In this SelectScience webinar available on demand, Dr. Mark Jordi, President of Jordi Labs, explores the regulations covering mutagenic impurities and reviews relevant analytical methodologies for their characterization.
Analysis of mutagenic impurities in pharmaceutical material is challenging due to the very high toxicity of these compounds which requires very low detection limits for quantitative methods (parts per billion). This also places a much higher burden on the analytical screening methods required to identify unexpected or unknown mutagenic impurities in pharmaceutical products, as screening at low levels requires high instrument sensitivity and low background noise. In addition, limitations on the commercial availability of impurity standards can require alternative quantitation strategies including relative quantitation versus surrogate standards. This has the potential to adversely affect quantitative accuracy.
Read on for highlights from the webinar Q&A session or register to watch the webinar on demand.
Q: What kinds of materials are most likely to contain mutagenic impurities?
MJ: High potency mutagenic impurities, particularly nitroso and azoxy compounds, are most commonly found in things that contain secondary amines because the process by which they're created is often due to a reduction of that amine group. With other members of the cohort of concern, for example aflatoxin-like species, they are most often due to exposure to a fungus and thus they are not as likely to be found in a pharmaceutical material.
Q: How can we confirm the identification of an impurity if we cannot obtain enough for NMR or FTIR confirmation?
MJ: It's very difficult when you can only obtain a trace level amount of a material to confirm its identification. Typically, the tools you are going to rely on most are HPLC and mass spectroscopy. You may have to resort to synthesis of a standard that you think is the correct structure and then confirm a retention time match. In the cases where you can’t readily collect fractions of the material by HPLC, one other option would be to begin to concentrate your sample, re-analyze it, confirm that it neither degrades nor volatilizes, and then continue to concentrate as needed until you can get a more sufficient amount of material. These are some of the strategies we use when identifying a very low-level impurity.
Q: How do you estimate limits of detection when the noise in a high-resolution mass spec is low to none?
MJ: Signal-to-noise does not work in those cases. There is an alternative strategy that has been put out by the EPA that involves the use of statistical analysis of the signal to define the method detection limit. This is done by analyzing a number of replicate injections of a standard near the method detection limit and determining the standard deviation. The associated Student’s t-value for the 99th percentile confidence interval is then multiplied by the standard deviation to obtain the method detection limit. This statistical approach is preferred in cases where the instrument effectively has no noise and thus the signal-to-noise ratio cannot be calculated.
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