Whitepaper

The Role of BioPhorum

Extractables Data in the Effective
Adoption of Single-Use Systems
Authors: Satish Kumar Mohanvelu, Marketing Manager, Emprove® Program,
MilliporeSigma, Jessica Shea, Global Technical Support, MilliporeSigma

The landscape for times and decrease the risk of and slip agents, pigments and
single-use systems cross-contamination. Further antioxidants such as BHT may
benefits include more dependable either leach out of the system or
Single-use systems are increas­ lead times and faster delivery be extracted by solvents used in
ingly popular in biomanufacturing. of higher-quality, lower-cost the manufacturing process. For
Their adoption offers numerous products. patient safety, biomanufacturers
advantages for greater efficiency must systematically assess and
and productivity in applications Despite these advantages, single- mitigate the risks posed by any
such as final filtration, mixing use systems can also contain extractables and leachables (E&L)
and aseptic connections. Single- risk. Polymeric materials in in these systems. Unfortunately,
use systems can reduce capital single-use systems can introduce the burden of obtaining E&L data
investment in facilities and a range of unwanted chemicals and assessing and mitigating any
equipment, eliminate the need into the manufacturing process risk found is the top reason some
for cleaning procedures and their fluid. Substances such as styrene researchers restrict their use of
required validation, reduce startup monomers, stabilizers, lubricants single-use systems.

Guidance for evaluating SUS for data. If the risk of maximum dosage of potential
extractables and leachables leachables remains, leachable evaluation and testing
may be necessary. Furthermore, if product quality
Although formal guidelines for E&L assessments could be affected by a potential leachable, studies
have not yet been enacted, there is nonetheless a may need to assess the effect on product quality,
regulatory expectation that researchers will test for including efficacy. These evaluations are possible
these potentially harmful contaminants. Agencies when the supplier provides extractables data, which
such as the USFDA’s Center for Biologics Evaluation can supplement final product quality assessments.
and Research (CBER) recommend a risk-based
approach to evaluation. In such an approach, Beyond the regulatory sphere, numerous industry
indication, safety, product characteristics, dosage, organizations have created best-practice strategies
formulation and stability are all factors. for implementing extractables studies. Prominent
among these coalitions is BioPhorum formerly known
If there appears to be lower risk with the materials as BioPhorum Operations Group (BPOG), a global
in question, the sponsor can submit supplier data, consortium of large biopharmaceutical manufacturers.
a detailed justification for applying this data and In the absence of official E&L-specific guidance, many
an explanation of why no more testing is required. manufacturers are fulfilling regulatory expectations
If there is relevant risk, the sponsor may have by following the BioPhorum and/or U.S. Pharmacopeia
to determine toxicity based on maximum dosage (USP) recommendations. An overview of these standard
of potential leachables derived from extractables extractables testing protocols is shown in Table 1.

The life science business of Merck operates as

MilliporeSigma in the U.S. and Canada.
Table 1: Comparison of BioPhorum and USP protocols for extractables testing

BioPhorum Requirements USP <665> draft (May 2019)

Scope Single-use components in contact with fluid Single-use and multi-use components and devices with fluid path
path (for biopharmaceutical manufacturing) contact (for pharmaceutical and biopharmaceutical manufacturing)
Solvents 1. 50% Ethanol 1. 50% Ethanol
2. 0.5N NaOH 2. 0.2M KCI, pH 3
3. 0.1M Phosphoric Acid 3. 0.1M Phosphate buffer, pH 10
4. WFI
Analytical methods HPLC-DAD / MS (APCI, ESI, +/–) NVR Described in USP <1663>. Broader scope in methods selection.
ICP / MS TOC
DI-GC / MS, HS-GC / MS pH
Time points 1–3, dependent on component 1, dependent on component
Pre-treatment If an item is pre-treated prior to actual use, Polymeric components are most appropriately tested when
the item should be pre-treated exactly the they have been conditioned or processed in a manner consistent
same way before being tested for extractables. with their intended use and as specified in the manufacturer’s
­instructions for use.
timing Revised BioPhorum Protocol published in Second draft comments phase completed in May 2019
Apr 2020

