Sponsored by Sartorius
Regulations in the United States and elsewhere require that 100% of parenteral drug products be inspected for defects and that released drug products be “essentially free” or “practically free” of visible particulate matter. Single-use systems (SUS) and equipment used in bioprocessing must meet the level of cleanliness required to enable biopharmaceutical manufacturers to meet current good manufacturing practice (CGMP) and pharmacopoeial requirements regarding particulate matter.
USP <788> and USP <790>
Historically, biomanufacturers have asked SUS suppliers to “meet USP <788> specifications for particulate matter” (1). However, as discussed in detail in a recent BPI article, the US Pharmacopeia’s (USP) chapter <788> is misapplied to SUS (2). The “Particulate Matter in Injections” chapter describes test methods and limits for subvisible particulate matter in parenteral drug products only — not SUS. Adaptation of those test methods to the measurement of particulate matter in SUS requires addition of nonstandardized procedures and making multiple assumptions to transform USP <788> acceptance criteria into acceptance criteria for SUS. In addition, the commonly applied particle-measurement technique of light obscuration cannot detect particulate matter reliably in the usually defined visible size range (≥100 µm) according to USP <1788.1> guidance (3). TA Barber’s text on particulate matter in pharmaceuticals manufacturing describes in detail the significant limitations of light obscuration in detection of large and fiber-shaped particulate matter (4).
In another unfortunate trend, biopharmaceutical manufacturers increasingly ask suppliers to assure that SUS are “free of visible particles” according to USP <790> “Visible Particles in Injections” (5). Like USP <788>, USP <790> does not describe test methods to measure visible particulate matter in SUS. A common misperception is that the latter describes a procedure and specifications for visual inspection of SUS to detect the presence of particulate matter. But USP <790> describes a procedure only for the manual visual inspection of parenteral drug products such as those in vials or syringes, not SUS. The method requires five seconds of inspection each against white and black backgrounds, with the lighting intensity between 2,000 and 3,750 lux.
In addition, a “sampling at batch release” of drug product is reinspected according to the sampling plan based on an acceptable quality limit (AQL). The USP <790> inspection conditions are not necessarily optimal for — and the AQL required is not realistically applicable to — visual inspections of SUS. Similarly, neither the European Pharmacopoeia’s EP 2.9.20 nor the Japanese Pharmacopoeia’s JP 6.06 is practically applicable to SUS (6, 7). However, some guidance in USP <1790> “Visual Inspection of Injections” is applicable to the development of visual inspection procedures for SUS (8).
Free of Visible Particles?
The expectation that SUS be “free of visible particles” presents SUS suppliers with multiple challenges. A requirement to manufacture SUS with zero visible particles will increase costs significantly. To meet such requirements, a visual inspection of each final SUS product would have to be capable of detecting 100% of visible particulate present in the SUS. As discussed below, that is an unrealistic requirement given the probabilistic nature of visual inspection processes and significant challenges associated with detecting particles on the inside (fluid-contacting) surfaces of single-use equipment and components.
A quality risk management (QRM) approach for determining risks of final drug products containing SUS-borne particulate matter requires consideration of three factors: detectability, probability of occurrence, and severity of harm (9). For a detailed discussion on reducing the probability of occurrence of particulate matter in SUS manufacturing, see the most recent recommendations from the Bio-Process Systems Alliance (BPSA) (10). Also in those recommendations is a detailed discussion of how the potential severity of harm from particulate matter in SUS strongly depends on the SUS application in a given bioprocess, which we summarize herein.
For SUS applied upstream of sterilizing-grade filters, all particulate matter larger than ~0.2 µm on the inside (fluid-contacting) surfaces of a SUS released during bioprocessing would be captured by the downstream filter. In such an application, the potential severity of harm to final drug products is considered to be minimal according to the rationale detailed in the BPSA recommendations (10).
However, the potential severity of harm would be significant in applications of SUS downstream of final sterile filters (final filling) or in aseptic bioprocesses (cell-therapy manufacturing). In such critical scenarios, particulate matter on the inside (fluid-contacting) surfaces of SUS could release during bioprocessing and directly contaminate final drug products.
Acceptable Risks and Realistic Expectations
Final parenteral drug products are subject to a highly regulated 100% visual inspection procedure (5, 11), so the potential harm from particulate matter in them is more often the cost associated with rejected drug products rather than a risk to patient safety. A bioprocess is considered to be under control when a very low (e.g., 1%) and stable number of rejects is discovered during visual inspection (12).
In critical SUS applications, the equipment and components must be “essentially free” or “practically free” of visible particulate matter with respect to their potential contribution to the overall burden of visible particulate matter in final drug products. Particulate matter released from a SUS does risk causing additional rejects during visual inspection of final drug products. For some bioprocesses, the risk that small numbers of drug products will be rejected for visible particulate matter released from SUS may be acceptable. The relationship between the levels of visible particulate matter in a SUS to the potential number of final drug products rejected in a visual inspection depends highly on the specifics of a given process.
