Quality from the start: A deep dive into analytical method development

In this webinar, Analytical Scientists Annie Wilt and Katlyn Jelosek at our North Carolina facility discuss their analytical method development process and some of the instruments used to test drug products, and ensure product quality and patient safety.

Hello and thank you for joining us today for “Quality from the start: A deep dive into analytical method development”. Today we’ll be discussing our analytical method development process and some of the instruments we use to test drug products and ensure product quality and patient safety. We’ll go through our considerations for phase appropriate test methods and clarify what to expect from analytical method development at Sterling Pharma Solutions.

I’m Annie Wilt and I’ve been an Analytical Scientist at Sterling for nearly three years, and I’ve got analytical method development and validation experience across the phases of pharmaceutical production and development, as well as in the biotechnology and textile industries.

My name is Katlyn Jelosek, and I’m and I’m an Analytical Scientist here at Sterling. The primary focus of my role at Sterling is method development for APIs. I’ve been developing methods for semiconductor materials and small molecule APIs for over six years.

Today we’ll go over the significance and purpose of analytical method development, some analytical objectives, and common pitfalls and how we avoid them, as well as some method requirements across the API development life cycle. We’ll also go over Sterling’s specific approach to analytical chemistry and how we build partnerships and facilitate communication for a better end product. We’ll also take a look at a case study that harnessed our analytical knowhow to solve a particularly difficult problem, and then we’ll answer some questions at the end.

There are a lot of reasons why we should focus on analytical method development right from the start of a project. Having dedicated analysts that focus on the development of robust test methods for the evaluation of intermediates or final APIs early on will aid in the understanding of the project’s needs and other necessary analytical techniques. For example, it’ll help us determine if the detector or instrument being used can show all of the intermediates or common impurities of the production process, or if any underlying impurities in a step, especially if there are any that will need an additional or separate method to make sure it’s under control and well monitored. Finding these potential stumbling blocks early on in your project will help avoid setbacks later on since we’ll have time to develop viable solutions to problems well before any deadlines. An early focus on development will also aid in the maintenance of product safety and efficacy because the work was put in to understand and characterise the molecule throughout R&D development. We take the time at the beginning to facilitate communication and build trust with our clients and internal departments to ensure alignment on project scope, priorities, challenges, and changes. We work as a team to build partnerships that maintain product quality and safety while keeping projects on track.

It can be challenging for organisations to discern how much and what types of methods to utilise in early development. Sterling can provide assistance with determining what customers need between too little and too much development. We truly aim for a sweet spot, zeroing in on phase appropriateness and the end use of a method. We aim to strike the right balance between being sufficiently thorough in our development and ensuring that the methods are fully developed and well tested without pushing for acceptance criteria that are simply not necessary for the phase or scope that is needed for your product. We strive to focus on the overall method goal to provide efficient method development that is suitable for the needs of the product. We have honest conversations about the end goals of the method and the project scope, especially if a project is at very early development or a one-off activity. This benefits everyone by keeping timelines on track, ensuring customer priorities are centred in our work. On the other hand, our overall goal is to provide our customers with a method that is easily modifiable for later stage validations. We aim for methods that are high quality and efficient, but we will not upsell you on levels of rigour that are simply not phase-appropriate. This can increase costs and increase timelines for our customers when it’s not necessary.

When Sterling develops a method for our customers, we do so with the intention that even if we are creating an early phase or more basic test method, our customers can return for later phase work and our method can be pulled forward to the next phase with as few changes as possible. Therefore, our development efforts usually have the same type of objectives for each project, with variations in the criteria for each phase. For chromatographic development, which makes up the vast majority of our work, common testing objectives are selectivity, which is a measure of how well the test method can isolate the target compound. It also evaluates if the method conditions cause the main compound to elute at a consistent retention time and if we are able to confirm the identity of the analyte peak. Related to that is specificity, which is a measure of how well the instrument method can separate the API or target analyte from its matrix. This is particularly important in mixed or highly impure samples and it ensures that we are quantifying our compound of interest and only the compound of interest. This provides certainty that our results and calculations are only considering the target analyte without interference from other compounds, preventing falsely inflated values or incorrect purity/potency calculations.

