Membrane filtration plays a central role in microbial testing and sterility checks across pharmaceutical, biotech, cosmetics, and other regulated industries. These workflows follow USP, EP, and JP pharmacopoeias and often use specific rinsing and diluting solutions (such as fluids A, D, and K) to support the recovery of microorganisms from a wide range of sample types. Additional solutions like PBS, saline, and sugar solutions are also used in some applications, particularly when handling more viscous materials.
In this application note, we evaluated the flow rate performance of our 47 mm ME25 (mixed cellulose ester, 0.45 µm) and Supor™ polyethersulfone (PES, 0.45 and 0.2 µm pore sizes) membrane discs with six test solutions: saline solution, sugar solution, PBS, Fluid A, Fluid D, and Fluid K.
Across the solutions tested, the 0.45 µm Supor™ PES membrane demonstrated the fastest flow rate performance, while the ME25 membrane delivered more consistent flow rates across fluids and volumes. Together, these results show that our ME25 0.45 µm and Supor™ PES membranes are well-suited for membrane filtration in bioburden and sterility testing workflows.
Introduction
Microbial enumeration and sterility testing are essential parts of quality control in pharmaceutical, biotech, cosmetics, and food and beverage industries.
Membrane filtration is an effective and accepted method widely employed to perform this analysis and is carried out in compliance with United States (USP), European (EP) and Japanese (JP) Pharmacopoeias as described in USP <71>(1), <61>(2), EP 2.6.1(3), 2.6.12(4), and JP 4.05(5), 4.06(6).
For these workflows, filtration requires a hydrophilic membrane with a porous structure, typically made from polymers such as polyethersulfone (PES) or mixed cellulose ester (ME), and a pore size of 0.45 µm or smaller with validated microorganism retention. Each membrane disc is individually packed and gamma irradiated to ensure sterility.
Pharmacopeial methods also specify the use of different rinsing and diluting solutions throughout the workflow. Saline solution and phosphate buffered saline (PBS) are used for both microbial enumeration and sterility testing. Fluids A, D and K are specific for sterility testing:
- Fluid A is a sterile washing solution formulated to dilute or dissolve samples without affecting microbial viability.
- Fluid D contains polysorbate 80 and is intended for oily substances, samples containing lecithin, and devices with a “sterile pathway.”
- Fluid K includes peptic digest, beef extract, and polysorbate 80, and is recommended for samples containing petrolatum or oils.
Sugar solution, commonly used in food and beverage testing, was included in this study as a representative viscous and more challenging-to-filter sample type.
In this application note, we assess the flow rate performance of our 47 mm Cytiva ME25 (0.45 µm) and Supor™ PES (0.45 and 0.2 µm) membrane discs with these six different fluids.
Material and methods
All experiments described in the materials and methods section were performed in the Cytiva Harbourgate site (Portsmouth, UK) in 2024 and 2025.
Membrane disc filters
We evaluated the flow rate performance of three sterile Cytiva membrane disc filters with a diameter of 47 mm:
- ME25 mixed cellulose ester membrane, 0.45 µm pore size
- Supor™ polyethersulfone (PES) membrane, 0.45 µm pore size
- Supor™ polyethersulfone (PES) membrane, 0.2 µm pore size
Key characteristics of the membranes are summarized in Table 1.
Table 1. Characteristics of the Cytiva membrane disc filters evaluated
| Membrane disc | |||
|---|---|---|---|
| Membrane type | ME25 | Supor™ PES | Supor™ PES |
| Membrane diameter (mm) | 47 | 47 | 47 |
| Color | White, gridded | White, gridded | White, gridded |
| Sterile | Yes | Yes | Yes |
| Membrane pore size (µm) | 0.45 | 0.45 | 0.2 |
Test solutions
We assessed flow rate using six solutions commonly used in sterility testing and microbial enumeration workflows. The composition and suppliers of each solution are listed in Table 2.
Table 2. Solutions used to evaluate flow rate for ME25 and Supor™ membrane disc filters
| Solution | Composition | Supplier |
|---|---|---|
| Sugar solution (0.95% w/v) | 9.5 g/L raw cane sugar in R.O. water | Whole Food Earth |
| Saline solution (1% w/v NaCl) | 10 g/L NaCl | VWR |
| PBS | PBS tablet dissolved in sterile water to prepare 1× solution | VWR |
| Fluid A | 1 g / L peptic digest of animal tissue;
pH 7.1 ± 0.2 |
Oxoid |
| Fluid D | Fluid A (1 L) with polysorbate 80 (1 mL) | Oxoid/Thermo |
| Fluid K | 5 g peptic digest of animal tissue, 3 g beef extract, 1 mL polysorbate 80 | Oxoid/Thermo/VWR |
Fluid A, D, and K were prepared at a concentration of 10× and autoclaved in accordance with USP <62> and <71> guidance. Prior to testing, solutions were diluted with RO water to the final working concentration.
