Analysis of mechanically labile protein crystals

B. Radel*, H. Nirschl, Karlsruhe Institute of Technology (KIT), Germany

Recent progresses in the biotechnology industries have led to much higher product yields after the fermentation. In the case of proteins the purification of such high titers have become a challenging task. The common method is to use several chromatography steps to separate the protein with high purity. However, the high concentration of protein in the mother liquor requires large amounts of toxic and explosive organic solvents and large chromatography columns. Hence, another approach like selective crystallization to separate the protein from the mother liquor is becoming increasingly interesting for the pharmaceutical and biotechnology industry. Crystallized proteins not only offer advantages in the downstream processing but also have other positive properties like extended shelf life, easier product handling and in the case of therapeutic proteins different drug release properties. After the crystallization, the next step is often a solid-liquid separation. In the case of protein crystals this separation is a challenging task because in contrast to common organic or inorganic crystals, protein crystals are much more fragile to mechanical stress. Even low pressure differences may lead to crystal breakage or crystal abrasion. Since therapeutic or enzymatic proteins are often expensive products a method to analyze the mechanical stability and the influence of crystal breakage on the cake filtration with low volumes and hence low product quantities is required.

To analyze the influence of crystal breakage on the cake filtration a special filtration cuvette has been constructed which allows the filtration of volumes less than one milliliter. This cuvette can be used in an optical, analytical centrifuge by LUM GmbH (Berlin, Germany) to perform and monitor the filtration by means of light transmission at the same time. As a model substance hen egg-white lysozyme crystals were investigated. The particle size distribution can be measured by automatic analysis of microscope images or by analysis of sedimentation velocity. By comparing the particle size distributions before and after the filtration the crystal breakage can be quantified at different centrifugal accelerations. In this contribution the experimental setup and the results of experiments with different crystal shapes of lysozyme will be presented in detail.

Session: L4 - Cake Filtration - Particle Properties and Analysis
Day: 14 March 2018
Time: 09:00 - 10:15 h

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