Accuracy determines if the amount measured by an instrument is the same as the true value by comparing the concentration of a known solution to the value calculated through the resulting chromatography. This is usually evaluated by measuring the recovery of a sample compared to a previously characterised standard. The resulting percent recovery value is used to quantify accuracy. Precision is the closeness of the results when a sample is tested repeatedly. We can look at precision in two parts: repeatability and reproducibility. Repeatability can show us how consistent our method is. We can see if the results are reliable under the same operating conditions and over a short period of time, and we can use this value to evaluate the stability of the method and the consistency between injections. Reproducibility shows that the method produces the same results across different analysts, instruments, and laboratories. This is included as part of phase three validation and later projects, but it’s a good idea to test at least informally at earlier stages. At the bare minimum, using a new column that’s the same part number as the original column that the method was developed for can show any changes to column chemistry caused by earlier projects or different analyses, and especially show any unknown or non-reproducible changes that can adversely impact the chromatography. This helps to avoid or prevent delays or the need for redevelopment if we discover it early instead of during production or midway through stability. Having multiple analysts execute a draft test method can help identify and close gaps in any written methodologies that the developer may not identify since they’ve become so familiar with the execution of the method. It also helps to clarify the testing procedures. Reproducibility testing also helps to identify and mitigate any quirks of instrumentation. We don’t want a situation where a project or compound can only be analysed on a single instrument. Our methods should be transferable to different instruments of the same type, allowing for variations in retention time, etc., caused by instrumentation or volume differences, but without having to do any redevelopment. It’s very difficult to justify in a GMP environment that a compound should only be tested on a single specific instrument in a single specific lab, so we ensure our methods produce good results wherever they are run.

Linearity measures the ability of an analytical procedure to obtain test results that are directly proportional to the concentration or amount of analyte in the sample within a predetermined range. We can extrapolate based on the calculated slope or a calibration curve to obtain the concentration or assay result, depending on the detector type. This range may either be linear or exponential. Typically, the expected range of a test method is between at least 80 to 120% of the nominal concentration of the API. However, later phase test methods or projects with isolated impurities may also evaluate a low-level linearity in order to accurately quantify the impurities or intermediates in the sample.

Peak symmetry is a marker of the efficiency of the elution of the peak. Better symmetry aids in the accurate quantitation of the compound and allows for better resolution between close eluting peaks. Good peak symmetry can also be an indicator of proper selectivity and specificity. Sudden changes in peak symmetry can also be an early indicator of declining column health and can be used as a system suitability marker during sample analysis.

The limit of quantitation (LOQ) is the lowest amount of analyte that can be reliably quantified. This is determined by measuring the lowest concentration or amount that yields a signal-to-noise ratio of greater than or equal to 10. Noise is simply the signal that is received from the detector when no analyte is detected. This range is calculated as the baseline noise range in chromatography. This is then compared to the signal of the peak detected. If the signal divided by the noise is greater than or equal to 10, it is considered reliably quantifiable. This is important to give the values detected validity. A small value has to be repeatedly quantifiable in order to be considered valid. If an impurity is detected that is lower than the LOQ, this is usually reported as less than LOQ instead of not detected. This is due to our limit of detection (LOD). This is the lowest concentration that can be detected. It is considered a signal with a signal-to-noise level of greater than or equal to three. This is usually calculated theoretically from the LOQ. Therefore, if the signal-to-noise is too low to quantify but is detected by the method, you can report your value as less than LOQ instead of not detected.

