Establishing Impurity Analysis Methods For Complex Samples Using Risk Assessment Strategies

In the actual development of separation methods, especially for the separation and analysis of complex pharmaceutical impurities, the following problems are often encountered:

How many impurities are present in the drug?

What are these impurities?

Can these impurities all be detected in the HPLC separation system?

Where do they peak?

These issues can be briefly summarized as:

“how many”, “what”, “where”

We not only care about “how many” impurities, “what” impurities, but also want to know “where” impurities.

However, the current separation method development and evaluation methods are difficult to answer these questions. In view of this gap, the following will correlate the HPLC separation method with the knowledge of the impurity profile. Based on the principle of risk analysis, the evaluation of HPLC analysis of complex samples is proposed. The new strategy of the system – the “four-quadrant” strategy.

By answering two questions:

1 Is the impurity known or unknown?

2 Are the impurities detected in the HPLC separation system?

The impurity analysis results are divided into four parts for consideration (as shown above).

The first quadrant represents: Detected known impurities

The second quadrant represents: Unknown impurities detected

The third quadrant represents: Undetected unknown impurities

The fourth quadrant represents: Undetected known impurities

The complexity of the analytical object determines the necessity of optimizing the HPLC separation conditions.

The “four-quadrant” strategy requires the optimization of existing separation conditions, the expansion of the first and second quadrants, and the evaluation of the method’s ability to detect impurities. Impurities in the four quadrants have different meanings and require different analytical strategies to gain insight into the relationship between analytical methods and impurity profiles.

The first quadrant characterizes the ability of the method to detect recognized impurities.

Identify the recognized impurities in the chromatogram using various information such as UV spectrum, mass spectrometry fragmentation, chromatographic retention rule, and degradation rule.

The second quadrant characterizes the analytical method’s ability to detect unknown impurities.

The structure of unknown impurities can be inferred from various information such as mass spectrometry fragmentation, degradation law, chromatographic retention law, and ultraviolet spectrum, which can gradually deepen the understanding of drug impurity spectra. In addition, theoretically, people have cognitive blind spots, where impurities are unknown and not detected by the method, which is the case of the third quadrant characterization.

Unknown impurities characterized in the third quadrant include new impurities that are not currently present in the sample, as well as new impurities that are objectively present in the sample but have never been discovered.

The former reflects people’s cognitive blind spot on impurity spectrum, that is, we cannot guarantee that new impurities that have never been encountered cannot appear when the current production process changes; the latter reflects people’s blind spot in cognition of analytical methods, that is, we cannot be sure. Current analytical methods can detect all impurities objectively present in the sample, or new impurities will not be found when the detection method is changed or other separation methods with different separation mechanisms are used.

The fourth quadrant is formed if the recognized impurity is not detected in the HPLC chromatographic system. The specific reason why the impurity was not detected should be further explored: the impurity is not present in the sample or the method is defective.

Impurities in the third quadrant represent blind spots in people’s cognition, which cannot be evaluated at the current stage, so they are a source of risk in the control of impurity spectrum. We can reduce the proportion of impurities in this part by strengthening the continuous understanding of the impurity spectrum, optimizing the impurity separation method and improving the detection ability, and promote the transfer of impurities from the third quadrant to the second and fourth quadrants. It is also possible to comprehensively evaluate all impurities contained in the analyte by developing complementary separation methods. “Four-quadrant” strategy system, expect impurities to exist in the third quadrant as little as possible.

In the next issue, we will introduce the construction of the “four-quadrant” strategy system based on risk assessment in detail. Pay attention to Welch Materials’ social media account, and we will continue to share related content of drug impurity profiling technology. The company provides high-quality innovative products and services that exceed customer expectations, and is committed to creating value for customers.