Citric acid production by Yarrowia lipolytica cultivated on olive-mill wastewater-based media Seraphim Papanikolaou a, * , Maria Galiotou-Panayotou a , Stylianos Fakas a , Michael Komaitis a , George Aggelis b a Department of Food Science and Technology, Agricultural University of Athens, 75 Iera Odos, 11855 Athens, Greece b Department of Biology, Division of Genetics, Cell and Development Biology, University of Patras, 26504 Patras, Greece Received 6 June 2006; received in revised form 2 May 2007; accepted 5 May 2007 Available online 2 July 2007 Abstract Yarrowia lipolytica ACA-DC 50109 cultivated on olive-mill wastewater (O.M.W.)-based media, enriched with commercial-industrial glucose, presented an efficient cell growth. Parameters of growth were unaffected by the presence of O.M.Ws in the growth medium. In diluted O.M.Ws enriched with high glucose amounts (initial sugar concentration, 65 g l 1 ), a notable quantity of total citric acid was produced (28.9 g l 1 ). O.M.W.-based media had a noteworthy stimulating effect on the production of citric acid, since both final citric acid concentration and conversion yield of citric acid produced per unit of sugar consumed were higher when compared with the respec- tive parameters obtained from trials without added O.M.W. Adaptation of the strain in O.M.W.-based media favoured the biosynthesis of cellular unsaturated fatty acids (principally of oleic and palmitoleic acids). Additionally, a non-negligible decrease of the phenolic compounds in the growth medium [up to 15% (wt/wt)], a slight decrease of the phyto-toxicity, and a remarkable decolourisation of the O.M.W. were observed. All these results suggest the potentiality of O.M.Ws utilisation in the fermentation process of citric acid production. Ó 2007 Elsevier Ltd. All rights reserved. Keywords: Yarrowia lipolytica; Fermentation; Olive-mill wastewater; Citric acid 1. Introduction The manufacturing process of olive oil production yields a liquid fraction, which is called olive-mill wastewater (O.M.W.). This important residue of the olive oil industry is one of the most difficult to treat wastes because of its high content in phenolic compounds (Gharsallah et al., 1999; Garcia Garcia et al., 2000; Aggelis et al., 2003; Ammar et al., 2005). Besides the presence of phenolic sub- stances, O.M.Ws cause serious environmental problems due to their potentially high concentration in sugars, tan- nins, pigments and emulsified oil that result in increased C.O.D. values, and their black colour that stains indelibly the soil (Scioli and Vollaro, 1997; Tsioulpas et al., 2002; Fadil et al., 2003). To reduce pollution in O.M.Ws, chem- ical or biological processes have been used (Mantzavinos and Kalogerakis, 2005; Crognale et al., 2006). One princi- pal strategy involves the use of moulds (e.g. Phanerochaete spp., Pleurotus spp., Panus tigrinus, Geotrichum spp., Len- tinula edodes, Trametes versicolor or Aspergillus spp.), which have been found capable of reducing C.O.D. values and breaking down phenolic compounds of the O.M.Ws treated (Sayadi and Ellouz, 1992, 1995; Gharsallah et al., 1999; Garcia Garcia et al., 2000; Tsioulpas et al., 2002; Aggelis et al., 2003; Fenice et al., 2003; D’Annibale et al., 2004; Ayed et al., 2005; Dhouib et al., 2006). Likewise, pro- karyotic microorganisms (e.g. Pseudomonas putida, Klebsi- ella oxytoca, Lactobacillus plantarum, Citrobacter diversus) have the capability to degrade phenolic compounds, in the presence or absence of sugars in the growth media (Seker et al., 1997; Lamia and Moktar, 2003; Ammar et al., 0960-8524/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.biortech.2007.05.005 * Corresponding author. Tel./fax: +30 210 5294700. E-mail address: spapanik@aua.gr (S. Papanikolaou). Available online at www.sciencedirect.com Bioresource Technology 99 (2008) 2419–2428