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Alternative Solvents for Extractables and Leachables Evaluation
Extractables and leachables evaluation of packaging components and components of bioprocessing systems is a crucial regulatory requirement. Solvents used for evaluation of process components may include surfactants that can interfere with chromatographic detection and contaminate the chromatographic system. The authors examine alternative solvents that provide extraction equivalence and do not interfere chromatographically.
To demonstrate that a container-closure system or a bioprocessing component is suitable for its intended use, the components typically undergo an initial extractables screening. The design of the extraction experiment should appropriately exaggerate the conditions of real-time use without breaking down or degrading the polymeric component under testing. Although strong polar and strong nonpolar solvents, such as 100% isopropanol (IPA) and 100% hexanes, are commonly used for highly aggressive reflux or soxhlet extractions of final container-closure systems that typically contain the final drug product for extended periods of time, these solvents are not always appropriate for the components of bioprocessing systems, such as bioprocess bags, filters, tubing, O-rings, diaphragms, and gaskets (3, 4).
For bioprocessing components, initial extractables screening involves filling or immersing the component in a variety of model solvents that more closely represent the formulation and exaggerate the conditions of real-time use. The components are incubated in the solvent for a predetermined length of time at an elevated temperature, such as 40–60 °C for several days, weeks, or even months. This type of extraction is recommended over an aggressive reflux extraction because exposure of the formulation to bioprocessing components is usually very short, and the temperatures of real-time use are typically at or below 25 °C. Reflux extraction of bioprocessing components using strong polar or nonpolar solvents is commonly not recommended unless the real-time exposure of the formulation to the component is long or at temperatures greater than 25 °C and the component is compatible with the solvent (3, 4).
Many extractables studies have instead used model solvents that are comprised of the same excipients that are present in the process buffers, drug substances, or final drug-product formulations. These excipients often include surfactants, which are common ingredients in the formulations of biologics and are regarded as essential components of the model solvents to be used to generate extractables profiles. Surfactants, however, pose major chromatographic interferences when screening for nonvolatile organic compounds by high-performance liquid chromatography–mass spectrometry (HPLC–MS). The detection of extractable compounds may be masked by co-eluting surfactant peaks. In addition, high concentrations of these surfactants are problematic, as they can contaminate the HPLC–MS system. Dilution is not always a viable solution because sensitivity can be greatly affected. Therefore, alternative solvents that provide extraction equivalence and do not interfere chromatographically were examined.
Materials and methods
The PP caps were chosen for this experiment due to the presence of known additives that could be easily tracked during extractables screening. Compounds previously observed through extractables studies that were targeted in this experiment include a di-tert-butyl(phenyl)phosphite (Irgafos 168, BASF); a phosphate oxidative degradant of Irgafos 168; ethylene bis(heptadecanamide); pentaerythritol tetrakis 3-(3,5-di-tert-butyl-4-hydroxyphenyl) propionate (Irganox 1010, BASF); and erucamide. Determination of extraction efficiency equivalence was made by comparing the responses for each of the targeted extractables observed in each solvent.
Figure 1a presents a visual representation overlay of LC–MS total ion chromatograms (TIC) of all the solvents used for the extraction study. The total ion chromatograms of the 60% IPA and 15% EGMB are similar; therefore, they cannot be distinguished in the figure but are shown as the gray/red line. Figure 1b presents a visual representation of just the overlay of 0.1% PS 80 and 60% IPA to show that any potential extractables would be masked by the PS 80 interference. Interferences are also observed with 0.1% PS 20 and 1% Triton X-100 as shown in Figure 1.
In addition to having greater extraction efficiency, as demonstrated in two separate studies using two different types of material, 60% IPA was also shown to eliminate interferences observed in the sample chromatography of surfactants. The use of extraction solvents that do not pose significant chromatographic interferences is critical so that potential extractable compounds are not missed during the extractables screening. In this case, erucamide was extracted in all of the solvents and was tracked using extracted ion analysis based upon the total ion chromatogram of the 60% IPA solvent. To perform extracted ion analysis, the ion of interest must be known. If only the total ion chromatograms of the surfactant solvents were used to screen for potential extractable compounds, and IPA was not used as one of the extraction solvents, erucamide would have been missed in the chromatograms of the surfactants. Erucamide elutes at approximately 7.7 minutes in Figure 1.
Jennifer M. Roark, Mai N. Jacques, PhD, Erica J. Tullo, PhD, Andrew T. Blakinger, and Thomas C. Lehman*, PhD are all in the Method Development & Validation group at Eurofins Lancaster Laboratories, 2425 New Holland Pike, Lancaster, PA 17601, USA, 1.717.656.2300, www.lancasterlabspharm.com.
*To whom all correspondence should be addressed.
1. FDA, Guidance for Industry: Container Closure Systems for Packaging Human Drugs and Biologics (Rockville, MD, May 1999).
2. CFR Title 21, Part 211.65 (Government Printing Office, Washington, DC, 2006).
3. Bio-Process Systems Alliance, Recommendations for Testing and Evaluation of Extractables from Single-Use Process Equipment, (Washington, DC, 2010).
4. Bio-Process Systems Alliance, BioProcess Intl. 5 (11) 36-49 (2007).
5. T. C. Lehman, "Evaluation of Model Solvents for Generating Extractable Compound Profiles from Single-Use Systems," presentation at AAPS Annual Meeting and Exposition (Washington DC, 2011).
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