Management of the impurity profile of an active pharmaceutical ingredient (API) is essential to manufacturing drug substance successfully in a manner that meets sponsor expectations for timeline and quality. The need to identify impurities in drug substance comes from the expectations set out in ICH Q3A (R2), with identification thresholds set based upon the dose of the final API.

Timely Impurity Identification

Impurities of principal concern are often first discovered through final product purity analysis, which for most small molecule drug substances utilizes high-performance liquid chromatography (HPLC) with UV detection. However, tracking and identification of impurities observed in in-process control testing, along with purity analysis of raw materials, can also be important to managing the total impurity profile of a drug — it is this control strategy that is most important to successful manufacturing and commercialization.

Timely identification of impurities requires a unique combination of process chemistry knowledge and considerable analytical capabilities. At Albemarle Fine Chemistry, these essential elements come together through combining decades of pharma research and development experience with teams who closely collaborate on all elements of process development, from the kilo lab to commercial production. These teams — formed early in process development — are then maintained through all phases of commercial development. This ensures that, when a need arises for prompt identification of an impurity, the puzzle pieces are already in place, and the team just needs to execute.

Understanding Customer Needs

The kickoff meeting also provides an opportunity for the CDMO and customer teams to get to know one another. Projects cannot be completed successfully unless the CDMO truly understands the short- and long-term goals of the customer. Only with the understanding of the customer’s primary concerns and essential factors for success can the CDMO establish the deliverables and determine what is needed to achieve those goals.

It is essential to be as flexible as possible so that important customer milestones can be achieved without issue, and so that robust, commercial-ready processes and methods can be established. It is also critical that, by the end of the kickoff meeting, all team members — from both the CDMO and customer — are very clear on what the deliverables are within the scope of the project and the timeline for those deliverables.

Putting Together the Puzzle — Technology is Just One Piece

While necessary for structural elucidation of impurities, technology alone cannot solve problems associated with identification and control of impurities. Technology, however, is an essential piece, and prompt access to modern analytical technology, including HPLC coupled with high-resolution mass spectrometry, nuclear magnetic resonance (NMR) spectrometers, gas chromatography – mass spectrometry (GCMS), liquid chromatograph (LC) fraction collectors, and semi-preparative LC is helpful in quickly identifying impurities. At Albemarle, not only have we made the needed investments in this technology, our lead analytical chemists are each trained in the use of these technologies, have walk-up access to the instruments, and have proven experience in applying the output of these technologies to solving chemical problems.

Completing the Puzzle — A Mass is Just the Beginning

Advances in modern mass spectrometry mean that, in many cases, it will be readily possible to obtain a mass of an impurity of interest. With the increased sensitivity of high-resolution instruments with tandem mass analyzers, such as a quadrupole time of flight mass spectrometer, a skilled analyst can have a parent mass along with several fragments — all with accurate mass — in a matter of hours. From here, the collaborative nature of our project teams shines through. Our analytical chemists then collaborate with team members from process chemistry about possible impurity formation mechanisms and propose potential structures for the impurity based on what is known. If needed, the same mass spectrometry techniques used to determine the mass of the unknown can be used to interrogate in-process control samples and raw material samples in order to identify upstream sources of impurities.

If additional information is still needed, work by both process chemists and analytical chemists is taken to further isolate the impurity or to chemically synthesize lab quantities of proposed structures through unambiguous routes. While fraction collection and preparative chromatography are valuable tools, it is worth considering that, in a 10-g sample of API, a 0.1% impurity is present at 10 mg (assuming similar response factors.) Therefore, these separation tools can be time consuming, and often do not yield sufficient material to perform further characterization. In these cases, it is often valuable to look at mother liquor streams for the impurity of interest — oftentimes, these impurities are enriched in such streams. These increased concentrations can help with attempts to isolate the impurity of interest or to obtain better quality mass spectrometry data.

Most often, however, we propose structures from the mass spectrometry data available, and, when straightforward, synthesize these compounds from known, unambiguous routes. This approach has multiple advantages: first, synthesis from a known route improves the confidence in the identification, and, second, produces enough material for work that comes after identification, including possible toxicology and response factor determinations. Once a potential impurity is synthesized, its structure can be confirmed by mass spectrometry and NMR. 

Putting the Pieces Together

After an impurity has been synthesized and its structure confirmed, the impurity should then be analyzed by the chromatographic methods which initially found the impurity, confirming the right entity was identified and synthesized. Work does not stop here, however. Occasionally, these impurities are already toxicologically qualified, or a sponsor may wish to qualify the impurity, but most often there is minimal toxicological coverage for new impurities, and therefore the formation of the impurity must be prevented. In these cases, the source of the impurity must be investigated — and this can involve raw material manufacturers, investigations of production campaigns, and development of new impurity control strategies; often, all three of these is necessary. Still the great collaboration and commitment by the team conducting the identification is called upon to complete these tasks, all with the goal of maintaining timelines to improve time to market for new medications.