During development, analysts will work to decrease baseline noise from various sources. This ensures a low LOQ is possible. This is a good metric for impurity methods and evaluating reaction completion in in-process control samples. Suitability is a series of tests used during sample analysis and qualification to prove that the instrument or system is performing as expected and running within acceptable limits for analysis. This can include parameters such as repeatability by analysing a standard in replicate, usually three or six times, and determining the percent relative standard deviation to make sure that there isn’t any variability between injections and showing that the system can run several analyses back to back and produce consistent results. We may also evaluate accuracy by making a second standard solution to compare the percent recovery from the first. If this is within an appropriate range, we know that the instrument is accurately reporting the true amounts of analyte in our unknown solutions. We can also screen for any diluent interference as a way of verifying specificity and to catch any latent system performance issues such as air bubbles or pump malfunctions that may be harder to see in sample injections. Resolution of our peaks can also be evaluated before performing analysis of samples with multiple components or action completion IPCs. This is especially important if the compounds of interest are closely eluting and may interfere with one another. If there is a failure of any of the system suitability criteria, the analysis is stopped and a root cause of the failure is evaluated. Once we’ve identified the source of the failure, we’ll work to solve the problem. We’ve got lots of tricks up our sleeves to improve system performance, from cleaning columns and preparing standard solutions to purging air bubbles and pumps to ensure that our analysis gets back on track. We can also take advantage of previously demonstrated reproducibility and move the analysis to a different instrument. If the work is especially time-critical, the rigour of system suitability testing can be tailored based on the method characteristics or project needs. If an in-process control sample has a closely eluting intermediate, an analyst may prepare a resolution sample to ensure that the instrument can accurately separate the two based on previously qualified parameters. We evaluate critical parameters of an instrument method and the purpose of the analysis before determining our system suitability criteria to ensure suitable results every time.

There are some common pitfalls in method development that we can modify our approach to avoid. Lack of robustness is a lack of reliable results between analysts, instruments, or even columns. Developing robust methods can ensure that slight variations in method conditions will still produce consistent results. Measuring this in qualification allows a thorough understanding of the instrument method and how precise preparations of mobile phases, measurements of pH, etc., need to be to achieve the same results between instruments, labs, or analysts. Reliability of our test methods is key, especially for long-term projects or stability studies, as we need to be able to compare results between time points or campaigns. If the instrument method is not consistent over time or between product batches, then it is hard to identify any potential changes to the product or catch any developing problems.

Another pitfall is doing too much too soon or performing significant amounts of testing that is far more than what’s needed for the phase of the project. This slows down timelines and can consume more API than necessary, increasing overall costs or delaying timelines. We’re happy to develop a bespoke testing plan for our clients and to guide you on what testing is needed for your product, depending on your phase and what future submission trials and further development you will be using your product for. Along with having too much testing too soon can come rushed testing. When turnaround times for analytical testing are not reasonable for the amount of work involved, analysts are pushed to complete testing faster, and what comes with rushing are mistakes. We’ve all heard the saying “slow is smooth and smooth is fast”. It’s also true in our industry. If we take time to do everything correctly now, even if it takes an extra hour of the day, that can prevent the need to do an extra day’s work in the future. Taking enough time to complete analytical work from the start will allow for shorter turnaround times overall when testing is completed properly the first time.

Performing analytical testing also requires sample or standard material. We also need adequate quantities of compounds to set down for stability protocols or to qualify as reference standards, and it can sometimes be overlooked when setting production minimums or deciding how a batch will be used by a client. Certain tests can take upwards of 20 grams per reform, and if a batch is going to be part of a stability study or used as a quality control retain sample, then we will need a set amount of material to perform the necessary testing at each time point, plus some extra for ad hoc investigations or additional testing.

Communication will always be a benefit during development for both R&D and analytical chemists. Having a thorough understanding of what each step of the process is trying to achieve will allow analytical chemists to have a better focus on the goal of their method. This can help guide the prioritisation of key method parameters. For example, if there’s a need to determine resolution between two peaks that are impurities, or are we able to prove that both impurities are removed at a later step? This saves time and allows for more efficient method development. Knowing what previous developers have tried allows for more efficient development as well and better overall methods because analysts have a better idea of the pathways that were attempted by another scientist and helps them avoid the repetition of failed development approaches and keeps timelines on track. Knowing these pitfalls in advance helps us to keep projects and timelines going smoothly, ensuring high-quality test methods and results.