Flow rate testing setup and conditions
To measure flow rate, each membrane disc was placed in a 500 mL Cytiva magnetic filter funnel and mounted on our three-position vacuum manifold (product code 4889). Filtration was performed at a constant vacuum of 20 inHg and at a controlled temperature of 20°C to 21°C.
For each solution, we evaluated two filtration volumes: 100 and 300 mL. The time required for each solution to pass through the membrane disc was recorded and used to calculate flow rate (mL/min).
A total of 12 membrane discs of each membrane type were tested per solution to ensure reproducibility.
Results and discussion
This study evaluated the flow rate performance of six different solutions through Cytiva membrane disc filters, including the 0.45 µm ME25, 0.2 µm Supor™ PES, and 0.45 µm Supor™ PES. Table 3 summarizes the complete dataset, while Fig 1 and Fig 2 shows the flow rate performance when filtering 100 and 300 mL of six test solutions, respectively.
Table 3. Flow rate (mL/min) values and standard deviation (SD) for Cytiva membrane disc filters obtained when filtering 100 and 300 mL of six test solutions
| Membrane disc | ||||
|---|---|---|---|---|
| Fluid | Volume | ME25 0.45 µm | Supor™ 0.45 µm | Supor™ 0.2 µm |
| Sugar solution
(0.95% w/v) |
100 mL | 193 ± 16 | 293 ± 17 | 125 ± 5 |
| 300 mL | 151 ± 13 | 186 ± 25 | 92 ± 7 | |
| Saline solution
(1%) |
100 mL | 206 ± 17 | 324 ± 34 | 153 ± 6 |
| 300 mL | 218 ± 27 | 349 ± 40 | 160 ± 9 | |
| PBS | 100 mL | 223 ± 12 | 339 ± 13 | 145 ± 9 |
| 300 mL | 228 ± 24 | 378 ± 14 | 155 ± 9 | |
| Fluid A | 100 mL | 212 ± 11 | 336 ± 14 | 158 ± 3 |
| 300 mL | 228 ±10 | 366 ± 11 | 167 ± 4 | |
| Fluid D | 100 mL | 216 ± 13 | 242 ± 15 | 101 ± 3 |
| 300 mL | 219 ± 15 | 216. ± 10 | 88 ± 4 | |
| Fluid K | 100 mL | 25 ± 10 | 248 ± 19 | 68 ± 14 |
| 300 mL | 179 ± 6 | 155 ± 12 | 22 ± 6 | |
Flow rate of 100 mL test solutions with ME25 and Supor™ PES membranes discs
We observed the highest flow rates when filtering the test solutions through 0.45 µm Supor™ PES membrane discs (Fig 1). Flow rates for saline solution, PBS, and Fluid A were comparable, with values of 324 ± 34 mL/min, 339 ± 13 mL/min and 335 ± 14 mL/min, respectively.
The flow rate obtained with sugar solution (0.95% w/v) was slightly lower at 293 ± 17 mL/min. The lowest flow rates for this membrane were recorded with Fluid D and Fluid K, at 242 ± 15 mL/min and 248 ± 19 mL/min, respectively.
Differences in flow rates for the 0.45 µm Supor™ PES membrane are likely influenced by the chemical properties of the test solutions. The reduced flow rate measured with sugar solution is consistent with its higher viscosity relative to PBS and Fluid A. In contrast, Fluids D and K contain surfactants that may influence membrane permeability and solution flow dynamics.
The 0.45 µm ME25 membrane discs showed intermediate flow rates across all six test solutions, with values consistently close to 200 mL/min (Fig. 1). The highest flow rate for this membrane was observed with Fluid K (225 ± 10 mL/min), while the lowest was measured with sugar solution (193 ± 16 mL/min). Compared with the 0.45 µm Supor™ PES membrane, the ME25 membrane demonstrated lower variability in flow rate across different solution types, indicating more uniform performance regardless of fluid composition.
