 µm Hypersil GOLD C18 column at 20 °C with a 30-minute water/methanol gradient (0.1% formic acid). The TSQ Fortis Plus MS operated in either HESI or APCI mode with optimized SRM transitions and a segmented quadrupole AIM+ collision cell for rapid polarity switching.

Instrumentation Used

• Thermo Scientific Vanquish Core UHPLC system (Binary Pump, Split Sampler CT, Column Compartment)
• Hypersil GOLD C18 column, 150 × 4.6 mm, 3 µm
• Thermo Scientific TSQ Fortis Plus triple-quadrupole mass spectrometer with AIM+ technology
• Chromeleon CDS 7.2.10 for compliant data acquisition and processing

Main Results and Discussion

The method achieved baseline separation of NDMA from DMF and resolved seven additional nitrosamines in a single run. Instrument LODs ranged from 0.2 to 1.0 ng/mL (2–10 ppb) and LOQs from 0.5 to 2.0 ng/mL (5–20 ppb) in HESI mode; similar or improved performance was observed in APCI. Calibration curves were linear (r² > 0.997) up to 1000 ppb. Spiked metformin samples at 20 ppb showed accuracy within 85–115% and precision (%RSD) below 15%. Compared to the previous TSQ Fortis platform, the Fortis Plus system delivered up to 4× higher peak areas and enhanced confidence in qualifier transitions.

Benefits and Practical Applications

• Single-run quantitation of eight nitrosamines, reducing analysis time.
• Regulatory compliance with FDA and EMA guidelines for genotoxic impurities.
• Robust performance suitable for routine QC/QA testing in pharmaceutical laboratories.
• Flexible ionization modes (HESI/APCI) for optimal sensitivity across analytes.

Future Trends and Applications

Emerging directions include expansion of the method to other drug classes, miniaturization of sample preparation workflows, and integration with automated data-handling platforms. Continued improvements in triple-quadrupole hardware and software will further lower detection limits and simplify method transfer across laboratories.

Conclusion

This application note presents a robust, compliance-ready LC-SRM-MS method on the Thermo Scientific Vanquish Core and TSQ Fortis Plus platform for sensitive quantitation of nitrosamine impurities in metformin. The workflow meets stringent regulatory limits, offers high throughput, and supports confident, routine monitoring of genotoxic residues in drug products.

References

1. U.S. FDA. Information about Nitrosamine Impurities in Medications. FDA; 2020.
2. European Medicines Agency. Nitrosamine impurities in human medicinal products: Assessment report EMA/369136/2020.
3. U.S. FDA. Control of Nitrosamine Impurities in Human Drugs. Guidance for Industry U.S. FDA-2020-D-1530.
4. U.S. FDA. LC-HRMS Method for Determination of NDMA in Ranitidine. FDA FY19-177-DPA-S.
5. U.S. FDA. LC-HRMS Method for Detection of Six Nitrosamines in ARBs. FDA FY19-107-DPA-S.
6. U.S. FDA. LC-HRMS Method for Nitrosamines in Metformin. FDA FY20-106-DPA-S.
7. U.S. FDA. LC-MS/MS Method for NDMA in Ranitidine Drug Substance and Product. FDA FY20-006-DPA-S.
8. Thermo Fisher Scientific. Application Note AN000239: Quantitation of Nitrosamine Impurities in Metformin.