As a theoretical example, consider that our study, previously published in BPI, showed that about 10 particles ≥100 µm in size were present in a market sampling of 20-L SUS bags (13). Consider two SUS application scenarios (Table 1): In a cell-therapy manufacturing scenario, 10 visible particles in a 20-L SUS bag could cause a ≤50% rejection rate for the final drug products. In a vaccine manufacturing scenario, the same number of visible particles released from the same type of SUS would lead to a 0.05% rejection rate at most. The estimations associated with these risk scenarios generate realistic expectations on the levels of particulate matter that can be tolerated in SUS, which differ significantly from the expectation that SUS be “free of visible particles.”
Table 1: Numbers of potential drug product rejects due to visible particulate matter in single-use system (SUS) bags.
What Is “Visible” in a Visual Inspection?
As TA Barber detailed, manual visual inspections are pass–fail attribute tests relying on human-operator judgment (4). Thus, manual visual inspection is a probabilistic and problematic process due to human subjectivity.
In visual inspection theory, good probability of detection (PoD) starts at 70%, and the PoD for particulate matter depends on many factors. No single particle size clearly defines “visible” versus “subvisible” (not visible), and visual inspection becomes more deterministic as particle size increases. As USP <1790> highlights, “The visible particulate detection process is probabilistic: The likelihood of detection is a cumulative function of visible attributes such as particle quantity, size, shape, color, density, and reflectivity as well as optical characteristics of the surrounding product and package” (8). A recent BioPhorum article provides more detail for consideration (14).
Given the probabilistic nature of visual inspection, it is impossible to guarantee an absolute requirement such as the absence of visible particulate matter using such a method. As USP <1790> confirms, “Although zero defects is the goal, and this should drive continuous process improvement, zero defects is not a feasible specification for visible particles given current packaging components, processing capability, and the probabilistic nature of the inspection process” (8).
As USP <1790> indicates, for particulate matter in glass vials, possibility of detection (PoD) >70% starts at 150 µm for particles and 500 µm for fibers. SUS are less transparent and much larger than glass vials. As published in the PDA journal, our company’s detailed study of the detectability of particulate matter in SUS demonstrates lower detectability (smaller PoD) in SUS compared than that found for glass vials (15).
That study details the development and validation of an optimized manual visual inspection procedure for detection of particulate matter on the interior surfaces in SUS assemblies of 2D bags with tubing lines. The procedure was validated for reliable detection of black and clear particles ≥1000 µm in size over the entire range of SUS examined (up to 50-L bags with up to 5-m long tubing lines attached). Fibers of ≤2000 µm in length yielded a PoD <10% and thus were poorly detectable in these SUS. The study confirmed that detectability depends strongly on color (e.g., black or clear) and shape (e.g., particles or fibers) of matter present. The study clearly showed that, as SUS size and complexity increases, the detectability of such matter decreases substantially. All SUS inspected were translucent; for SUS components that are nontransparent, a visual inspection of interior surfaces is not possible. Note that the optimal visual inspection conditions described in our study (Table 2) differ significantly from those required by USP <790>.
Table 2: Comparison of optimal visual inspection conditions for single-use systems (SUS) compared with USP <790> requirements (15); AQL = acceptable quality limit.
Because visual inspection is clearly probabilistic, SUS suppliers might miss particles during final visual inspection of SUS products, but biomanufacturing users could find those particles in a visual inspection before SUS implementation. Given the absence of standards specific to SUS, an effort is currently underway in the American Society for Testing and Materials (ASTM) E55 committee on pharmaceutical processes to develop a standard for development and validation of visual inspection procedures for SUS components and assemblies.
“Visible” in a Sartorius Visible Particle Test
The advantage of visual inspection is nondestructive testing for 100% of SUS equipment and components produced. However, given the limited detectability of particulate matter in a visual inspection, a sample of SUS produced should be tested destructively using liquid extraction according to BPSA recommendations (10). As discussed above, no such test is described in USP <788> (2).
In a recent paper in the PDA Journal, we detail our recommendations for “best practices” in quantifying and identifying particles on interior SUS surfaces (16). Three procedures are required for particle extraction, sample preparation, and particle measurement. ASTM E3230-20 describes development and qualification of effective extraction procedures (rinsing and/or agitation) for detaching particles from the interior surfaces of SUS (17).
In the sample-preparation procedure, liquid extract containing particles from the extraction procedure is filtered through a membrane. In the particle-measurement procedure, particles captured on that membrane’s surface are counted and sized using automated membrane microscopy. If particles only ≥100 µm are found, then at Sartorius we denote that as a “visible particle test” or Sartorius VPT.
Manual visual inspection requires a human inspector to detect the (usually) low levels of particulate matter dispersed over the (usually) large interior surfaces of a SUS. The Sartorius VPT concentrates particulate matter extracted from a SUS onto the small surface of a membrane filter. Rather than depending on human eyes, a microscope with an imaging system (motorized stage, digital camera, and imaging software) automatically counts and sizes particles present on that filter’s surface. Note that in addition to efforts toward developing standards specific to the measurement of particulate matter in SUS, another effort is underway in the ASTM E55 committee on pharmaceutical processes to develop standards for test methods based on automated membrane microscopy.