Note that while these protocols are not identical, analyses to determine what levels of volatile, semi-
the general idea is similar and at least one or volatile and nonvolatile organic compounds, and
the other can apply to most drug products and metals, have been extracted. Either approach will
substances. Testing is performed at various time produce an array of data that must then be evaluated
points and temperatures. Resulting extraction in the context of the proposed process.
solutions are subjected to robust and extensive

Supplier-provided single-use system extractables data saves time and resources

The task of gathering all the product information required to assess risk and optimize E&L test strategies for
a process can be daunting. Consider the filtration setup shown in Figure 1. This one assembly exposes the
contact fluid to multiple components as indicated.

10
FLOW

4
15 31 20 30 13 30 21 29 12 29 21 30 13 30 20 31 15 31 1 29
31 16 31 19 31 17 28

30 31 8
29 14 23
18
2 31 26
6 27
26
30
11 11
29 29
5 5
STOP!
3

23
3 9
8
9 22
5

Figure 1. S
 chematic of a 32-component Mobius® filtration assembly containing various types of single-use filters and bags

A proper E&L assessment must account for the risk Finally, sample analysis yields an extractables profile
imposed by each component in the fluid path. Once quantifying all volatile, semi-volatile and nonvolatile
each component’s materials of construction (MOC), organic compounds found, as well as elemental
resin identity and manufacturing process have been impurities.
documented, similar components may be grouped This complex process is much simpler when the
together for analysis. Two to three lots of worst-case supplier provides comprehensive, organized datasets
representative samples are extracted as per the for its single-use components, such as filters and
chosen testing protocol, under worst-case conditions. bags.
2
The overall strategy for E&L validation

Determining risk and interpreting extractables data to construct a reasonable validation strategy is a
complex process; expert assistance can be valuable. The general steps in evaluating each single-use
component are as outlined in Figure 2.

Assess the Risk

Assess the Risk Re-assess the Risk
•D
 efine the process –
to Patient Safety to Patient Safety
feed formulation,
filtration and process • Patient safety • Update ­patient
conditions ­evaluation safety ­evaluation

A B

Identify and Identify and ­Quantity Regulatory

Quantity the individually the ­Submission
Extractables ­Leachables
• Extractables • ­Leachables tests
data
• ­Flushing / risk
­mitigation
Figure 2. Risk-based approach to E&L assessment

In general, drug manufacturers must evaluate the the other hand, if there is a risk to the patient, a
single-use components that are in product contact leachables study may be conducted under normal
under the given process conditions such as duration, product application or storage conditions. Risk
temperature, solvents, material characteristics, may also be mitigated through a process-step
etc. Any risk-mitigating steps occurring later in the modification, such as including an additional flush.
process can be taken into consideration. At this A final option could be to change the material of
stage, an extractables profile, if available, would the component.
facilitate product, process and dosage-specific
assessment. If, thus far, the patient safety evaluation The following case study demonstrates this approach
indicates no risk, the findings may simply be to risk assessment.
reported and monitored for future changes. On

Case study: A risk-based approach to E&L assessment

of a final sterilization, single-use assembly

Figure 3. Monoclonal antibody final sterilization system including a single-use

Millipore Express® SHC filter and a PureFlex™ Plus Mobius® single-use bag

3
Figure 3 illustrates a representative, single-use Extractables test data prepared by the single-use
system for which E&L assessment might be required: system component manufacturer can be used to
a single-use final sterilization assembly including analyze this system. The availability of such data
a capsule filter and a product collection bag. This in an easy-access format saves significant time
assembly serves a commercial monoclonal antibody and effort. If these components were evaluated in
(mAb) application with typical excipients. The accordance with BPOG guidelines, they were tested
aqueous drug product solution includes 2% API, under worst-case conditions, as shown in Table 2.
0.02% PS80 surfactant and 2% dextrose, buffered
to a pH of 5. Filter contact time is eight hours with
a filtration temperature of 25 °C.

