Standardized Extractables Testing Protocol for Single-Use Systems in Biomanufacturing Part 4
This article presents a consensus standardized extractables testing protocol for single-use systems in biomanufacturing
This article was originally published in the November – December 2014 issue of Pharmaceutical Engineering® magazine. Missed Part one, two, and three? Catch up now – Standardized Extractables Testing Protocol for Single-Use Systems in Biomanufacturing – Part 1, Part 2, and Part 3. Get part five delivered straight to your inbox by subscribing to iSpeak.
The goal of the analytical techniques used in extractables testing is to identify and quantitatively assess compounds resulting from the extraction of SUS components. The results can then be used for safety assessments.20 In cases where quantitation is not possible, semiquantitative values should be reported. Extracts referenced in this section on analytical techniques are the solutions generated by the use of solvents on SUS components during extractables testing studies.
The analytical techniques proposed in this article were selected to detect the widest possible range of chemical compounds. An individual compound detected at a concentration of 0.1 µg/mL or greater should be identified, confirmed and quantified by use of an authentic reference compound (e.g., extractables known to result from component raw materials). Compounds observed at a concentration below 0.1 µg/mL should be identified by mass spectral library match and confirmed with quantitation if an authentic reference compound is available. When an authentic reference compound is not available, a chemically similar compound may be used although this will result in semiquantitative values in the results. (See Appendix: “Recommended Analytical Techniques for Extractables Identification and Quantification.”)
Analysis by high-performance liquid chromatography (HPLC) or Ultra-high-performance liquid chromatography (UHPLC) coupled with photodiode array (PDA) detection and mass spectrometry (MS) is required for all extractables testing. It is acknowledged that certain extraction solvents may present challenges in detection (i.e., PS-80 extracts). Dilution of the extracts to acceptable matrix interference concentrations is acceptable in these cases (e.g., 0.1% PS-80).
Mass spectrometric analysis should be conducted in both positive and negative mode with electrospray ionization (ESI) as well as atmospheric pressure chemical ionization (APCI) techniques. Use of two ionization methods provides complementary data and allows detection of the maximum range of potential extractable compounds resulting not only from bulk component material, but from additives and degradation products as well.
Gas chromatography (GC) with headspace inlets for volatiles and direct injection inlets for semivolatiles is also required for all extractables testing. Mass spectrometric detection should be performed in conjunction with either technique to permit compound identification via mass spectral libraries. Alternate detectors (e.g., nitrogen phosphorus, flame ionization, or nitrogen chemiluminescence) for specific classes of compounds may be used in addition to MS detection if required due to the nature of the specific component materials and potential extractables involved.
Inductively coupled plasma mass spectrometry (ICPMS) also should be performed to detect and quantify extractable metals. Optical emission spectroscopy (OES) as an alternate detection method may be used provided specificity and required detection limits can be achieved. Extracts should be analyzed intact unless dilution of the samples allows the required detection limits to be met for all metals of interest. In cases where the extract matrix would produce known interferences in detecting particular metals, a different isotope should be selected to minimize the interference. At a minimum, the amounts of all metals appearing in extracts that are specified in USP <232>,21 EMEA,22 and ICH guidelines23 should be quantified and reported. While it is only required to record results from the final extractables testing time point, additional time points may be analyzed as necessary.
The detected and identified compounds should be named based on International Union of Pure and Applied Chemistry (IUPAC) nomenclature, and reported with Chemical Abstracts Service (CAS) registry number, empirical formulas, chemical structures, and molecular weights, when possible.
Additional analytical techniques should be used to supplement the required data, in particular, to determine the total organic carbon (TOC) and pH of extracts when the test solvent does not interfere. Nonvolatile residue determination may be necessary in addition to the required analytical techniques when the test solvent is volatile. Resulting extractables testing data should be compiled into an extractables test report with representative chromatograms and raw data tables of the results. The extractables test report should include the amount and identity of known compounds and the estimated amount and class of compound for unknowns. The extractables test report also should include the analytical conditions for each technique as well as any additional discussion necessary to provide enough context such that the results are readily interpretable by end users. Specific analytical parameters and method sensitivity criteria are presented in the Appendix.
The authors sincerely thank the more than 40 colleagues from 18 companies in the BPOG Extractables Work Group for their contribution in developing our proposal, and in particular the following individuals: Tony White, Director BioPhorum Operations Group; Gerry McAuley, Facilitator BioPhorum Operations Group; Bobbijo V. Redler, Ph.D. Associate Principal Scientist Merck & Co., Inc.; Nancy Sweeney, Senior Scientist MTS Gallus Biopharmaceuticals; Ping Wang, Ph.D. Principal Scientist Johnson & Johnson; Russell Wong, PhD Sr. Manager Manufacturing Sciences – Raw Materials, Bayer Healthcare; Sally A. Kline, Scientific Director Amgen; Cara Weitzsacker, PhD QC Senior Specialist Bayer HealthCare; Amy J. Stitt, M.S. Scientist I Bristol-Myers Squibb; and Robert Repetto, Senior Director External Affairs Pfizer. The authors appreciate the detailed constructive review of the manuscript from Dr. Duncan Low from Amgen and Dr. Dennis Jenke from Baxter. We also thank Terry Hudson from Genentech, Dr. Michael T. Jones from Pfizer for reviewing the article, and Jim Sayer from Amgen for providing useful input on gamma irradiation.
Weibing Ding, PhD, is a Principal Scientist in Process Development at Amgen Inc.
Gary Madsen, PhD, is a Senior Principal Scientist with Pfizer Analytical R&D Bio-Therapeutics Pharmaceutical Sciences.
Ekta Mahajan is a Senior Engineer in Pharmaceutical Technical Development Engineering group at Genentech/ Roche in South San Francisco, CA.
Seamus O’Connor, PhD, is an Associate Manager, Analytical Sciences, Industrial Operations and Product Supply at Regeneron Pharmaceuticals, Inc.
Ken Wong is a Deputy Director in Process Technology group at Sanofi Pasteur in Swiftwater, PA where he serves as the site E/L SME.
20. Product Quality Research Institute, Leachables and Extractables Working Group, “Safety Thresholds and Best Practices for Extractables and Leachables in Orally Inhaled and Nasal Drug Products,” 2006. http://www.pqri.org/pdfs/LE_Recommendations_to_FDA_09-29-06.pdf.
21. United States Pharmacopeial Convention, “Elemental Impurities – Limits,” United States Pharmacopeia 35 NF30, <232>, 2013. http://www.usp.org/sites/default/files/usp_pdf/EN/USPNF/key issues/c232_final.pdf.
22. European Medicines Agency, “Guideline on the Specification Limits for Residues of Metal Catalysts or Metal Reagents,” EMEA/CHMP/SWP/4446/2000, 2008. http://www.ema.europa.eu/docs/en_GB/document_library/Scientific_guideline/2009/09/WC500003586.pdf.
23. The International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use, “Guideline for Elemental Impurities,” Q3D, Current Step 2b version, 2013. http://www.ich.org/fileadmin/Public_Web_Site/ICH_Products/Guidelines/Quality/Q3D/Q3D_Step2b.pdf.