Commercial sugars as substrates for lipid accumulation in Cunninghamella echinulata and Mortierella isabellina fungi

Cunninghamella echinulata and Mortierella isabellina, fungi capable of accumulating single cell oil (SCO) containing γ-linolenic acid (GLA), were cultivated on sugar-based media, at initial substrate concentration 60 g/L. It was demonstrated that the carbon source plays an essential role on lipid accumulation process. Especially, on glucose, C. echinulata produced 3.9 g/L lipids containing 19.5% GLA, whereas M. isabellina achieved 9.9 g/L of lipid, containing 3.5% GLA. Growth on fructose was also satisfactory for both microorganisms. Although M. isabellina failed to grow adequately on saccharose, C. echinulata presented appreciable biomass production of 17.6 g/L, containing 18% of fat. Extra-cellular invertase activity was also detected in the case of C. echinulata cultures, achieving 0.5 U/mL. Both molds presented satisfactory growth on molasses, with significant amounts of SCO and GLA produced, accompanied by non-negligible substrate decolorization. Scaling-up in a 3-L bioreactor of M. isabellina with glucose as a substrate resulted in 17.5 g/L of biomass, containing 12.7 g/L of lipid. Finally, fractionation of cellular lipid for both strains showed that neutral lipids represented about 90-92% w/w upon total accumulated lipids, whilst fractions of glycolipids plus sphingolipids and phospholipids were 6-7 and 3-4%, respectively.

[1]  S. Papanikolaou,et al.  Yarrowia lipolytica: A model microorganism used for the production of tailor‐made lipids , 2010 .

[2]  Seraphim Papanikolaou,et al.  Single cell oil (SCO) production by Mortierella isabellina grown on high-sugar content media. , 2004, Bioresource technology.

[3]  S. Kamzolova,et al.  Succinic Acid Synthesis by Ethanol-Grown Yeasts , 2009 .

[4]  G. L. Miller.  Use of Dinitrosalicylic Acid Reagent for Determination of Reducing Sugar , 1959 .

[5]  S. Narine,et al.  Maximizing the production of γ-linolenic acid in Mortierella ramanniana var. ramanniana as a function of pH, temperature and carbon source, nitrogen source, metal ions and oil supplementation , 2005 .

[6]  A. Navarro,et al.  Regulation of invertase synthesis in Aspergillus niger , 2006 .

[7]  Ram Chandra,et al.  Decolourisation and detoxification of synthetic molasses melanoidins by individual and mixed cultures of Bacillus spp. , 2006, Bioresource technology.

[8]  S. Papanikolaou,et al.  Organic nitrogen of tomato waste hydrolysate enhances glucose uptake and lipid accumulation in Cunninghamella echinulata , 2008, Journal of applied microbiology.

[9]  S. Gascón,et al.  Invertase isozymes and their localization in yeast. , 1967, Comptes-rendus des travaux du Laboratoire Carlsberg.

[10]  Hung-Chang Chen,et al.  Production of γ‐Linolenic Acid by the Fungus Cunninghamella echinulata CCRC 31840 , 1996 .

[11]  F. Smith,et al.  COLORIMETRIC METHOD FOR DETER-MINATION OF SUGAR AND RELATED SUBSTANCE , 1956 .

[12]  I. Ellis,et al.  Gamma linolenic acid with tamoxifen as primary therapy in breast cancer , 1998, International journal of cancer.

[13]  Sudheer Kumar Singh,et al.  Effects of various process parameters on the production of gamma-linolenic acid in submerged fermentation , 2006 .

[14]  Seraphim Papanikolaou,et al.  Lipid production by oleaginous Mucorales cultivated on renewable carbon sources , 2007 .

[15]  M. Čertík,et al.  Lipid formation and γ‐linolenic acid production by Mucorales fungi grown on sunflower oil , 1997 .

[16]  Sudheer Kumar Singh,et al.  Enrichment of γ-linolenic acid in the lipid extracted from Mucor zychae MTCC 5420 , 2009 .

[17]  M. Komaitis,et al.  Lipid and γ-linolenic acid accumulation in strains of zygomycetes growing on glucose , 2001 .

