(8 minute read)
You’ve sanitized your system. You have sterilizing grade filters installed at key points in your process. Your bioburden measurements are low, but you still have endotoxin issues. What’s going on?
How is Endotoxin Created
Endotoxins or lipopolysaccharides (LPS) are released into the environment during active growth, mechanical damage, lysis, and death of gram-negative bacteria cells. Therefore, endotoxins will likely be present in your process stream coming from the original water source. Sterilization processes can release endotoxins in cases where gram-negative bacteria are destroyed. Considering the intrinsic pyrogenicity, endotoxin concentrations need to be reduced to the lowest level possible. Endotoxins are heat stable and persist even after bacterial death. These cell fragments are typically smaller than 0.01 micron, and will not be removed by a 0.22 or even a 0.10-micron sterilizing filter.
Figure 1: Endotoxin in a membrane cell
How is Endotoxin Removed from the Process Stream?
Removal based strictly on size exclusion could be done using a tight Ultra Filtration (UF) process step. This would require high pressure and potentially large surface area filters depending on the required process flow. This would involve modifying the process plumbing to incorporate the UF filter step, and installation of a larger pump to build the required pressure. This would increase both the capital and operating costs of your system.
Fortunately, there is a simpler solution. Critical Process Filtration's charged PES Filter (CPF), a positively charged sterilizing grade filter, can even perform double duty by removing both bacteria and endotoxin from the system. The endotoxins, at pH > 2 are negatively charged. This allows a positively charged filter to remove the endotoxin by adsorption rather than relying solely on the size exclusion.
Figure 2: Positively charged membrane mechanism
Some of the benefits of using the Critical Process Filtration's charged PES Filter are:
- Our charged filter has a Log Endotoxin Removal greater than 4 when challenged with 100 EU/mL of purified coli Type O113:H10 endotoxin.
- Each individual module is integrity tested.
- The cartridges can be sanitized with 1% Minncare and/or 90 °C water.
- The filter materials meet the standards of the United States Pharmacopeia Class VI (USP VI).
- You can expect excellent filtration efficiency, outstanding flow rates, and high throughputs.
- Implementing this technology will require fewer (if any) changes to your process, and will be a far more economical solution.
Endotoxin removal using charged membranes
Capacity measurements were performed to test the flux and endotoxin removal of a Critical Process Filtration's charged PES membrane and comparable units from some of our competitors.
To be able to observe differences in the binding capacities of evaluated membranes, the endotoxin spike was increased to levels that would not normally be observed in biotech manufacturing processes. The tested feed stream contained endotoxin levels of 1000 EU/mL.
The water for dialysis must be ≤0.25 EU/ml. This value was used as a threshold for acceptance.
The plots show the results of the experiments designed to evaluate the efficiency of endotoxin removal by a Critical Process Filtration's charged PES Filter and competitors.
The results show the highest level of endotoxin removal by the Critical Process Filtration's charged PES membrane followed by competitor A and competitor B, respectively. Because endotoxins are significantly smaller than the pore size of the Critical Process Filtration's charged PES filter, the high level of observed endotoxin removal is attributed to the filter’s positive charge. The enhanced endotoxin removal by positive charge is based on electrokinetic interaction.
Selecting the right filters
Several factors must be considered when selecting Sterilizing and Endotoxin removal filters:
Process flow rate
The required process flow rate will determine the number of filter elements and housing size required.
Potential bacteria load in the process stream
The potential for bacteria contamination is a consideration in whether an in-line sterilization process (UV, etc.) is required. If a high load is anticipated, the in-line process will help reduce the load on the sterilizing filters. If the anticipated load is low, then the filters alone could be sufficient.
Potential endotoxin load in the process stream
If the potential for endotoxin contamination is low, a positively charged sterilizing filter could be an option. If the expected load is high, the implementation of separate sterilizing and endotoxin removal filters would allow for the replacement of only the endotoxin filter as the charge is depleted.
Acceptable endotoxin concentration
Most systems target < 0.25 EU/ml (the limit for Water for Injection). Some system limits may be even more stringent. The specified limit will help determine the appropriate endotoxin filter. For most applications, a positively charge 0.22-micron filter will be sufficient. For stricter applications, a dual layer 0.22 or even 0.10 micron charged filter might be a better option.
Testing frequency
How often the process will be tested for bacteria and endotoxin will play a role in filter selection. If testing will be infrequent, then the filters selected will need to have a high assurance of performance - perhaps a 0.10-micron rating instead of 0.22-micron. This may also require some level of validation. If testing is conducted frequently, then you will have ongoing evidence that the filters are performing as expected.
Expected filter life
How long you expect your filters to last will have an impact on the size of your filter process. This will also be affected by the expected levels of bacterial and endotoxin contamination.
Filter change-out strategy
Do you test your process frequently and change filters based on performance? Or do you expect the filters to last a certain time and change them proactively based on a schedule? Here again, this can impact which filters to use and how many will be required in your system.
Where to place filters
In a recirculating system, it is usually advisable to install separate sterilizing and endotoxin removal filters. The sterilizing filter will remove bacteria and other contaminants that might affect the performance of the endotoxin removal filter. This also allows placement of the filters where they will be more efficient and allows for the replacement of the individual filters as needed. For example, in a system utilizing a tank for recirculation, the sterilizing filter could be placed on the inlet side of the tank. The endotoxin removal filter would be better placed toward the outlet side of the recirculating loop, after any in-line sterilization steps (UV, etc.). Critical Process Filtration's charged filter will remove endotoxins in a stand-alone position.
When considering a point-of-use filter, a positively charged sterilizing grade filter would be a good choice. This will remove any unintended contamination of both bacteria and endotoxin and provide added assurance for process and product safety.
In a non-recirculating system, the decision to use separate or combined sterilizing and endotoxin removal filters would depend on potential bacteria and endotoxin load, where in the process it is critical to prevent contamination, and where any in-line sterilization steps are installed.
Applications of the Critical Process Filtration's charged PES Filter include:
- Reduction in endotoxin levels in pharmaceutical-grade water systems;
- Pharmaceutical and medical device manufacturing;
- Active pharmaceutical preparations;
- High-purity chemicals;
- Large volume parenteral;
- Complex solutions.
DOES YOUR APPLICATION CALL FOR ENDOTOXIN REMOVAL?
If so, let us help you configure the best solution. We have an experienced Applications and Process Support staff that will identify the appropriate sterilizing and/or endotoxin removal filters for your process while optimizing the cost of your filters. And we deliver fast - products ship in weeks, not months!
From presales consulting to testing and analysis to full validation services, Critical Process Filtration delivers quality products and support that meet your filtration performance requirements.
CONTACT US TODAY AT (603) 880-4420 OR SEND US YOUR ENDOTOXIN FILTRATION QUESTIONS HERE AT SALES@CRITICALPROCESS.COM
