Integrity Testing Methods

Integrity testing sterilizing filters is a fundamental requirement of critical process filtration applications in the pharmaceutical industry. FDA Guidelines require integrity testing of filters used in the processing of sterile solutions such as large volume paren terals (LVPs) and small volume parenterals (SVPs). The FDA also requires corresponding testing documentation be included with batch product records.

Two classifications of integrity testing are destructive and non-destructive. Millipore's practice is to perform destructive testing as a lot release criteria on samples from each manufacturing lot of all fabricated sterilizing-grade filter products, and non-destructive testing on each sterilizing-grade filter prior to sale to insure its integrity.

Destructive Testing

Millipore performs destructive bacterial challenge testing in accordance with ASTM F838-83 methodology. Destructive challenge testing is the best way to determine a sterilizing filter's ability to retain bacteria. Bacterial challenge testing provides assurance that the membrane and fabricated device meet the critical performance criteria of a sterilizing filter. The test is performed on a statistical sample of each lot of membrane and fabricated devices produced.

During the Millipore bacterial retention test, 0.22 µm filter discs and devices are challenged with a solution of culture medium containing bacteria (Brevundimonas diminuta ATCC 19146) at a minimum challenge of 10⁷ per cm². The effluent is then passed through a second 0.45 µm assay filter disc that is placed on an agar plate and incubated.

Non-Destructive Testing

Non-destructive testing may be done on filters before and after use. Integrity testing sterilizing filters before use monitors filter integrity prior to batch processing, preventing use of a non-integral filter.
Integrity testing sterilizing filters after a batch has been filtered can detect if the integrity of the filter has been compromised during the process. Detecting a failed filter alerts operators to a problem immediately after batch processing, eliminating delay and allowing rapid reprocessing.

There are three types of non-destructive testing – the bubble point test, the diffusion test, and the waterflow integrity test for hydrophobic filters (HydroCorr™ Test). The pressure hold, forward flow, and pressure decay tests are variations of the diffusion test. The stringent requirements of the pharmaceutical industry dictate that non-destructive filter integrity testing must be performed in each sterilizing application.

To be able to use an in-process non-destructive integrity test, physical tests were developed that correlate to the bacterial challenge test. A specification for the physical test correlates directly to the bacterial challenge test. Once this correlation is established, it is determined that a cartridge passing the physical test is an integral sterilizing filter.

Bubble Point Test

The most widely used non-destructive integrity test is the bubble point test. Bubble point is based on the fact that liquid is held in the pores of the filter by surface tension and capillary forces. The minimum pressure required to force liquid out of the pores is a measure of the pore diameter (see formula).

P = bubble point pressure
d = pore diameter
k = shape correction factor
cos Θ = liquid-solid contact angle
Σ = surface tension

Bubble Point Procedure

1. Wet the filter with the appropriate fluid, typically water for hydrophilic membranes or an alcohol/water mixture for hydrophobic membranes.

2. Pressurize the system to about 80% of the expected bubble point pressure which is stated in the manufacturer's literature.

3. Slowly increase the pressure until rapid continuous bubbling is observed at the outlet.

4. A bubble point value lower than the specification is an indication of one of the following:
- fluid with different surface tension than the recommended test fluid
- integral filter, but wrong pore size
- high temperature
- incompletely wetted membrane
- non-integral membrane or seal

Diffusion Test

At differential gas pressures below the bubble point, gas molecules migrate through the water-filled pores of a wetted membrane following Fick's Law of Diffusion. The gas diffusional flow rate for a filter is proportional to the differential pressure and the total surface area of the filter. At a pressure approximately 80% of the minimum bubble point, the gas which diffuses through the membrane is measured to determine a filter's integrity. The flow of gas is very low in small area filters, but it is significant in large area filters. Maximum diffusional flow specifications have been determined for specific membranes and devices and are used to predict bacterial retention test results.

Diffusion Test Procedure

1. Thoroughly wet the filter with appropriate test fluid, typically water for hydrophilic membranes or an alcohol/water mixture for hydrophobic membranes.

2. Slowly increase pressure on the upstream side of the filter to the recommended test pressure provided by the manufacturer, typically at least 80% of the minimum bubble point specification.

3. Allow the system to equilibrate.

4. Measure the gas flow at the outlet for one minute with an inverted graduated cylinder or a flow meter.

5. A diffusional flow reading higher than the specification is an indication of one of the following:
- wrong pore size
- temperature other than ambient
- incompletely wetted membrane
- non-integral membrane or seal
- liquid/gas combination different than the recommended fluids
- inadequate stabilization time

Pressure Hold Test

The Pressure Hold Test, also known as pressure decay or pressure drop test, is a variation of the diffusion test. In this test, a highly accurate gauge is used to monitor upstream pressure changes due to gas diffusion through the filter. Because there is no need to measure gas flow downstream of the filter, any risk to downstream sterility is eliminated.

The pressure hold value is dependent on the diffusional flow and upstream volume. It can be calculated using the following equation:

Where:
D = Diffusion rate (mL/min)
T = Time (minutes)
Pa = Atmosphere pressure (1 Atm or 14.7 psi)
Vh = Upstream volume of apparatus (mL)
ΔP = Pressure Drop (bar or psi)