26/05/2023, 16:07 Rapid Quantitation of Four Nitrosamine Impurities in Angiotensin Receptor Blocker Drug Substances - ScienceDirect

Journal of Pharmaceutical Sciences

Volume 112, Issue 5, May 2023, Pages 1246-1254

Pharmaceutics, Drug Delivery and Pharmaceutical Technology

Rapid Quantitation of Four Nitrosamine Impurities in Angiotensin Receptor

Blocker Drug Substances
Jinhui Zhang, Susan Daniela Selaya, Diaa Shakleya, Adil Mohammad, Patrick J. Faustino

Show more

Share Cite

https://doi.org/10.1016/j.xphs.2022.12.005 ↗
Get rights and content ↗

Abstract

Starting in July 2018, the FDA alerted patients and health care professionals to the recall of ARBs such as valsartan
by several pharmaceutical companies because of their potential contamination with carcinogenic nitrosamine
impurities, including: (1) N-nitrosodimethylamine (NDMA), (2) N-nitrosodiethylamine (NDEA), (3) N-
nitrosoethylisopropylamine (NEIPA), (4) N-nitrosodiisopropylamine (NDIPA), (5) N-nitrosodibutylamine (NDBA) and
(6) N-nitroso-N-methyl-4-aminobutyric acid (NMBA). The FDA initiated a laboratory investigation to develop
analytical procedures to test multiple lots of marketed ARB drugs to determine the possible presence of
carcinogenic impurities and, if present, quantitate the levels of these impurities.

Here the FDA laboratory developed and validated an automated micro-solid phase extraction MS/MS method,
where all the analytes are not separated prior to elution to the MS, to simultaneously quantify NEIPA, NDIPA, NDBA
and NMBA in ARB drug substances with an instrument sample analysis time of 12 seconds. The method was
validated according to the ICH Q2(R1) guideline, and was determined to be specific, accurate, precise and linear
over the corresponding nitrosamine analytical ranges. The method has been successfully implemented to quantitate
the four nitrosamine impurities in 129 generic losartan, valsartan, olmesartan, irbesartan and telmisartan drug
substance samples from 32 lots; and 32 losartan and valsartan drug product samples from 6 lots.

Introduction

Angiotensin II receptor blocker (ARB) drugs, including valsartan, losartan, olmesartan, irbesartan and telmisartan
are commonly used to treat hypertension and cardiovascular diseases.1,2 On July 13th, 2018, the FDA announced a
voluntary recall of several valsartan drug products following detection of a nitrosamine impurity, N-
nitrosodimethylamine (NDMA).3 On September 13th, 2018, FDA reported the finding of an additional nitrosamine
impurity, N-nitrosodiethylamine (NDEA), in an already recalled valsartan product.3 On March 1st, 2019, FDA
announced the first ARB recall of a drug product, losartan, because of the presence of a third nitrosamine impurity,
N-nitroso-N-methyl-4-aminobutyric acid (NMBA).3 NDMA, NDEA and NMBA are all classified as probable human
carcinogens.4 The presence of these nitrosamine impurities was unexpected and was later reported to be generated
during the manufacturing of the drug substances when amines such as dimethylamine reacted with a nitrosating
agent.5 Subsequently, the combination of sources of vulnerable amines and nitrosation conditions posed a risk for

https://www.sciencedirect.com/science/article/abs/pii/S0022354922005688 1/5
26/05/2023, 16:07 Rapid Quantitation of Four Nitrosamine Impurities in Angiotensin Receptor Blocker Drug Substances - ScienceDirect

the formation of nitrosamine impurities.5 Following additional risk assessment of commonly used manufacturing
processes and reagents, three other nitrosamine impurities might also be present in ARB drugs, namely N-ethyl-N-
nitroso-2-propanamine (NEIPA), N-nitroso-diisopropylamine (NDIPA), and N-nitrosodibutylamine (NDBA).3

