Sterilisation by membrane filtration

Question 1

What is the principle of membrane filtration?

Answer 1

Membrane filtration is used for reduction of viable and non-viable particles in gases and fluid products that are not amenable to sterilisation by heat or irradiation. In contrast to other sterilisation methods, the principle of membrane filtration is not inactivation but removal of microorganisms from the product.

Question 2

What makes sterilisation by filtration different from other sterilisation methods e.g. heat sterilisation? How is this accomplished?

Answer 2

Does not involve killing microorganisms or inhibiting their growth

• Living & non-living particles are removed by passage through fibrous, granular or synthetic membrane filters of appropriate retention efficiency.

Question 3

What type of medicaments (active ingredients and products) cannot be terminally sterilised (e.g. by heat sterilisation/irradiation) and may be subjected to a filtration procedure using a filter?

Answer 3

Thermolabile medicaments

Question 4

What are some conditions of sterilisation by filtration

Answer 4

• The production process and environment are designed to minimise microbial contamination and are regularly subjected to appropriate monitoring procedures.
• The equipment, containers and closures and, wherever possible, the ingredients are subjected to an appropriate sterilisation process.
• It is recommended that the filtration process is carried out as close as possible to the filling point. The operations following filtration are carried out under aseptic conditions.
• The sterility and integrity of the equipment downstream from the point of filtration, the qualified environmental conditions and the validated aseptic procedures applied in the handling of the filtered product all contribute to preventing recontamination of the product

Question 5

What are some negatives/positives of sterilisation by filtration

Answer 5

Negatives

• Cold, wet (involves liquid, gas)
• Not as reliable as asepetic transfer
• Not terminal sterilisation
• Thermolabile material must be stable in solution

Positives:

• Aseptic process
• Multi-step processes

Question 6

What are the FOUR steps in sterilisation by filtration?

Answer 6

1. Filtration of the solution through a suitable sterile, bacteria-retaining filter
2. Aseptic distribution of the filtered solution into previously sterilised containers – includes a measurement phase
3. Aseptic closure of the containers
4. Testing of the batch for sterility

Question 7

For prefiltration;

A) Why is it done

B) What type of filter is used

C) Does this pre-filter have to be related to the final filter

D) When is an upstream prefiltration unit favorable

Answer 7

A)

• Potentially extend the working life of final filters
• Retain a large number of the particulates, prevent premature clogging of the final filters

B)

• A type of depth filter is generally used for the purpose

C)

• The performance characteristics of the prefilter should be related to those of the final filter
• In order to ensure an adequate flow rate, the area of the prefilter should be greater than that of the final filter

D)

• When the volume is small

Question 8

For depth filters (pre-filter);

A) What are they made from?

B) What is the pore size relative to the particles removed? What diameter are the channels?

C) What determines the size of particles retained?

D) What conditions are filtration carried out under?

E) What measurement should the pressure differential be?

Answer 8

A)

• Pads containing compacted polypropylene, cellulose or glass fibres.

B)

• The pores are much larger than the particles removed but the channels are of uneven diameter and change direction frequently within the filter bed.

C)

• The maximum pore diameter and retention efficiency of the filter determine the size of particles retained.

D)

• Filtration may be carried out under vacuum or positive pressure

E)

• Uniform pressure is important and the pressure differential should not exceed 35-70 kPa because compression of the matrix would decrease the flow rate
• High dirt holding capacity due to matrix

Question 9

What are disadvantages of depth filters?

Answer 9

• A significant volume of the liquid may be lost by retention in the filter pad
• Components of the liquid (e.g. proteins) may be adsorbed, reducing their concentration in the filtrate
• Filter fibres may be shed into the filtrate: may necessitate rejection of the first portion that passes through the filter or pre-flushing –> may have to discard first portion
• Penetration of microorganisms if the pressure differentials increase (due to decreased flow rate).

Question 10

For screen filter (final filtration step);

A) What are they? How does it relate to the depth pre-filter?

B) What are they also known as?

Answer 10

A)

• Membrane filters or “membranes” are microporous films with specific pore size ratings
• Anything bigger than nominal pore size isn’t going to go through the filter and therefore will accumulate on top and reduce flow rate that’s why pre-filter is used to protect premature clotting of screen filter.

B)

• Also known as screen, sieve, or microporous filters,
• membranes retain particles or microorganisms larger than their pore size primarily by surface capture

Question 11

Complete the following sentence

Where multiple bioburden-reduction filters are used to increase the efficacy of the filtration process, the filter closest to the filling point in the final container is characterised as the………..

Answer 11

sterilising filter

Question 12

How is sterilisation by membrane filtration done?

Clue: relating to pore size

Answer 12

Sterilisation by membrane filtration is performed by passage of the product through a microporous membrane with a nominal pore size not greater than 0.22 µm

Question 13

For membrane filters;

A) What is their thickness

B) What is their regularly space spore size

C) How do they differ from depth filters?