[18]  M. Dostálek,et al.  Effect of culture conditions on mycelial growth and production of γ-linolenic acid by the fungus Mortierella ramanniana , 1988, Applied Microbiology and Biotechnology.

[19]  M. Zong,et al.  Efficient lipid production with Trichosporon fermentans and its use for biodiesel preparation. , 2008, Bioresource technology.

[20]  S. Papanikolaou,et al.  Gamma-linolenic acid production by Cunninghamella echinulata growing on complex organic nitrogen sources. , 2008, Bioresource technology.

[21]  Mark Horton,et al.  Volatile Compounds in Archaeological Plant Remains and the Maillard Reaction During Decay of Organic Matter , 1997 .

[22]  C. Ratledge,et al.  The biochemistry and molecular biology of lipid accumulation in oleaginous microorganisms. , 2002, Advances in applied microbiology.

[23]  Fengwu Bai,et al.  High-density cultivation of oleaginous yeast Rhodosporidium toruloides Y4 in fed-batch culture , 2007 .

[24]  Hideo Tanaka,et al.  Application of Maxblend Fermentor® for microbial processes , 1997 .

[25]  D. Somashekar,et al.  Effect of culture conditions on lipid and gamma-linolenic acid production by mucoraceous fungi , 2003 .

[26]  Guanglei Liu,et al.  Single cell oil production from hydrolysate of cassava starch by marine-derived yeast Rhodotorula mucilaginosa TJY15a. , 2010 .

[27]  T. Ng,et al.  Peptides and proteins from fungi , 2004, Peptides.

[28]  Seraphim Papanikolaou,et al.  Evaluating renewable carbon sources as substrates for single cell oil production by Cunninghamella echinulata and Mortierella isabellina. , 2009 .

[29]  M. Čertík,et al.  Influence of different carbon sources on growth, lipid content and fatty acid composition in four strains belonging to mucorales , 1988, Biotechnology Letters.

[30]  M. C. Rubio,et al.  Purification and characterization of invertase from Aspergillus niger , 1995, Current Microbiology.

[31]  M. Kennedy,et al.  Fatty acid production characteristics of fungi with particular emphasis on gamma linolenic acid production , 1993, Biotechnology and bioengineering.

[32]  D. Adhikari,et al.  Utilization of molasses for the production of fat by an oleaginous yeast,Rhodotorula glutinis IIP-30 , 2005, Journal of Industrial Microbiology.

[33]  David A. Rhoades,et al.  The scale up of mycelial shake flask fermentations: A case study of gamma linolenic acid production byMucor hiemalis IRL 51 , 1994, Journal of Industrial Microbiology.

[34]  S. Papanikolaou,et al.  Lipids of Cunninghamella echinulata with emphasis to γ-linolenic acid distribution among lipid classes , 2006, Applied Microbiology and Biotechnology.

[35]  S. Narine,et al.  Implications for the use of Mortierella fungi in the industrial production of essential fatty acids , 2005 .

[36]  Seraphim Papanikolaou,et al.  Compositional shifts in lipid fractions during lipid turnover in Cunninghamella echinulata , 2007 .

[37]  Hung-Chang Chen,et al.  Inoculum effects on the production of γ-linolenic acid by the shake culture of Cunninghamella echinulata CCRC 31840 , 1997 .

[38]  S. Kamzolova,et al.  Microbiological Production of Citric and Isocitric Acids from Sunflower Oil , 2008 .

[39]  H. Jang,et al.  Effect of culture media and conditions on polyunsaturated fatty acids production by Mortierella alpina. , 2005, Bioresource technology.

[40]  D. Horrobin.  Nutritional and medical importance of gamma-linolenic acid. , 1992, Progress in lipid research.

[41]  C. Ratledge,et al.  17 Microbial Production of Oils and Fats , 2005 .

[42]  Xin Zhao,et al.  Medium optimization for lipid production through co-fermentation of glucose and xylose by the oleaginous yeast Lipomyces starkeyi , 2008 .

[43]  S. Papanikolaou,et al.  Repression of reserve lipid turnover in Cunninghamella echinulata and Mortierella isabellina cultivated in multiple‐limited media , 2004, Journal of applied microbiology.