The detection of the nitrosamine impurities in ARB drugs resulted in numerous recalls by manufacturers and some
ARB drug shortages were identified in the United States.3 In response to concerns about public health exposure to
nitrosamines impurities, FDA developed methods for determining nitrosamine impurities in ARB drug substance
and drug products. For example, headspace GC/MS methods to quantitate trace amounts (ng per mg, or parts-per-
million (ppm)) of NDMA were validated and publicly shared, Later, NDEA was added to the method when this
nitrosamine was found in valsartan drug substance and drug product.6,7 However, the newly developed GC/MS
methods did not detect NMBA, a nitrosamine impurity found in losartan from one manufacturer.8 Manufacturing
processes using sodium nitrite and N-methyl-2-pyrrolidone (NMP) during the synthesis of losartan drug substance
were discovered to have the potential to form NMBA. Given the limitations of gas chromatography for NMBA and
NDBA with low calculated vapor pressures9 relative to NDMA, NDEA, NEIPA and NDIPA and the fact that NMBA
contains a carboxyl group, an additional method which could screen and quantify the two additional nitrosamines
was desired.

Therefore, an automated micro-solid phase extraction MS/MS platform, where all the analytes are not separated
prior to elution to the MS, was strategically implemented to simultaneously quantify the four nitrosamines in drug
substance to provide rapid and accurate analytical data. A high throughput mass spectrometry system, Agilent
RapidFire™-MS/MS, that utilizes an automated trap and elute strategy to enable desalting and direct sample
transfer to a tandem mass spectrometer without a liquid chromatography interface was applied for this work.10 The
system employed a unique feature that allows for a typical turnaround time of only 2 to 20 seconds per analytical
run. FDA had already successfully implemented this platform for clinical studies, IVPT studies using complex
receptor media, in vitro drug-drug interaction, monitoring drug degradation such as cold chain product oxytocin, as
well as in-depth investigation of key parameters in pharmaceutical analysis and bioanalytical method development
and validation.11 In the work reported here, we describe the development and implementation of a method for the
simultaneous quantitation of four nitrosamine impurities NMBA/NDBA/NDIPA/NEIPA in ARB drug substance, with a
turnaround time less than 12 seconds per sample.12 (Fig. 1).

Section snippets

Reagents

Standards for four impurities, NEIPA, NDIPA, NDBA and NMBA, and three internal standards (IS) d4-NDEA, d18-
NDBA and d3-NMBA were purchased from Toronto Research Chemicals (North York, Canada). d3-NMBA was the IS
for NMBA, d18-NDBA was the IS for NDBA whereas d4-NDEA was the IS for NEIPA and NDIPA. Stock solutions of all
analytes and internal standards were prepared in methanol. Working solutions of all analytes and internal
standards were made by diluting stock solutions with 10% methanol in…

Results and Discussion

FDA laboratories developed, validated and implemented an analytical procedure using an automated micro-solid
phase extraction-MS/MS system, where all the analytes are not separated prior to elution to the MS, for the
simultaneously screening and qualifying of four nitrosamine impurities in ARB drug substances and drug products.
The method was shared publicly (https://www.fda.gov/media/125477/download ↗).12 Since there was previously no
orthogonal method for all four nitrosamine impurities of…

Conclusion

https://www.sciencedirect.com/science/article/abs/pii/S0022354922005688 2/5
26/05/2023, 16:07 Rapid Quantitation of Four Nitrosamine Impurities in Angiotensin Receptor Blocker Drug Substances - ScienceDirect

A high throughput automated micro-solid phase extraction MS/MS method, where all the analytes are not
separated prior to elution to the MS, for screening and qualifying four nitrosamine impurities in ARB drug
substances was developed and fully validated, to test for the possible presence of nitrosamine impurities in generic
losartan, valsartan, olmesartan, irbesartan, telmisartan drug substances and drug products. The method has been
published on the FDA public website (//www.fda.gov/media/125477/download ↗…

Acknowledgements

This work was supported by FDA's Valsartan Task Force with intramural funding provided by FDA's Office of Testing
and Research. The authors also wish to thank Dr. Jin Zhang, Office of Policy for Pharmaceutical Quality, Office of
Pharmaceutical Quality, FDA/CDER, for helpful discussions on the scope of the study.…

