Enzyme and Microbial Technology 40 (2007) 273–277 Laccase isolation by foam fractionation—New prospects of an old process Diana Linke a , Holger Zorn a , Birte Gerken b , Harun Parlar b , Ralf G. Berger a,∗ a Universit¨ at Hannover, Inst. f. Lebensmittelchemie, Wunstorfer Str. 14, D-30453 Hannover, Germany b TU M ¨ unchen, Lehrstuhl f. Chemisch-Technische Analyse und Chemische Lebensmitteltechnologie, Weihenstephaner Steig 23, D-85354 Freising, Germany Received 3 January 2006; received in revised form 10 April 2006; accepted 17 April 2006 Abstract A laccase (E.C. 1.10.3.2) from Trametes spec. was isolated from aqueous media using foam fractionation. The pH value, superficial velocity, foaming period, and temperature were varied to optimise the transport of the active enzyme into the foam phase. Several detergents were added in varying concentrations to form and stabilize the foam, and the cationic detergent cetyltrimethylammonium bromide (CTAB) proved to be the most appropriate. From water as a model system, maximum recovery rates of 94% of laccase activity were achieved at pH 6.0 in 6min. For separation of the enzyme from protein rich culture media, the operation conditions had to be adjusted. At pH 5.4, 89% of laccase activity was transported into the foam phase after 15 min. The method established was successfully applied to the isolation of an active laccase isoenzyme from submerged cultures of the basidiomycete Pleurotus sapidus. © 2006 Elsevier Inc. All rights reserved. Keywords: Foam fractionation; Downstream process; Laccase; Basidiomycete 1. Introduction Foam fractionation is a gentle, environmentally compatible, inexpensive, and selective method for the effective separation of surface-active compounds from diluted aqueous solutions [1,2]. This process was first patented in 1920 [3] and received renewed interest in separation of biological materials such as proteins [4–6], microorganisms [7], suspended solids [8], aromatic sub- stances [9], and pigments [10] in recent years. A mathematical model of the foaming process of albumines was described by Du et al. [11]. Foam systems consist of a thin liquid film including a gaseous phase. The two phases are separated by a thin, anisotropic interface region [12]. Surface-active compounds adsorb to the interface of the gas–liquid dispersion and thus stabilize the dis- persed gas phase by self-orientation. Proteins, as amphiphilic compounds, strongly interact with the interface and are thereby separated from a feed phase into the foam phase. While the hydrophobic parts of the protein attach to the gas phase, the hydrophilic parts are orientated towards the liquid phase. Several parameters such as pH value and protein concentration influence the protein adsorption, and thus the separation efficiency and the ∗ Corresponding author. Tel.: +49 511 762 4581; fax: +49 511 762 4547. E-mail address: rg.berger@lci.uni-hannover.de (R.G. Berger). success of a foaming process. The adsorption to the interface has been described as a three-step process, comprising protein denaturation in the second step [13]. For this reason, foam frac- tionation of active enzymes has been considered impossible in the past. Numerous enzymes for the food, pharmaceutical and cos- metic industry are currently isolated by multistage processes such as precipitation, ultrafiltration, and dialysis, followed by several liquid chromatographic steps. These methods are gen- erally time-consuming, expensive, and above all, often come along with a high loss of enzymatic activity. In this study, foam fractionation of a commercial laccase preparation from the basidiomycete Trametes spec. and from submerged cultures of Pleurotus sapidus was investigated. Laccases currently gain a great deal of attention for bioremediation, beverage (wine, fruit juice, tea, beer) processing, ascorbic acid detection, pectin gela- tion, baking, and as biosensor [14,15]. The aim was to develop a foaming process for the separation of extracellular laccases from fungal culture while maintaining the full enzymatic activity. 2. Materials and methods 2.1. Materials Laccase (E.C. 1.10.3.2) from Trametes sp. was obtained from ASA Spezialenzyme (Wolfenb¨ uttel, Germany). Cetyltrimethylammonium bromide 0141-0229/$ – see front matter © 2006 Elsevier Inc. All rights reserved. doi:10.1016/j.enzmictec.2006.04.010