Table 2. BioPhorum conditions for extractables testing of the single-use components in the mAb system

Gamma-compatible Millipore Express® SHC Filter PureFlex™ Plus Mobius® Single-Use Bags

• Three different lots • Three different lots

• Gamma-irradiated at 45–55 kGy • Gamma-irradiated at 45–55 kGy
• Two time points at 1 day/7 days • Three time points at 1 day/21 day/70 days
• Temparature 40 °C • Temperature 40 °C
• SA/V = 4:1 • SA/V = 6:1

Summary results of this testing are shown in Figures 4 and 5. The data includes numerous compounds that
were produced through extraction using the solvents listed. Darker dots indicate compounds found in the
highest concentrations.

Component 1
Extractables Data for Gamma-Sterilized Millipore Express® SHC Filter
32 Individual Compounds Identified

WFI • • • • •

0.1M H3PO4 • • • • •

50% EtOH • • • • • • • • • • • • • • • • • • • • • •

• Highest Concentrations

• Below highest Concentrations

Individual Quantitation
Compound Evaluation

WFI • • • • •

0.1M H3PO4 • • • • •

50% EtOH • • • • • • • • • • • • • • • • • • • • • •

Figure 4. Summary of extractables test results for the single-use filter.

The highlighted compounds are present in multiple components.

4
Component 2
Extractables Data for PureFlex™ Plus Film
22 Individual Compounds Identified

WFI • • •

0.1M H3PO4 •

0.5N NaOH • • • • •

50% EtOH • • • • • • • • • • • • • •

• Highest Concentrations

• Below highest Concentrations

Individual Quantitation
Compound Evaluation

WFI • • •

0.1M H3PO4 •

50% EtOH • • • • • • • • • • • • • • •

Figure 5. Summary of extractables test results for the film used to make the single-use bag.
The highlighted compounds are present in multiple components.

Water (WFI), phosphoric acid (0.1 M H2PO4) and Similarly, the extractables data for the bag must be
ethanol (50% EtOH) are most applicable to this scaled, as it is reported per cm2. The 10L bag has
process based on the pH and formulation of the a surface area of 3,077 cm2, so the bag’s contribution
drug product. Hence, the focus is on the compounds is:
extracted by these solvents. One observation is
that the two compounds highlighted – 2,4-di-tert- (2.81 µg/cm2) 3,077 cm2 = 8,646 µg
butylphenol and 1,3-di-tert-butylbenzene – are
common degradants and appear in the extraction
data for both the filter and the single-use bag. In Note that the bag contributes the majority of the
such cases, once the single-use system contributions total 2,4-di-tert-butylphenol, which is 8,799 µg/
have been scaled to the actual system in question, assembly.
it is the sum of these contributions that must be
evaluated for toxicity. The next consideration is, how much of this will
be delivered to patients? If the minimum process
To illustrate this calculation, here is the analysis volume is 4L, and a 1L flush is added, the total
of 2,4-di-tert-butylphenol. The first step is to volume is 5L. The final concentration of 2,4-di-tert-
scale the data for each component’s contribution. butylphenol is:
Filter data is provided as 0.36 mg for a 10” filter,
which has a surface area of 0.54 m2. However, the 8,799 µg/5,000 mL = 1.76 µg/mL
5” device in use has a surface area of 0.23 m2.
The proportionately scaled contribution of this
compound by the filter, then, is:

(0.23/0.54) 0.36 mg = 0.153 mg = 153 µg

5
Aspects of posology such as dosage, frequency and route of administration are then used to calculate the
actual patient exposure, as shown in Figure 6.