As the compound progresses through development, the level of rigour and burden of proof for the results generated increases. Methods should be acceptable and able to be validated for the next phase as is or with a few minor changes. For pre-clinical or phase one methods, we require that methods are fit for purpose and scientifically sound. We will generally qualify the methods through a memo-style process that focuses on the key parameters of the test method. These typically include specificity, precision, and accuracy at a minimum to verify that the results obtained are consistent and true. We can also evaluate linearity, especially for assay methods and limits of quantitation. If additional tests are required by the project or the specific analysis, we can modify the qualification process as needed. The qualification of a test method can also be used in setting acceptance criteria for system suitability or for exposing weaknesses in a test method that can be easily modified before implementation or use.

For example, during the qualification of a recent HPLC method, we realised that the components in a retention time marker solution were reacting with one another to form a new impurity. Using the data generated during qualification, we were able to update our test methods and release specifications with a modified retention time marker solution before product testing began.

Once we get into phase two and beyond, validation of analytical test methods will be done through a formal protocol and report process. The acceptance criteria used for these protocols will also be tighter for the different parameters being tested. For example, the precision of a phase one method could be considered acceptable for use with an RSD of 5%, but in phase two, we might tighten that acceptance to 2 to 3% RSD because there is a higher bar to clear to ensure that the data we generate is consistent each time. This trend continues through phase three and to commercial validations, but it’s not always necessary. Phase three and commercial methods must also demonstrate their robustness across different analysts and laboratories as part of their formal validation. This is to ensure that the method can be picked up by different labs or scientists and produce the same data, as well as really stress test a method and find critical parameters for commercial methods. All test methods in use are required to be validated, and validation reports can and should be included with regulatory filings. If there are additional methods needed at this stage, they would need to be validated prior to implementation.

When analytical scientists at Sterling are creating even a pre-clinical or phase one method, our goal is to make a method that will be easily validated at a later stage with few or no changes. We’re always open to improvements of our methods, especially as we learn more about a compound or if a production process changes, but we strive for a method that will be suitable for use for more than just its current phase.

As discussed in the previous slide, analytical methods are validated with higher levels of rigour as they progress into higher phases. Below are some of the regulatory requirements for validation or qualification for use at each stage. All that’s required to qualify preclinical methods is selectivity. As we mentioned previously, this is the ability to consistently determine the API peak of interest against any other peak in the sample matrix. During this qualification, we will look for repeatability, linearity, and most of the other requirements for at least a phase one method. We may have broader acceptance criteria at earlier phases as we hone in on the method and the molecule, but we strive for a method that can deliver reliable and repeatable results throughout the R&D process.

At phase one, there are some additional requirements. Here we include accuracy, repeatability, linearity, LOQ, and solution stability. Accuracy and precision, specifically repeatability in this case, help determine the consistency and accuracy of the method. Linearity provides evidence of the relationship between concentration and the result. For example, even if your method shows that the sample is only 80% pure, the established linear relationship proves that the assay result is true. An acceptable LOQ is important here as well, as we tend to take a closer look at specific impurities, even those at a low level at this stage.

In phase two, accuracy, selectivity, precision, and linearity are required to be evaluated, and the additional factor is specificity. This has to do with the matrix and other contents in the sample. As we mentioned in our previous example, the sample matrix can have an effect on the results of the method by generating background noise or additional impurity peaks that cannot be attributed to the analyte.

