Cocoa butter-like lipid production ability of non-oleaginous and oleaginous yeasts under nitrogen-limited culture conditions

Applied Microbiology and Biotechnology, Feb 2017

Cocoa butter (CB) extracted from cocoa beans is the main raw material for chocolate production. However, growing chocolate demands and limited CB production has resulted in a shortage of CB supply. CB is mainly composed of three different kinds of triacylglycerols (TAGs), POP (C16:0–C18:1–C16:0), POS (C16:0–C18:1–C18:0), and SOS (C18:0–C18:1–C18:0). The storage lipids of yeasts, mainly TAGs, also contain relative high-level of C16 and C18 fatty acids and might be used as CB-like lipids (CBL). In this study, we cultivated six different yeasts, including one non-oleaginous yeast strain, Saccharomyces cerevisiae CEN.PK113-7D, and five oleaginous yeast strains, Trichosporon oleaginosus DSM11815, Rhodotorula graminis DSM 27356, Lipomyces starkeyi DSM 70296, Rhodosporidium toruloides DSM 70398, and Yarrowia lipolytica CBS 6124, in nitrogen-limited medium and compared their CBL production ability. Under the same growth conditions, we found that TAGs were the main lipids in all six yeasts and that T. oleaginosus can produce more TAGs than the other five yeasts. Less than 3% of the total TAGs were identified as potential SOS in the six yeasts. However, T. oleaginosus produced 27.8% potential POP and POS at levels of 378 mg TAGs/g dry cell weight, hinting that this yeast may have potential as a CBL production host after further metabolic engineering in future.

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Cocoa butter-like lipid production ability of non-oleaginous and oleaginous yeasts under nitrogen-limited culture conditions

Cocoa butter-like lipid production ability of non-oleaginous and oleaginous yeasts under nitrogen-limited culture conditions Yongjun Wei 0 1 2 Verena Siewers 0 1 2 Jens Nielsen 0 1 2 0 Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark , 2800 Kgs. Lyngby , Denmark 1 Novo Nordisk Foundation Center for Biosustainability, Chalmers University of Technology , SE-412 96 Gothenburg , Sweden 2 Department of Biology and Biological Engineering, Chalmers University of Technology , SE-412 96 Gothenburg , Sweden Cocoa butter (CB) extracted from cocoa beans is the main raw material for chocolate production. However, growing chocolate demands and limited CB production has resulted in a shortage of CB supply. CB is mainly composed of three different kinds of triacylglycerols (TAGs), POP (C16:0-C18:1C16:0), POS (C16:0-C18:1-C18:0), and SOS (C18:0C18:1-C18:0). The storage lipids of yeasts, mainly TAGs, also contain relative high-level of C16 and C18 fatty acids and might be used as CB-like lipids (CBL). In this study, we cultivated six different yeasts, including one non-oleaginous yeast strain, Saccharomyces cerevisiae CEN.PK113-7D, and five oleaginous yeast strains, Trichosporon oleaginosus DSM11815, Rhodotorula graminis DSM 27356, Lipomyces starkeyi DSM 70296, Rhodosporidium toruloides DSM 70398, and Yarrowia lipolytica CBS 6124, in nitrogen-limited medium and compared their CBL production ability. Under the same growth conditions, we found that TAGs were the main lipids in all six yeasts and that T. oleaginosus can produce more TAGs than the other five yeasts. Less than 3% of the total TAGs were identified as potential SOS in the six yeasts. However, T. oleaginosus produced 27.8% potential POP and POS at levels of 378 mg TAGs/g dry cell weight, hinting that this yeast may have potential as a CBL production host after further metabolic engineering in future. Cocoa butter-like lipids; Oleaginous yeasts; Lipid production; TAG profiles - Cocoa butter (CB) is extracted from cocoa beans of the cocoa tree Theobroma cacao, and besides being used as food flavor and cosmetics additive, CB is the main component of chocolate (Lipp and Anklam 1998). Due to variations in CB production and an increasing chocolate demand by consumers, the CB supply is currently insufficient and its price has increased in recent years (Clough et al. 2009). Although some other vegetable oils can be used as CB equivalents, for example, Illipe butter, shea butter, and kokum butter, their production is also limited (Jahurul et al. 2013; Verstringe et al. 2012). Therefore, there are interests in developing other routes for production of CB-like lipids (CBL), which can be used as a stable and economically feasible supply for chocolate production (Clough et al. 2009). CB mainly contains three different kinds of triacylglycerols (TAGs), which are esters formed with one glycerol and three fatty acids (Jahurul et al. 2013). The fatty acids in the sn-1 and sn-3 positions of CB TAG backbone glycerol are mainly palmitic acid (C16:0) or stearic acid (C18:0), and the fatty acid in sn-2 position is predominantly oleic acid (C18:1). POP (C16:0–C18:1–C16:0), POS (C16:0–C18:1–C18:0), and SOS (C18:0–C18:1–C18:0) are the three main CB TAGs, and they are therefore also the desired TAGs of CBL (Jahurul et al. 2013). POP, POS, and SOS ratios in CB are 14–16.4, 34.6–38.3, and 23.7–28.4%, respectively (Lipp and Anklam 1998). Moreover, the fatty acid distributions of CB are C16:0 (24.1–25.8%), C18:0 (33.3–37.6%), and C18:1 (32.7–36.5%) (Lipp and Anklam 1998). Currently, enzymatic re-esterification of vegetable oils using lipases is used for CBL production (Ferreira-Dias et al. 2013; Matsuo et al. 1981; Mohamed 2013; Verstringe et al. 2012; Xu 2000). However, re-esterification requires hydrogenation of large amounts of plant oils which have limited production (FerreiraDias et al. 2013; Wang et al. 2006). In addition to the lipaseassisted method, yeasts, especially oleaginous yeasts, also have potential application for CBL production, as the main fatty acids produced by yeasts are C16 and C18 fatty acids (Beopoulos et al. 2011; Papanikolaou and Aggelis 2011). For the model yeast, Saccharomyces cerevisiae, global analysis of its lipidome, has shown that only small amounts of CBL were produced (Ejsing et al. 2009). On the other hand, several different oleaginous yeast strains have been reported as candidates for CBL production, such as Yarrowia lipolytica, Rhodosporidium toruloides, Lipomyces starkeyi, and Cryptococcus curvatus (Hassan et al. 1994; Hassan et al. 1995; Papanikolaou et al. 2001; Papanikolaou et al. 2003; Wu et al. 2011). Though the cultivation conditions that affect fatty acid production have been examined in several yeasts (Kolouchová et al. 2016), a detailed comparison of the CBL production ability in different yeasts and given conditions has rarely been reported. In this study, we compared the compositions of total lipids, (...truncated)


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Yongjun Wei, Verena Siewers, Jens Nielsen. Cocoa butter-like lipid production ability of non-oleaginous and oleaginous yeasts under nitrogen-limited culture conditions, Applied Microbiology and Biotechnology, 2017, pp. 3577-3585, Volume 101, Issue 9, DOI: 10.1007/s00253-017-8126-7