As expected, the 0.2 µm Supor™ PES membrane discs produced the lowest flow rates among the membranes evaluated (Fig 1), reflecting the smaller pore size. Flow rates for saline solution, PBS, and Fluid A were similar, at 153 ± 6 mL/min, 145 ± 9 mL/min, and 158 ± 3 mL/min, respectively.
In comparison, sugar solution filtered more slowly, with an average flow rate of 125 ± 5 mL/min. The lowest flow rates for this membrane were recorded with Fluid D and Fluid K, at 101 ± 3 mL/min and 68 ± 14 mL/min, respectively.
Our results indicate that while pore size affects absolute flow rate, solution-dependent trends remain consistent across Supor™ PES membranes.
Fig 1. Flow rate comparison of Cytiva membrane disc filters using 100 mL of test solutions. Each bar represents the mean flow rate obtained from 12 replicate membrane discs.
Flow rate performance with 300 mL test solutions
Similar trends were observed when filtering 300 mL of the test solutions (Fig 2). The highest flow rates were again measured with the 0.45 µm Supor™ PES membrane discs when filtering saline solution, PBS, and Fluid A, with values of 349 ± 40 mL/min, 378 ± 14 mL/min, and 366 ± 11 mL/min, respectively. These results are consistent with those obtained at the 100 mL volume.
For the sugar solution, the 0.45 µm Supor™ PES membrane showed a lower flow rate compared with the other test solutions, and a decrease in flow rate was observed as the filtered volume increased. Flow rates of 293 ± 17 mL/min and 186 ± 25 mL/min were measured when filtering 100 mL and 300 mL, respectively. A similar reduction in flow rate with increasing filtered volume was observed for Fluids D and K (Table 3).
As with the 100 mL study, the 0.45 µm ME25 membrane demonstrated intermediate flow rates when filtering 300 mL. Flow rates for saline solution, PBS, Fluid A, and Fluid D ranged from 218 ± 27 mL/min to 228 ± 10 mL/min. The lowest flow rate for this membrane was observed with sugar solution, at 151 ± 13 mL/min, which may be attributed to the higher viscosity of this solution over longer filtration times.
Overall, we found that increasing the filtered volume of lower viscosity solutions (saline solution, PBS, and Fluid A) resulted in higher flow rates for Supor™ PES membranes, whereas flow rates for the 0.45 µm ME25 membrane remained more consistent.
These differences may be influenced by membrane structure, including asymmetry and pore architecture, as well as interactions between the membrane material and solution chemistry.
Notably, when filtering 300 mL of Fluid K, the 0.45 µm Supor™ PES membrane exhibited a lower flow rate than the 0.45 µm ME25 membrane. This behaviour may be associated with the higher surfactant concentration in Fluid K, which may interact more strongly with the Supor™ PES membrane.
Across all solutions and volumes tested, the slowest flow rates were consistently observed with the 0.2 µm Supor™ PES membrane, as expected based on its smaller pore size.
Fig 2. Flow rate comparison of Cytiva membrane disc filters using 300 mL of test solutions. Each bar represents the mean flow rate obtained from 12 replicate membrane discs.
Conclusion
This study evaluated the flow rate performance of Cytiva ME25 and Supor™ PES membrane disc filters using solutions commonly applied in bioburden and sterility testing workflows. The results show that membrane material, pore size, solution composition, and filtered volume all influence filtration performance.
Key findings include:
- The 0.45 µm Supor™ PES membrane delivered the highest flow rates across most test solutions, particularly with lower viscosity fluids such as saline solution, PBS, and Fluid A.
- Flow rate performance of the 0.45 µm Supor™ PES membrane varied with solution type and filtered volume, especially for more viscous or surfactant-containing fluids.
- The 0.45 µm ME25 membrane demonstrated more consistent flow rates across all solutions and volumes tested, indicating stable performance regardless of sample composition.
- As expected, the 0.2 µm Supor™ PES membrane showed lower flow rates than the 0.45 µm membranes due to its smaller pore size, while maintaining similar solution-dependent trends.
Together, these results demonstrate that our ME25 0.45 µm and Supor™ PES membranes (0.45 µm and 0.2 µm) provide reliable and robust options for membrane filtration in bioburden and sterility testing applications, supporting informed membrane selection based on workflow requirements.
Frequently asked questions
What is the purpose of this study?