Peaks that cannot be attributed to the analyte. The overall limits of the testing are more stringent in earlier phases. A thorough explanation and scientifically sound reasoning can be used to decide or modify the necessary acceptance criteria for a method through the validation process. In later phases, more proof of the necessity of these changes is required if there is a need to allow for wider acceptance criteria. We may also validate test methods for their ability to quantify specific isolated impurities or intermediates in addition to the final product. Sometimes executing separate validation protocols solely focused on these intermediate compounds can ensure that their amounts are being accurately reported.

For phase three in commercial manufacturing, this phase requires all of the previous levels of validation plus extra analytical requirements such as robustness and the inter-lab analyst testing to prove it. Robustness will determine the allowable variance of the preparations of the method without affecting the performance of the method in chromatography. This could include the mobile prep, the diluent prep, or even the sample weight variance. This is measured to give more certainty in the preparation for future analysts and labs. For robustness testing, an analyst may prepare multiple different mobile phases at slightly different ratios of solvents or with different pH levels to mimic the differences between different analysts or glassware variations. They will use at least three different columns of different lots for analysis, and a lab will have at least one other analyst perform the testing from start to finish, and the data will be compared. All of this is done to ensure methods are fit for extensive use and that any changes to the results generated, such as changes to retention time or the appearance of new impurities, likely have a root cause that is not test method-based.

All of this validation and verification of the test method is done to ensure that the results produced are accurate and provide true information about the analytes being tested. Our focus is always on the safety and quality of the product. Part of ensuring that is proving that we’ve made what we said we did and that it’s as pure as the compound needs to be. Validation provides evidence and proof that your test results reflect reality and that the tested product is safe and of high quality before being used in the clinic.

Our approach to analytical method development is built upon a foundation of reliability and collaboration, and it’s grounded in our company values of caring, reliability, transparency, and willingness. We work to foster partnerships between all project stakeholders to develop an analytical programme that is efficient and provides support throughout process development and GMP production, or even to external formulators, by ensuring that our methods have been validated, qualified, or otherwise proven to be scientifically sound. By having all reported data go through technical review and by using GMP-qualified equipment regardless of phase to perform our testing, we can make sure that test methods and results we provide are reliable and dependable. We take pride in our methods being robust, working properly, and producing good data every time. We work collaboratively throughout all stages of the process by asking questions and being aware of process changes, and we’re transparent and honest about our challenges. This enables us to get to the root of our problems quickly and focus on solutions rather than blame or avoiding the truth. We work with our different departments to leverage what we’ve learned about a molecule and collaborate to solve problems across our areas of expertise.

For a recent production example, we had an intermediate solution that surprised us with relatively low stability, but we were able to use our learnings from how analytical samples had behaved when stored in different conditions to avoid a very large-scale product loss by quickly changing track and refrigerating all of our samples.

By keeping collaboration and communication as cornerstones of our project work, we can avoid some of the pitfalls of method and process development, such as ensuring that production minimums keep analytical testing needs in mind from the very start. If the analyst is alerted to any changes in the process that will call for a different test method, they can help monitor any new impurities. Collaboration also helps to ensure that priorities are aligned across all parties and that we aren’t spending time solving problems that are no longer relevant. This keeps projects on track and avoids wasted time.

At Sterling, we work very hard to share expertise and strive to develop partnerships with our clients and fellow scientists that enable us to produce the best methods and products possible. To give an example of method development here at Sterling, we’ll showcase an example of chiral method development and optimisation for an API with two chiral centres. This means there are four possible enantiomers. When analysing a racemic sample, meaning that equivalent amounts of each enantiomer are present, there should be a total of four distinct peaks. Analysts set out to create a chiral method capable of doing so.

When I started working on this project, this method development had started at the phase one level. This project was now returning for phase two work and was in need of further improvement for a new phase. In this case, we had four peaks we needed to fully resolve from one another, where we currently had three peaks resolved but two peaks did not have phase-appropriate resolution for the higher phase. As we mentioned in the previous slide, this customer’s API process selectively created one enantiomer of the final API. To test our final product and prove that the process worked to create only that enantiomer, we need to test it by separating each enantiomer through the use of chiral chromatography. We use chiral columns that have selective stationary phases capable of separating enantiomers of the same compound.

