Carbohydrate carbon sources induce loss of flocculation of an ale-brewing yeast strain E.V. Soares 1 , A. Vroman 1,2 , J. Mortier 1,3 , K. Rijsbrack 1,3 and M. Mota 4 1 CIEA-Departamento de Engenharia Quı´mica, Instituto Superior de Engenharia do Instituto Polite´cnico do Porto, Rua Dr Anto´nio Bernardino de Almeida, Porto, Portugal, 2 KdGH-Antwerpen, Departement Industriele Wetenschappen en Technologie, Campus KIHA, Salesianenlaan, Hoboken, Belgium, 3 KaHo St.-Lieven, Industrial Engineering, Department of Biochemistry-Microbiology, Gebroeders Desmetstraat, Gent, Belgium, and 4 Departamento de Engenharia Biolo´gica, Universidade do Minho, Campus de Gualtar, Braga, Portugal 2003/0656: received 25 July 2003, revised 16 October 2003 and accepted 27 January 2004 ABSTRACT E.V. SOARES, A. VROMAN, J. MORTIER, K. RIJSBRACK AND M. MOTA. 2004. Aims: To identify the nutrients that can trigger the loss of flocculation under growth conditions in an ale-brewing strain, Saccharomyces cerevisiae NCYC 1195. Methods and Results: Flocculation was evaluated using the method of Soares, E.V. and Vroman, A. [Journal of Applied Microbiology (2003) 95, 325]. Yeast growth with metabolizable carbon sources (glucose, fructose, galactose, maltose or sucrose) at 2% (w/v), induced the loss of flocculation in yeast that had previously been allowed to flocculate. The yeast remained flocculent when transferred to a medium containing the required nutrients for yeast growth and a sole nonmetabolizable carbon source (lactose). Transfer of flocculent yeast into a growth medium with ethanol (4% v/v), as the sole carbon source did not induce the loss of flocculation. Even the addition of glucose (2% w/v) or glucose and antimycin A (0Æ1 mg l )1 ) to this culture did not bring about loss of flocculation. Cycloheximide addition (15 mgl )1 ) to glucose-growing cells stopped flocculation loss. Conclusions: Carbohydrates were the nutrients responsible for stimulating the loss of flocculation in flocculent yeast cells transferred to growing conditions. The glucose-induced loss of flocculation required de novo protein synthesis. Ethanol prevented glucose-induced loss of flocculation. This protective effect of ethanol was independent of the respiratory function of the yeast. Significance and Impact of the Study: This work contributes to the elucidation of the role of nutrients in the control of the flocculation cycle in NewFlo phenotype yeast strains. Keywords: carbon source, flocculation, NewFlo phenotype, protein synthesis, Saccharomyces cerevisiae. INTRODUCTION Yeast flocculation has been traditionally exploited in brew- ing industry (Stewart and Russell 1981). However, this characteristic might also be useful in modern biotechnology as it is an easy and cheap way of separating the cell biomass from the media at the end of fermentations. According to lectin-like theory, flocculation occurs as a consequence of the interaction between the specific floccu- lation proteins (lectins) present only in the flocculent cells and the carbohydrate residues (receptors) of the walls of neighbouring cells (Miki et al. 1982); in this process, Ca 2+ seems to induce the correct conformation of the lectins (Miki et al. 1982; Stratford 1989). Four phenotypes were described taking into account pH and sugar inhibition sensitiveness or the requirement of ethanol to induce flocculation, they were: Flo1 phenotype, the flocculation of these strains was only inhibited by mannose; NewFlo phenotype, inhibited by Correspondence to: E.V. Soares, CIEA-Departamento de Engenharia Quı´mica, Instituto Superior de Engenharia do Instituto Polite´cnico do Porto, Rua Dr Anto´nio Bernardino de Almeida, 431, 4200-072 Porto, Portugal (e-mail: evs@isep.ipp.pt). ª 2004 The Society for Applied Microbiology Journal of Applied Microbiology 2004, 96, 1117–1123 doi:10.1111/j.1365-2672.2004.02240.x