In a manual visual inspection of SUS, particles ≥1000 µm might be “visible” and detectable with a PoD >70%. With the Sartorius VPT, particles ≥100 µm are highly detectable (PoD of nearly 100%), so all particles ≥100 µm are made “visible” through microscopy. Only a small fraction of those measured in the Sartorius VPT are potentially visible in a visual inspection of SUS.
The Sartorius VPT is an essential tool for quantitatively assessing the risk to final drug products from particulate matter present on the inside (fluid-contacting) surfaces of SUS. Only by analyzing quantitative data with calculations specific to a bioprocess of interest (Table 1) can suppliers and users determine whether SUS will be clean enough to produce final parenteral drug products that are “essentially” or “practically” free of particulate matter.
References
1 USP General Chapter <788> Particulate Matter in Injections. USP–NF. United States Pharmacopeia: Rockville, MD, 2024.
2 Vogel JD, et al. Measurement of Particulates in Single-Use Systems for Cell and Gene Therapies Manufacturing, Part 1: Misapplication of USP <788>. BioProcess Int. 21(6) 2023: 18–21; https://www.bioprocessintl.com/cell-line-development/measurement-of-particulates-in-single-use-systems-for-cell-and-gene-therapies-manufacturing-part-1-misapplication-of-usp-788-.
3 USP General Chapter <1788.1> Light Obscuration Method for the Determination of Subvisible Particulate Matter. USP–NF. United States Pharmacopeia: Rockville, MD, 2024.
4 Barber TA. Control of Particulate Contamination in Healthcare Manufacturing. CRC Press: Boca Raton, FL, 1999; https://doi.org/10.1201/9780429246692.
5 USP General Chapter <790> Visible Particulates in Injections. USP–NF. United States Pharmacopeia: Rockville, MD, 2024.
6 EP 2.9.20. Particulate Contamination: Visible Particles. European Pharmacopoeia 6.0, 2008; http://www.uspbpep.com/ep60/2.9.20. particulate contamination- visible particles 20920e.pdf.
7 JP 6.06. Foreign Insoluble Matter Test for Injections. Japanese Pharmacopoeia June 2021; https://www.mhlw.go.jp/stf/seisakunitsuite/bunya/0000066597.html.
8 USP General Chapter <1790> Visual Inspection of Injections. USP–NF. United States Pharmacopeia: Rockville, MD, 2023; https://doi.org/10.31003/USPNF_M7198_06_01.
9 ICH Q9(R1). Quality Risk Management. International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use: Geneva, Switzerland, 2023; https://database.ich.org/sites/default/files/ICH_Q9%28R1%29_Guideline_Step4_2023_0126_0.pdf.
10 2020 Recommendations for Testing, Evaluation, and Control of Particulates from Single-Use Process Equipment. Bio-Process Systems Alliance: Arlington, VA, 2020; https://www2.bpsalliance.org/forms/store/ProductFormPublic/2020-Recommendations-for-Testing-Evaluation-and-Control-of-Particulates-from-Single-Use-Process-Equipment.
11 CBER/CDER/CVM. Inspection of Injectable Products for Visible Particulates: Draft Guidance for Industry. US Food and Drug Administration: Rockville, MD, 2021; https://www.fda.gov/media/154868/download.
12 Posset T, et al. Good Practice Paper: Visual Inspection of Medicinal Products for Parenteral Use. ECA Visual Inspection Working Group. ECA Academy: Heidelberg, Germany, 2014; https://www.visual-inspection.org/visual-inspection-best-practice.html.
13 Wormuth K, et al. Visible Particulate Matter in Single-Use Bags: From Measurement to Prevention. BioProcess Int. 17(4) 2019: 50–53; https://www.bioprocessintl.com/sponsored-content/visible-particulate-matter-in-single-use-bags-from-measurement-to-prevention.
14 Barnes C, et al. 100% Inspection Does Not Mean 100% Defect Detection. BioPhorum Operations Group: Sheffield, UK, 2023; https://doi.org/10.46220/2022FF002.
15 Wormuth K, et al. Challenges in the Manual Visual Inspection of the Interior Surfaces of Single-Use Systems for the Presence of Particulate Matter. PDA J. Pharm. Sci. Technol. 75(4) 2021: 332–340; https://doi.org/10.5731/pdajpst.2020.012211.
16 Wormuth K, et al. Best Practices To Quantify and Identify Particulate Matter on the Interior Surfaces of Single-Use Systems. PDA J. Pharm. Sci. Technol. 78(1) 2024: 90–99; https://doi.org/10.5731/pdajpst.2022.012755.
17 ASTM E3230-20. Standard Practice for Extraction of Particulate Matter from the Surfaces of Single-Use Components and Assemblies Designed for Use in Biopharmaceutical Manufacturing. ASTM: West Conshohocken, PA, 2023.
Corresponding author Klaus Wormuth, PhD, is a principal scientist at Sartorius Stedim Biotech in Goettingen, Germany ([email protected]). Fanny Gaston, PhD, is a laboratory manager at Sartorius Stedim Biotech in Aubagne, France ([email protected]).
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