The separation we had with the original phase one method is shown in the chromatogram below. You can see that we have four peaks, but only some of those have proper resolution. What I mean by resolution is that the peak returns fully to the baseline before the next peak elutes. The final two peaks on the right have this, whereas the left two do not. Resolution is a mathematical measure that I won’t fully get into, but it is measured using chromatography software or can be done by hand.

Now that we had a goal in mind, I set out to tweak the instrument method by adjusting the ratio of mobile phases used, along with updating additives of the mobile phase to improve separation. We started out by changing columns, updating the types of mobile phase solvents used, and even changing additives that allow for pH adjustment or affect the affinity of the molecule to the column. After trying many different approaches with several other methods to work on simultaneously, the phase two project was quite a big one. We reached out to a vendor for assistance. Companies that create chromatography columns have resources that offer assistance with tricky separations. In this case, ours was quite tricky, and while we continued to work on other steps and methods for the process, we allowed our vendor to attempt this separation or to at least narrow down our column search. However, their analyst working on the method was only able to separate three of the four peaks of the final product, and they were surprised that there was another peak hiding underneath the first, shown in the chromatogram below. We held a meeting to discuss the results they obtained. They had even more trouble with this separation and landed on the same column as we were originally using. This told us to continue with this column and focus on other aspects of the method to improve separation.

So you may say, you can see all four peaks, why do you need more separation than what you obtained at phase one, especially when analysts from a vendor weren’t able to improve separation? Looking at the chromatograms below, we have an overlay of the racemic mixture of the final product seen in blue. This is our original separation and a chromatogram of the final product of only the desired enantiomer in pink using the same method. You can tell from the overlay that there is some tailing of the final product peak in pink that overlaps with one of the enantiomers in blue. This is concerning because we wouldn’t be able to measure the enantiomer that may elute underneath the tail of the main enantiomer peak, leading to a falsely passing result. The USP resolution we measure using our chromatographic software was 0.87. For most methods at the phase two level and higher, we need a resolution of 1.5 or separation such that reliable and consistent integration can be made. In our case, we need improved separation in the blue chromatogram. The last two peaks have a lot of tailing, and while this indicates a lack of symmetry of these peaks, we have separation. In this case, having resolution is more important than symmetry of these undesired enantiomers, but improving the peak shape was also a goal of the method development.

Here are some of the adjustments that we made to test out which parameters had the greatest effect on separation. This is also after many attempts to change the method parameters entirely and determining that we had the correct column types of mobile phase but maybe not the correct ratios of the mobile phases, additives to those mobile phases, or column temperatures used. We made various adjustments to the ratios of three mobile phases, changing each of the mobile phases to determine which would allow for separation of the earliest eluting peaks without causing a very long run time for the latest eluting peaks. We adjusted the temperature of the column compartment. This usually can decrease retention time and sometimes improves peak shapes, but some compounds are quite sensitive to changes in heat, so this could affect the separation of these two peaks. We did one last round of closely related columns to confirm no others were better suited for the separation. What we found was that by changing the column, no improvements and no peaks were found in the resulting chromatograms, meaning either the peaks eluted much too fast and none were separated, or the resolution was not improved, or nothing at all, meaning that the compounds were fully retained.

