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Nitrosamine Ames Test (NDSRIs) | OECD 471

4 min de lecture

N-nitrosamines and NDSRIs have become one of the most scrutinised topics in pharmaceutical genotoxicity. To help sponsors meet tightening FDA and EMA expectations, GenEvolutioN — a GLP-accredited European genotoxicity CRO — now runs the OECD 471 Ames test under both standard and enhanced conditions, and links it to a full mutagenic-impurity strategy under ICH M7.

Why the standard Ames test falls short for nitrosamines

The OECD 471 Ames test is the most widely requested genotoxicity assay in regulatory frameworks, detecting DNA-reactive mutagens by measuring reverse mutations in Salmonella and E. coli strains. But standard guideline conditions show reduced sensitivity for some N-nitrosamines and NDSRIs — which is exactly why regulators now expect a sensitivity-optimised, two-step strategy rather than a single assay.

Enhanced Ames test conditions at GenEvolutioN

GenEvolutioN performs the OECD 471 Ames test with enhanced conditions designed to reliably detect the mutagenic potential of N-nitrosamines and NDSRIs.

Study format Single mutagenicity test
Tester strains TA98, TA100, TA1535, TA1537, WP2 uvrA pKM101
Method Pre-incubation, with extended pre-incubation time (30 minutes)
Metabolic activation 30% rat S9 and 30% hamster S9 (induced with phenobarbital and β-naphthoflavone)
Solvent Water or the lowest possible volume of organic solvent
Nitrosamine positive controls NDMA (N-nitrosodimethylamine), CPN (1-Cyclopentyl-4-nitrosopiperazine), DPN (N-nitroso-di-N-propylamine)
Historical control data Separate historical ranges for 30% rat S9 and 30% hamster S9, using the same pre-incubation format and acceptance criteria — including mean revertant counts, SD, min/max values, number of experiments and period covered

The standard GLP OECD 471 Ames test

For routine regulatory submissions, GenEvolutioN runs the full standard OECD 471 panel to GLP.

Strains TA98, TA100, TA1535, TA1537 and TA102
Metabolic activation Induced rat liver S9 (other sources on request)
Test format Plate incorporation and pre-incubation
Test item requirement 1 g
Analyse de la formulation Available on request
Endpoint Mutagenicity

From assay to regulatory dossier: an integrated mutagenic-impurity strategy

What sets GenEvolutioN apart is the link between the assay and the dossier. Beyond the Ames test itself, our analytical chemistry team supports mutagenic impurity assessment under ICH M7 — including identification and quantification of N-nitrosamine impurities — so that genotoxicity data and analytical evidence are built into a single, submission-ready package. This integrated approach reduces back-and-forth and shortens the path to regulatory acceptance.

Why GenEvolutioN

Key figures

50+

years of scientific heritage

5

genotoxicity assays under one roof

GLP

accredited regulatory testing

Highlights

GLP accreditation with expertise from former Sanofi and Covance preclinical centres
Genotoxicity suite covering OECD 471 (Ames), OECD 487 (MN), OECD 490 (MLA), OECD 473 (CA) and the Comet assay
Ames + analytical chemistry combined into one submission-ready package under ICH M7
Strict scientific validation process on every study

Foire aux questions

Nitrosamines & enhanced Ames — common questions

What is an NDSRI?
A nitrosamine drug substance-related impurity (NDSRI) is a nitrosamine formed from the active substance itself, rather than introduced from an external source. Because of their structure, several NDSRIs are potent DNA-reactive mutagens and require sensitivity-optimised testing.
Why is a second Ames test needed for nitrosamines?
Standard OECD 471 conditions can under-detect some nitrosamines. FDA and EMA recommend confirming a negative standard result with a second Ames test run under enhanced conditions before concluding that a substance is non-mutagenic.
What makes the enhanced Ames conditions different?
Enhanced conditions use 30% rat and 30% hamster S9 (phenobarbital and β-naphthoflavone induced), an extended 30-minute pre-incubation, and nitrosamine-specific positive controls (NDMA, CPN, DPN), supported by dedicated historical control data.
Does GenEvolutioN support ICH M7 mutagenic impurity assessment?
Yes. Our analytical chemistry team handles identification and quantification of N-nitrosamine impurities, so genotoxicity and analytical data are delivered together in a single, submission-ready package.

Discuss your nitrosamine / NDSRI testing strategy → Book a call