In September 2024, the FDA released Revision 2 of their guidance document titled “Control of Nitrosamine Impurities in Human Drugs Guidance for Industry.” This updated guidance represents a substantial expansion from the previous version, particularly in its approach to detecting and mitigating nitrosamines and their precursors. The revision introduces new directives for API manufacturing, which are applicable to all drug substance and component syntheses.

Water used in API manufacturing can vary in purity based on its intended use and the level of subsequent purification. Non-compendial waters are often utilized, with potable water being the minimum standard. Historically, API and drug substance manufacturers have not tested process water for nitrosamine impurities or their precursors. However, the new guidance makes it clear that testing process water for these impurities is now expected, as outlined in the updated document as follows:

API manufacturers should be aware that potable water used in API manufacture may contain low levels of nitrites and even nitrosamines from environmental sources. The existence of nitrites in processing water may lead to nitrosamine formation in API manufacture. Therefore, to avoid unacceptable levels of nitrosamine impurities in APIs, API manufacturers should analyze nitrite and nitrosamine levels in water and use water that has been purified to remove unacceptable impurities.

This language appears to be consistent with recent guidance from World Health Organization that we recently reviewed here:

Nitrosamine Testing Requirements in Pharmaceutical Waters

Potable water is more likely to contain nitrosating impurities, such as nitrite, rather than nitrosamines themselves. Therefore, monitoring for these nitrosamine precursors is a logical step. If nitrosamine precursors are detected, it is important to implement a treatment regimen specifically designed to reduce their levels in potable water, even if the water meets established drinking water standards (e.g., EPA National Primary Standards or WHO potable water standards).

MONITORING SUGGESTIONS

In pharmaceutical water treatment systems, which often involve multiple treatment stages, a sampling program for nitrosamines and their precursors should extend beyond just the final product water. At a minimum, the source water for these systems should be tested regularly. If nitrosamine precursors are detected, additional treatment methods may be necessary to specifically remove nitrogenous compounds. As previously mentioned, risk assessments should be conducted to identify and mitigate potential contamination sources, particularly from ion-exchange systems.

For API synthesis using potable water, a more comprehensive sampling program is recommended. Similar to the testing of other critical quality attributes, a thorough sampling regimen during Process Performance Qualification is expected. A suggested sampling matrix might include the following elements:

Impurity	Sampling Location	Suggested Early Phase PQ Testing	Ongoing Testing
Nitrate / Nitrite	Feed water	Daily	Quarterly
	Downstream of DI or ion-exchange processes	Daily	Quarterly
	Product water	Daily	Weekly
Chloramine	Feed water	Daily	Daily  (for microbial control purposes)
	Downstream of chlorination step	Daily	Continuous on-line
N-nitrosodimethylamine (NDMA) or indicator compound	Feed water	Daily	Quarterly
	Product water	Daily	Quarterly

A lack of process control or variability in feed water sources may necessitate more frequent testing. Conversely, if a robust and validated water treatment process is in place, sampling frequencies could be reduced. Testing for every nitrosamine compound may be impractical; instead, if no specific nitrosamine impurity has been identified, examining a suggested “indicator compound” could be useful.

Industry and regulatory organizations have recommended Acceptable Intake (AI) levels for nitrosamines based on daily exposure for final drug products. However, no specific maximum acceptable concentration for nitrosamines is established for raw materials or ingredients such as water. Target concentrations can vary depending on the application, but an Action Level of 10 ppb for nitrosamines in process waters is a reasonable benchmark. For APIs, acceptable concentrations of nitrosamines and their precursors may be higher, depending on the extent of downstream purification.

Regardless of the sampling frequency for nitrosamines and related compounds in pharmaceutical waters, it is crucial to recognize that water can be a potential source of contamination. Therefore, a risk analysis should be conducted to determine whether routine monitoring is necessary.