Disclosure of Potential Conflicts of Interest

None of the authors has any personal or financial conflict of interest or has entered into any agreement that could
potentially result in a conflict of interest due to their access to the data presented here, or on their ability to analyze
the data independently, or to prepare and publish future manuscripts, based wholly or partially on these data.…

References (17)

SG Chrysant
Angiotensin II receptor blockers in the treatment of the cardiovascular disease continuum
Clinic Therap (2008)

H Siragy
Angiotensin II receptor blockers: review of the binding characteristics
Am J Cardiol (1999)

DJ Snodin et al.
Short commentary on NDMA (N-nitrosodimethylamine) contamination of valsartan products
Regul Toxicol Pharmacol (2019)

RG Klein
Calculations and measurements on the volatility of N-nitrosamines and their aqueous solutions
Toxicology (1982)

FDA
FDA Updates and Press Announcements on Angiotensin II Receptor Blocker (ARB) Recalls (Valsartan,
Losartan, and Irbesartan)
(2019)

Control of Nitrosamine Impurities in Human Drugs: Guidance for Industry

(2021)

Combined Direct Injection N-Nitrosodimethylamine (NDMA), N-Nitrosodiethylamine (NDEA), N-

Nitrosoethylisopropylamine (NEIPA), N-Nitrosodiisopropylamine (NDIPA), and N-Nitrosodibutylamine
(NDBA) Impurity Assay by GC-MS/MS
(2019)

Combined Headspace N-Nitrosodimethylamine (NDMA), N-Nitrosodiethylamine (NDEA), N-

Nitrosoethylisopropylamine (NEIPA), and N-Nitrosodiisopropylamine (NDIPA) Impurity Assay by GC-
MS/MS

https://www.sciencedirect.com/science/article/abs/pii/S0022354922005688 3/5
26/05/2023, 16:07 Rapid Quantitation of Four Nitrosamine Impurities in Angiotensin Receptor Blocker Drug Substances - ScienceDirect

(2019)
There are more references available in the full text version of this article.

Cited by (0)

Recommended articles (6)

Research article

N-Nitrosamines Impurities in Pharmaceuticals The Abrupt Challenges that Resulted, the Evolving
Science, and the Regulatory Framework
Journal of Pharmaceutical Sciences, Volume 112, Issue 5, 2023, pp. 1161-1162

Research article

Ranitidine: A Proposed Mechanistic Rationale for NDMA Formation and a Potential Control Strategy
Journal of Pharmaceutical Sciences, Volume 112, Issue 5, 2023, pp. 1220-1224
Show abstract

Research article

European Pharmacopoeia Activities on Control of Nitrosamines and other DNA-Reactive Impurities

Journal of Pharmaceutical Sciences, Volume 112, Issue 5, 2023, pp. 1163-1165

Research article

Current Threat of Nitrosamines in Pharmaceuticals and Scientific Strategies for Risk Mitigation
Journal of Pharmaceutical Sciences, Volume 112, Issue 5, 2023, pp. 1192-1209

Show abstract

Research article

Nitrosated Active Pharmaceutical Ingredients – Lessons Learned?

Journal of Pharmaceutical Sciences, Volume 112, Issue 5, 2023, pp. 1210-1215
Show abstract

Research article

N-Nitrosamines Impurities in Pharmaceuticals the Abrupt Challenges They Bring and Approaches to
Tackle the Risk
Journal of Pharmaceutical Sciences, Volume 111, Issue 10, 2022, p. 2651

This scientific publication reflects the views of the authors and should not be construed to represent FDA's views or policies.

View full text

© 2022 Published by Elsevier Inc. on behalf of American Pharmacists Association.

https://www.sciencedirect.com/science/article/abs/pii/S0022354922005688 4/5
26/05/2023, 16:07 Rapid Quantitation of Four Nitrosamine Impurities in Angiotensin Receptor Blocker Drug Substances - ScienceDirect

Copyright © 2023 Elsevier B.V. or its licensors or contributors.

ScienceDirect® is a registered trademark of Elsevier B.V.

https://www.sciencedirect.com/science/article/abs/pii/S0022354922005688 5/5