How are endotoxins removed?

Traditionally, endotoxins have been removed by reverse osmosis or ultrafiltration membranes of a 10,000 NMWL cutoff12. However, this approach is not practical when a component of a drug product has a molecular weight greater than 10 kDa.

How are endotoxins removed from water?

Current methods for removing endotoxin include distillation and reverse osmosis, both of which are resource intensive processes. Membranes that present an absolute barrier to macromolecular passage may be capable of delivering pure water for biomedical applications.

What is DEAE ion exchange chromatography?

Anion-exchange chromatography is a process that separates substances based on their charges using an ion-exchange resin containing positively charged groups, such as diethyl-aminoethyl groups (DEAE). In solution, the resin is coated with positively charged counter-ions (cations).

What is the pI of endotoxin?

Petsch showed the pI to be around 8 to 9.5 for the IgG used in Table 1. 1, 2 Therefore, at pH 8 to 9.5 depending upon the specific protein, samples below pH 8 (commonly parenteral drugs are formulated around pH 6 – 7) can be treated with base to neutralize their charge and thereby free up endotoxin in solution.

How do you get rid of bacterial endotoxins?

Endotoxin is continuously shed from the outer membrane of viable gram-negative bacteria and is released when the bacterial cell dies. Although bacteria are often removed by using a 0.2 μm sterilizing grade filter, LPS itself is difficult to remove or inactivate because it is extremely heat and pH stable.

How can bacterial endotoxins be reduced?

The most common depyrogenation procedures for physical components include incineration and removal by washing, also termed dilution. The literature has shown other procedures, such as filtration, irradiation and ethylene oxide treatment to have limited effect in reducing pyrogen/endotoxin levels.

Can endotoxins be removed by filtration?

Endotoxins are negatively charged at pH >2 and can be efficiently removed by positively charged filters such as the ELGA LabWater Biofilter.

Does UV remove endotoxin?

Endotoxins, which are present in the cell walls of Gram-negative bacteria, pose a potential health hazard and must also be removed from process water, especially in high purity applications like pharmaceutical manufacturing and food processing. …

What is DEAE Sepharose chromatography?

DEAE Sepharose Fast Flow is a weak anion exchange chromatography resin (medium) for fast protein purification. Well-proven weak anion exchanger developed for industrial downstream processes. Used extensively for capture and intermediate purification of a wide range of approved biopharmaceuticals.

What is DEAE cellulose chromatography?

Diethylaminoethyl cellulose (DEAE-C) is a positively charged resin used in ion-exchange chromatography, a type of column chromatography, for the separation and purification of proteins and nucleic acids.

How can I lower my endotoxin levels?

Endotoxin can be inactivated when exposed at temperature of 250º C for more than 30 minutes or 180º C for more than 3 hours (28, 30). Acids or alkalis of at least 0.1 M strength can also be used to destroy endotoxin in laboratory scale (17).

How do you remove LPS from protein?

EDTA strips most of the LPS of the cell surface. This way there will be lower LPS content in cell lysate to begin with. If your protein/purification protocol can tolerate acidic pH this would also decrease LPS contamination. Polymixin B resin is very efficient in LPS removal.

How do you remove endotoxins from proteins?

In the presence of proteins, affinity chromatography and two-phase extraction methods can take advantage of the physical-chemical interaction between endotoxin and protein to completely remove endotoxin [ 6 F. B. Anspach and O. Hilbeck, “Removal of endotoxins by affinity sorbents,” Journal of Chromatography A, vol. 711, no. 1, pp. 81–92, 1995.

Is two-phase partitioning an effective method of endotoxin removal?

With endotoxin removal, the hydrophobic nature of endotoxin means that two-phase partitioning serves as an effective endotoxin removal method.

What is endendotoxin and why should you care?

Endotoxins are heat stable, making them resistant to most conventional sterilization processes and thus necessitating separate tests for viable cells (bioburden) and endotoxin. Pharmaceutical processes and equipment are at risk from endotoxin.

How do affaffinity adsorbents remove endotoxins?

Affinity adsorbents based on a polymyxin B ligand ( Talmadge and Siebert, 1989, Anspach and Hilbeck, 1995 ), intended to remove endotoxin by binding the endotoxin rather than the product, were tested.