From manufacturing to product storage, there are many opportunities for pharmaceutical products to become contaminated with organic volatile impurities capable of compromising product safety and quality. Extractables and leachables (E&L) and residual solvents are among the most common volatile impurities. There are several sources of these impurities; E&L can originate from medical devices, container materials and single-use systems, while the widespread use of solvents during manufacturing presents another potential source of contamination. For example, the dissolution of ethyl cellulose in N-methyl pyrrolidone, a volatile organic solvent, is a key step in the production of a gel for periodontitis treatment1.
Regulations are in place to help safeguard public health and are only becoming stricter following major recall incidents. Therefore, there is an increasing need for sensitive and efficient analytical methods to demonstrate the purity of active pharmaceutical ingredients (APIs) and finished products. When coupled to gas chromatography (GC), automated headspace sampling (HS) simplifies impurities testing workflows and generates the high-quality data needed to produce results, with confidence.
Regulations Reflect the Importance of Ensuring Product Purity
Pharmaceutical manufacturers and testing laboratories must ensure products do not contain toxicologically significant levels of residual volatile compounds used in manufacturing processes. Acceptable exposure limits are set according to the health hazard severity posed by impurities and are derived from a combination of in vivo toxicity studies, tumorigenicity tests and mathematical modeling.
Following the assessment of products against these limits, several carcinogens have come into the spotlight, triggering a reassessment of regulations and detection methods. In 2018, nitrosamines, known as mutagenic carcinogens, were detected at unacceptable levels in generic drugs containing the API valsartan. A major recall ensued and sparked the development and refinement of sophisticated sampling methods for impurity control, as well as an update on related regulations.
Following the valsartan incident, the U.S. Food and Drug Administration (FDA) also highlighted the need for better change control processes and awareness of cross-contamination risk2. Currently, the International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use3 refers to the US Pharmacopeia (USP) <467> for the analysis of volatile organic impurities4, which was updated at the end of 2019. Among the changes aimed at improving patient safety, the latest version of USP <467> defines acceptable amounts of residual solvents and outlines procedures for their analysis – triggering a global reassessment of current methods for impurity analysis.
The Challenges of Quantifying Volatile Impurities in Solid Products
Those quantifying organic volatile impurities will be familiar with the time-consuming process needed to mitigate the common concern known as “carryover”. Injectors in early static HS-GC systems retain high-boiling point solvents used to dissolve samples for headspace analysis, compromising the accuracy of results. To overcome this issue, the injector needs to be routinely cleaned with blank runs between test samples. However, this risk-reducing step is tedious and adds to the overall burden laboratories face, including increasing work complexity and volume, outdated and inefficient informatics tools, and pressure to meet performance metrics without compromising data integrity. Analysts must also regularly record relevant metadata, enter data manually, review documents, calibrate instruments—and the list goes on. To ensure deadlines are met, it is common for instrumentation to run unattended overnight; therefore, robustness and reliability are non-negotiable qualities.
However, the ideal analytical tool needs to offer more than just high throughput capabilities and reliable results. In research and development, flexibility is another rare yet desirable trait; having the ability to easily change and monitor methods for process development would greatly enhance productivity. Yet, some headspace systems for volatile impurity quantitation require tedious and complex manual configuration to accommodate different vial sizes, and restrict runs to only one vial size at a time.
Not only does impurity analysis require the detection of known contaminants and residuals; there is also a need to monitor for previously unknown E&L or impurities resulting from process changes. Without the ability to obtain accurate mass and structural data, the detection of novel compounds at low concentrations remains a significant challenge.
Headspace Sampling Coupled to GC Transforms Impurities Testing Workflows
HS sampling is a separation technique that relies on organic volatile compounds reaching an equilibrium between the solid (e.g. the API) and gas phase. Based on this principle, volatile compounds are removed for analysis, leaving behind heavier, non-volatile matrix compounds. Together with GC, HS sampling provides a sophisticated testing method for the analysis of E&L and residual solvents in APIs and finished products.
Drive Productivity with a Simplified, Dynamic Approach to Headspace Sampling
Harnessing sensitive technology that boosts sample throughput and accelerates productivity is critical to the success of busy analytical laboratories. With several decades of experience in testing for E&L across a wide range of industries, SGS is a company highly familiar with these concepts. SGS is recognized as the global benchmark for quality and integrity, and operates a network of more than 2,600 offices and laboratories around the world. To maintain its status as the world’s leading inspection, verification, testing and certification company, SGS has been using automated HS coupled to GC for the analysis of organic volatile impurities.