Patient Exposure
Posology Concentration
of 2,4-di-tert-­ Drug dosage
butylphenol = = 100 mL
1.76 µg/mL

Maximum Drug Dosage 100 mL

Route of Administration Intravenous Compound-
Quantity above the
Yes specific
Frequency Once per week TTC (ICH M7)
176 µg/person/ evaluation
Expected Duration Lifetime day of 2,4-di-
tert-­butylphenol

No

Risk is acceptable

Figure 6. Patient exposure as a function of posology

The result is then assessed for risk. If the risk were do that, results from pre-existing rat toxicology
acceptable in reference to the threshold of studies are converted to equivalent human values
toxicological concern (TTC), that finding could be using the ICH Q3C modifying factors for relating the
reported and no further investigation would be data to humans, F1–F5. An additional bioavailability
required. However, in this case, the exposure correction accounts for the incomplete absorption
exceeds the TTC. Further analysis is needed to seen when drugs are administered orally, as they
determine the compound-specific permitted daily were in the rat study. The overall approach is shown
exposure (PDE) for this particular compound. To in Table 3.

Table 3. Converting the result of an animal study of 2,4-di-tert-butylphenol

toxicity to a compound-specific PDE for humans

Factors: Starting No-observed-

dosage: adverse-effect level
75 mg/kg (NOAEL) when orally
bw/day administered to rats
Interspecies extrapolation F1 5 Rats -> humans
Intraspecies variability F2 10 Default
Exposure duration F3 10 4 weeks -> chronic
Severity of effects F4 1 No severe effects
LO(A)EL -> NO(A)EL F5 1 NOAEL available
Bioavailability correction F6 10 Oral -> parenteral, no data
Overall safety factors 5,000
Permitted daily exposure (PDE) 75 mg/kg bw/day / 5,000 = 15 µg/kg bw/day
PDE for 50 kg adult human 750 µg/day

6
The newly calculated PDE for 2,4-di-tert-butylphenol reported as such. If the risk were still too high, a
– 750 µg/day – can now be compared to the total method for mitigating risk elsewhere in the process
exposure calculated above – 176 µg/day. The risk would need to be applied. The overall process of risk
has now been shown to be acceptable and can be evaluation is summarized in Figure 7.
Acceptable Risk
Patient Safety

176 µg/person/ Quantity above the Quantity above

day of 2,4-di- Yes Risk is acceptable No Risk is acceptable
TTC (ICH M7) The PDE?
tert-­butylphenol

No Yes

Risk Mitigation
Risk is acceptable
Leachables?
Dilution?
Process Change?
Figure 7. E&L risk evaluation decision tree

Summary qualifications, risk assessments and process

optimization. E&L risk analysis is a complex process,
Despite the current absence of guidance, and expert guidance is helpful to ensure compliance
regulatory expectation does require patient safety and drug safety. While the task of analytical E&L test
evaluations with supporting data for manufacturing data interpretation and submission can be daunting,
components that directly come into contact with the right single-use system supplier can save time
drug manufacturing process streams. Readily by providing single-use system E&L data in well-
available extractables data can help manufacturers organized, easy-to-use formats.
using single-use technology to accelerate product

THE EMPROVE® PROGRAM starting materials. Material Qualification Dossiers

report on a material’s fundamental properties. They
The Emprove® Program for Accelerating also feature a manufacturing flow chart, product
Drug Regulatory Approvals characterization, regulatory statements and more.
Quality Management Dossiers include a quality
The Emprove® Program at MilliporeSigma entails self-assessment while documenting detailed supply
organized, detailed product information and information for the material, including its complete
assistance to help customers with material chain-of-custody from manufacture to final release.
qualification and risk assessment of biomanufacturing These dossiers present additional useful information
devices, as well as process optimization. By avoiding such as shelf-life data, sterilization validation and
the need to develop test methods, search quality- packaging requirements. Operational Excellence
related documents or evaluate scientific data, Dossiers include product quality reports with
customers can reduce the time, effort and resources elemental impurities information and extractables
needed to prepare documents for drug approval profiles, and specify the analytical procedures
submissions. used to obtain this data. Convenient 24/7 online
document access and customer support ensure
This program offers three categories of thoughtfully that biopharmaceutical investigators may take full
organized, comprehensive dossiers for our filters and advantage of the program any time.
single-use components as well as pharma raw and

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Lit. No. MK_WP3014EN 05/2020