D) how do they become clogged

E) What does air locking of filters mean

Answer 13

A)

• 125-150 um in thickness

B)

• Regularly spaced pores 0.22 µm GS; 0.20 µm OR available as discs ranging from 13 to 293 mm in diameter

C)

• Low dirt-handling capacity compared with fibrous depth filters

D)

• Easily clogged by particles just larger than pore diameter

E)

• after wetting the membrane filter, if there is any air, air may not be able to go through, the liquid cannot go through due to air.

Question 14

Advantages of membrane filters

Answer 14

• All particles larger than the pore diameter are retained and bacteria cannot pass through
• A large open volume (80% of the filter) ensures an adequate flow rate
• The volume of liquid retained in the membrane is small and can be expelled without breaking sterility compared to depth filter
• The quality of the filtrate is not altered by adsorption of solutes or contamination with foreign material = no shedding of fibres

Question 15

What are the types of membrane filters?

Answer 15

• Hydrophilic filters = Cellulose esters (acetate and nitrate)
• Surfactant-free filters
• Low protein binding filters –> use this when dealing with peptides (biotech product)
• Hydrophobic filters

> vacuum line protection, sterilising gases, venting sterile containers, and sterilising or clarifying organic solutions

> Polytetrafluoroethylene (PTFE), polyvinyl chloride and polyvinylidene difluoride (PVDF)

Question 16

When are hydrophilic filters with hydrophobic edge membranes/rims used? How do they work?

Answer 16

• Sterility testing of liquids containing antimicrobial agents
• Hydrophobic edge membranes are standard MF-Type filters with edges rendered hydrophobic to 3 or 6 mm widths –> solution may get trapped at edge and cannot be washed therefore cannot be washed and get rid of bacteriostatic material in product
• In antibiotic and bacteriostatic drug sterility testing by membrane filter method, the 6 mm wide non-wetting edge prevents drug intrusion under holder sealing rim –> all the solution goes in the middle section and nothing is trapped

Question 17

How to setup vacuum filter holders and pumps?

Answer 17

Vacuum operated unit

• All have to be sterile
• Product in the top, collect filtrate at the bottom

Question 18

How to setup pressure filter holders?

Answer 18

Pressure operated united

• Sealed tank
• Collect in a sterile vessel
• Supply-side is closed to apply pressure, delivery side is open

Question 19

What are the advantages of prefabricated units used in sterilisation by membrane filtration?

Answer 19

• Consistent „
• Convenient „
• Surfactant free „
• Disposable „
• Filtration areas „
• Bidirectional support „
• Inlet Luer-Lock „
• Outlet Luer-Slip

Question 20

What are the integrity tests used in membrane filtration

Answer 20

The integrity of an assembled sterilising filter is verified before use and confirmed after use by carrying out tests appropriate to the type of filter used and the stage of testing, for example bubble-point, pressure hold or diffusion rate tests

• pre-sterilisation microbial contamination, pre-filtration integrity test results, duration of filtration, volume filtered, differential pressure and post-filtration integrity test results.
• Suitable integrity test procedures (e.g. diffusive flow measurement, bubble-point determination or water-intrusion testing) are employed, as recommended by filter manufacturers.

Question 21

For bubble point;

A) What is it?

B) What does the test consist of?

C) What is the procedure?

D) What does a smaller pore size mean for the bubble point?

Answer 21

A)

• 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.

B)

• The integrity of membrane filters can be tested by measuring the bubble point or the diffusional flow of a gas through a wetted filter
• The bubble point, or bubble pressure, is the pressure at which the liquid contained in the channels of the filter is driven out by the test gas.
• Non-destructive test to the filter paper

C)

• Wet the filter with the appropriate fluid, typically water for hydrophilic membranes or an alcohol/water mixture for hydrophobic membranes
• Pressurise the system to about 80% of the expected bubble point pressure which is stated in the manufacturer’s literature.
• Slowly increase the pressure until rapid continuous bubbling is observed at the outlet.

D)

• Smaller pore size = higher bubble point due to higher pressure required to get liquid from the pore.

Question 22

What are the possible reasons when the bubble point value is lower than the specification?

Answer 22

Conduct bubble point test at 0.22 um pore size, bubble point should be 14.5 psi but it is 10 psi

• What are some factors affecting the pressure that is applied to get rid of liquid from the pore?

1. Filter paper is broken
2. Chose wrong filter paper
3. Conducted test at higher temperature, therefore, reduced tension = fluid doesn’t stay in place
4. Check if filter paper is wet completely –> if not wet, air goes through the pore, not water.
5. Use the same fluid when comparing results as there is different surface tension with different liquids

Question 23

SA questions

1. What are the principles of membrane filtration sterilization?
2. Describe the process of membrane filtration sterilisation for an aqueous product
3. Suggest what quality control test should be in place to ensure the process is successful