Tanja Mehling

Micellar liquid chromatography (MLC) is an alternative to conventional reversed-phase chromatography (RPLC) [1]. MLC is an already well-established method for the separation of organic molecules out of different aqueous product streams [2]. The mobile phases thereby are aqueous solutions of surfactants at a concentration above the critical micellar concentration (cmc). Due to the variety of possible interactions between micelles, solutes and stationary phase MLC is characterised as a technique adjustable for many different substances. It has been shown, that biomolecules can be purificated by means of micellar systems [3]. Here the separation is based on hydrophobic and electrostatic interactions between micelles and the solvent molecules. The extraction is influenced by different factors [3], like type and concentration of surfactant, solute properties, pH of the solution, ionic strength and temperature. For this reason, a reliable modelling allowing to describe these effects is required.

For enzymes and proteins the separation by means of a two-stage extraction has already been described [4].

Using MLC the separation of target molecules from the aqueous phase can be done in a single process step. Concerning solute concentration, equilibrium is reached between micelles, bulk phase and stationary phase. The relation of concentrations is given by the partition coefficients.

The goal of this work is the separation and purification of proteins and enzymes of different product streams by means of micellar liquid chromatography. The partition coefficients are predicted by means of thermodynamics (COSMO-RS), and based on this information suitable surfactants are selected. To validate the modelling, the calculated partition coefficients are confirmed experimentally.
In the final step chromatographic separation for different product streams is executed and optimised.

• [1] Berthod, Alain; Girard, Ines; Gonnet, Colette (1986): Micellar liquid chromatography. Retention study of solutes of various polarities. In: Anal. Chem., Jg. 58, H. 7, S. 1359–1362.
• [2] Kadam, Kiran L. (1986): Reverse micelles as a bioseparation tool. In: Enzyme and Microbial Technology, Jg. 8, H. 5, S. 266–273.
• [3] Mazzola, Priscila G.; Lopes, Andre M.; Hasmann, Francislene A.; Jozala, Angela F.; Penna, Thereza C. V.; Magalhaes, Perola O. et al. (2008): Liquid-liquid extraction of biomolecules: an overview and update of the main techniques. In: Journal of Chemical Technology & Biotechnology, Jg. 83, H. 2, S. 143–157.
• [4] Noh, Kyung-Hyun; Imm, Jee-Young (2005): One-step separation of lysozyme by reverse micelles formed by the cationic surfactant, cetyldimethylammonium bromide. In: Food Chemistry, Jg. 93, H. 1, S. 95–101.