At one point, I realised there was another option I could try, but that was not normal for chiral methods. Most chiral methods are isocratic, meaning the ratio of mobile phase solvents is the same during the entire runtime of the method, but we hadn’t tried a gradient of this method, which is common for our other assay methods. So we separated each solvent out into different lines using a quaternary pump on our HPLC and began creating methods that would increase one of the solvents along the way and then another, sometimes two at once, and allowing another to decrease over time. What we found was that we were consistently using mobile phase A as the main solvent in the beginning, but we changed to having more of our mobile phase B, and this gave us the correct affinity to allow for separation of our peaks. And voila! I am very proud of this chromatography, and it was a long time coming. The chromatograms below show the results of the updates to this method, and you can see very well-resolved peaks in the racemic sample shown at the top and in the spiked samples. These spikes are of 0.5% of the undesired enantiomers. Those are also very well resolved. The resolution of the racemic mixture is 1.5, hitting what we wanted for adequate resolution, and the arguably more important separation is that of the spike sample, what you might see in a real sample used in release. That USP resolution between those first two peaks is 2.18, exceeding what’s necessary. Success was achieved, and the method was validated for use at the phase two level. This was the culmination of work that happened over a year, along with many other processes and methods. We were finally achieving success in an unconventional way, and this was a great win to hand over to our clients, who were very pleased with our work.

And now we’ll take some questions from the group. What are some regulations and guidelines that offer guidance on which tests should be conducted at each stage? So USP 1225 and 1226 are the validation and verification of compendial methods from the USP. We also have internal SOPs that provide guidance regarding phase-appropriate validations, protocols, and the like, but they are all based off of these two USP protocols. Can you elaborate on how cross-team collaboration comes into play for analytical method development, particularly with the chemistry and R&D team? Cross-team collaboration is continuous at Sterling. During development, analytical scientists use materials provided by either the customers or by R&D to separate and quantify compounds of interest, intermediates, and final products. During the time of development, analysts also provide R&D support in the form of testing samples provided by R&D of their process development. This allows for analytical to get continuous data to see any differences between the known compounds and typical impurity profiles. One example of this is when analysts were synthesising a specific isomer of a product during development of a small demonstration batch. The final product was tested for purity. It was noted that the final product did not elute or come off the column at the retention time previously seen in samples of the final product. There was also a smaller peak at the area of the desired product. R&D was quickly informed and shown these results. The retention time was confirmed by a previous injection of the desired material, so we were able to prove the method and instrument were working and suitable. After discussion with R&D, some confirmatory testing was done by NMR, and we were able to determine that the wrong isomer had been formed, and the smaller peak seen at the correct retention time was the correct isomer. We were able to quickly catch an issue and optimise the synthesis process for the better and ensure yield of the desired enantiomer.

What is the most common question you get from customers when you’re engaging in analytical method development, and how do you address it? The most common question we get from customers usually has to do with the amount of sample quantity needed for analytical testing and retention, and why. I think we covered fairly well why we need sample material to troubleshoot, develop methods, and test release or stability. We need to have extra on hand for investigation or unexpected testing problems, such as an instrumentation or system suitability failure, or even if a sample is prepared incorrectly. We need to have enough to re-prepare without having to source more material and cause delays. We always want to plan ahead for this need and make sure that everyone is aware early on, and that can also influence batch sizes and customers’ expectations of what they can do with their material when they get it. But we always want to be above board and make sure we can allot enough for proper tests.

What would cause scientists to go back and fix analytical method development later in the process, and how can they avoid this? If proper scale-up studies weren’t available or a large change in scale has taken place and the synthesis isn’t proceeding as usual, this could result in different impurities forming or the need for tighter specifications. In some situations, analysts have developed a method using this isolated impurity, and the reaction mixture that’s being tested isn’t provided until much later. So that could also create a reason they’re not getting real-time process samples. Communication between R&D and analytical is always needed to provide a thorough understanding of the process and what’s needed from testing.

How does Sterling’s approach to analytical method development differ from other outsourced partners? As we’ve reiterated, there’s definitely more openness and collaboration between R&D, production, and our customers than we’ve seen at other CDMOs. There’s an increased presence of analytical project meetings along every step of the way, and analytical is definitely more in the development conversation and brought on earlier in the project than I’ve experienced in other CDMOs or companies. It really shows how Sterling is a partnership developer rather than a contract researcher.

I’m Katlyn Jelosek, and I’m Annie Wilt. Thank you for joining us today for this look into our collaboration and quality-driven approach to analytical method development at Sterling.