This study evaluated the flow rate performance of Cytiva ME25 and Supor™ PES membrane disc filters when filtering a range of solutions commonly used in bioburden and sterility testing workflows. The goal was to understand how membrane material, pore size, solution composition, and filtered volume influence filtration performance.
Which membranes were evaluated in this application note?
The study assessed three Cytiva membrane disc filters with a diameter of 47 mm:
- ME25 mixed cellulose ester membrane, 0.45 µm pore size
- Supor™ PES membrane, 0.45 µm pore size
- Supor™ PES membrane, 0.2 µm pore size
All membrane discs were sterile, individually packed, and gamma irradiated.
Which test solutions were used, and why were they selected?
Six solutions were selected based on their common use in regulated microbiological testing:
- Saline solution and PBS, used in both microbial enumeration and sterility testing
- Fluids A, D, and K, specified in pharmacopeial sterility testing methods for different sample types
- Sugar solution, included as a representative viscous and more challenging to filter sample, commonly encountered in food and beverage testing
How does membrane material affect flow rate performance?
The results show that membrane material influences flow rate behaviour, particularly when filtering viscous or surfactant containing solutions. In this study, the Supor™ PES membranes delivered higher flow rates overall, while the ME25 membrane showed more consistent performance across different fluids and filtered volumes.
Why does filtered volume affect flow rate?
Flow rate can change as the filtered volume increases due to interactions between the solution and the membrane surface, including viscosity effects and potential fouling or wetting behaviour. In this study, increasing filtered volume had a greater impact on Supor™ PES membranes than on the ME25 membrane, particularly for higher viscosity solutions.
How does pore size influence filtration performance?
Smaller pore sizes generally result in lower flow rates due to increased flow resistance. As expected, the 0.2 µm Supor™ PES membrane showed lower flow rates than the 0.45 µm membranes across all test solutions, while maintaining similar solution dependent trends.
Which membrane should I choose for my sterility or bioburden testing workflow?
Membrane selection depends on your application requirements:
- Choose Supor™ PES membranes when higher flow rates are a priority, particularly with lower viscosity solutions.
- Choose ME25 membranes when consistent flow performance across a wide range of solution types is required.
- Select 0.2 µm membranes when smaller pore size is needed for retention requirements, recognizing the associated reduction in flow rate.
Are these results applicable to pharmacopeial methods?
Yes. The test solutions and workflows evaluated in this study align with those described in USP, EP, and JP pharmacopeial guidance for sterility testing and microbial enumeration. However, users should always follow their site specific procedures and applicable regulatory requirements.
Where can I find more information about Cytiva membrane filters?
Additional information on Cytiva membrane filters, including specifications and application guidance, is available on cytiva.com and in related Cytiva application notes and technical resources.
REFERENCES
- United States Pharmacopeial Convention. United States Pharmacopeia and National Formulary (USP 43–NF 38). Chapter <71>: Sterility Tests. Rockville, MD: United States Pharmacopeial Convention; 2025. Available at: http://ftp.uspbpep.com/v29240/usp29nf24s0_c71.html
- United States Pharmacopeial Convention. United States Pharmacopeia and National Formulary (USP 43–NF 38). Chapter <61>: Microbiological Examination of Nonsterile Products—Microbial Enumeration Tests. Rockville, MD: United States Pharmacopeial Convention; 2025. Available at: https://www.usp.org/harmonization-standards/pdg/general-methods/microbial-enumeration
- European Directorate for the Quality of Medicines & Healthcare (EDQM). European Pharmacopoeia. Chapter 2.6.1: Sterility. Strasbourg, France: EDQM. Available at: EDQM Microbiology Chapters PDF.
- European Directorate for the Quality of Medicines & Healthcare (EDQM). European Pharmacopoeia. Chapter 2.6.12: Microbiological Examination of Non Sterile Products—Microbial Enumeration Tests. Strasbourg, France: EDQM. Available at: EDQM Microbiology Chapters PDF.
- Pharmaceuticals and Medical Devices Agency (PMDA). Japanese Pharmacopoeia, 18th Edition. Chapter 4.06: Sterility Test. Tokyo, Japan: PMDA. Available at: https://www.pmda.go.jp/english/rs-sb-std/standards-development/jp/0029.html
- Pharmaceuticals and Medical Devices Agency (PMDA). Japanese Pharmacopoeia, 18th Edition. Chapter 4.05: Microbiological Examination of Non Sterile Products. Tokyo, Japan: PMDA. Available at: https://www.pmda.go.jp/files/000152672.pdf