A Method for Removal of Endotoxin from Pharmaceutical Formulation - The authors describe a simple method to remove endotoxins from highly viscous formulations. - BioPharm International

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A Method for Removal of Endotoxin from Pharmaceutical Formulation
The authors describe a simple method to remove endotoxins from highly viscous formulations.


BioPharm International
Volume 25, Issue 9, pp. 26-29

Sampling for endotoxin analysis

Because of the high viscosity, it was crucial to mix the formulation thoroughly after elution through the Sartobind Q15 anion exchanger. As the absorption efficiency of the Q15 will decline rather sharply when the capacity is exceeded, a marked difference in the endotoxin level can be observed between the start and end of the removal process.

Analysis of endotoxin content


Table I: Values of endotoxin, pH and Gd/Iodine concentrations of the formulations before and after treatment with Sartobind Q15. Note that the values of endotoxin are the measured ones and differ from the theoretical values cited in the experimental section.
All reagents and standards were supplied by Lonza (Basel, Switzerland). The chromogenic method was used for determination of the endotoxin content with a sensitivity of 0.005 EU/mL, as described in the pharmacopeia. A dilution of 1:200 was prepared by diluting 50 µl formulation in 9.95 mL WFI, and the sample was analyzed on the kinetic QCL instrument together with a positive control. If the positive control was positive, the test was considered valid and the results were automatically calculated by the software. No endotoxin present would give a result of NMT 1 EU/mL.

Application to formulations containing Na + and Ca 2+


Figure 1: The capacity of Sartobind Q15 for lipopolysaccharide (LPS) of Escherichia Coli. Comparison of the endotoxin content pre- and post-filtration with the anion exchange membrane disclose that the approximate capacity for each cartridge is around 450–460 European Units (EU). Within this capacity range the filtrate has a measured endotoxin concentration < 0.5 EU/mL that is safe for intravenous injection into experimental animals.
Occasionally, unintentional contamination of sterile formulations takes place. The following describes such an example where the method was used to depyrogenize formulations of X-ray agents having endotoxin levels far above values accepted for injection. The formulations were made with different ratios of Na+ and Ca2+ that were important to preserve. In addition to the equilibration and sterilization procecures described above, the anion exchange membranes were equilibrated with TRIS-buffers having the same Ca2+/Na+ ratios as the formulations. The volumes of formulations were 100 mL, and Sartobind Q100 (100 cm2 ) membrane filters having a theoretical capacity for endotoxin of 4750 EU were used.


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