Using the Thermo Scientific TriPlus 500 Headspace Autosampler coupled to the Thermo Scientific TRACE 1310 GC and Q Exactive GC Orbitrap GC-MS/MS systems, SGS has been reaping the benefits of enhanced productivity. With a 120-vial capacity, combined with the ability to include 10 and 20 mL vials in the same run, it is easy for SGS to achieve higher throughput. Automated HS has made it possible to overcome the need for tedious cleansing of the injector at SGS, as the autosampler is connected directly from the valve manifold to the GC and GC-MS/MS systems. Such a setup shortens the sample path and addresses the carryover problem, as evidenced by the lack of residual solvent detected in blank runs (Figure 1). While headspace samplers with long transfer lines can suffer from cold spots — where samples condense and cause contamination — short transfer lines are heated with greater efficiency and consistency, which translates to more reliable results. Improved system inertness is another major benefit of the direct connection, which enables better sample integrity during transfer into the column with less dead volume.
Figure 1. The TriPlus 500 HS autosampler GC-FID chromatogram of a blank injection (bottom) immediately following injection of pure Isopropanol Alcohol (IPA) (top) shows virtually no carryover. Chromatograms courtesy of SGS.
With the issue of ‘carryover’ eliminated, scientists at SGS can spend less time optimizing methods and running blanks, and instead direct their energy toward activities that make better use of their specialist skills and experience. To execute analyses efficiently and deliver client results on time, having a user-friendly software that can be programmed easily is critical. At SGS, the greater flexibility afforded by the associated Thermo Scientific TraceFinder software is significant, particularly for bioprocess optimization where methods need to be changed regularly.
Quantitate and Detect Organic Impurities with Greater Confidence in Your Data
One of the key benefits of the direct-connect HS autosampler design is the enhanced confidence that analytical scientists can have in their data. Greater sample integrity and system inertness translates to a better chromatographic peak shape and high repeatability of results, which is important not only for the routine analysis of known impurities. High-resolution accurate-mass GC-MS data also helps with the identification and structural elucidation of unknown compounds, which is increasingly needed for the analysis of E&L (Figure 2).
Figure 2. TriPlus 500 HS autosampler-Q Exactive Orbitrap GC-MS/MS mass spectrometer analysis of a negative control sample in which E&L are not present (top) and a test sample (bottom) showing an unknown peak in the MS chromatogram. Chromatograms courtesy of SGS.
With the 1-ppm mass accuracy of the Thermo Scientific Q Exactive Plus, SGS can be confident in determining the chemical composition of low-concentration components, and avoid the need to repeat runs (Figure 3).
Figure 3. Based on the high-resolution accurate-mass spectrum of the unknown peak provided by the Q Exactive GC Orbitrap GC-MS mass spectrometer, the formula of each fragment can be confirmed and the unknown compound structure elucidated. The table is reporting the 1-isopropenyl-2,2,4,4-tetramethylcyclohexane fragments information, showing exceptional high mass accuracy provided by the Orbitrap technology even for low masses (< 100 m/z). Chromatogram courtesy of SGS.
Advanced software tools are now also available to help laboratories make real-time decisions and avoid the manual review and re-analysis of suspect samples. SGS uses the Thermo Scientific TraceFinder software, for example, to trigger preconfigured actions on flagged results for real-time rejection of samples, insertion of blanks, and to halt sample sequences in which data has fallen outside of predefined criteria — all of which are particularly valuable for laboratories undertaking substantial method development.
Conclusion
Rigorous chemical analysis is critical to ensuring pharmaceutical products do not contain toxicologically significant levels of volatile organic impurities, such as E&L and residual solvents. While sensitive and reliable methods are needed to ensure product quality and safety, pharmaceutical manufacturers also require greater flexibility and simplified workflows. Automated HS sampling coupled to GC meets this need and helps manufacturers to remain compliant while achieving greater analytical efficiency. A shortened sample path is achieved through direct-connect HS autosampler designs, which translates to a sharper chromatographic peak shape and more reproducible results. The method also allows data to be generated with greater confidence and enables manufacturers to ensure the purity of pharmaceutical products and safeguard patient health.
References
1) Wasilewska, K., Winnicka, K. Ethylcellulose—A pharmaceutical excipient with multidirectional application in drug dosage forms development. Materials, 12 (20) 2019.
2) FDA Statement on the FDA’s ongoing investigation into valsartan and ARB class impurities and the agency’s steps to address the root causes of the safety issues, 2019.
3) Impurities: Guideline for Residual Solvents Q3C(R7), ICH Harmonised Guideline, International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use, 2018.
4) General Chapter USP <467> Organic Volatile Impurities, Chemical Tests, United States Pharmacopeia, 2012.
About the Author:
Dr. Dujuan Lu is the Technical Client Manager/Global Lead for Extractable and Leachable Testing at SGS.