Yeast Lipid Produced through Glycerol Conversions and Its Use for Enzymatic Synthesis of Amino Acid-Based Biosurfactants

The aim of the present work was to obtain microbial lipids (single-cell oils and SCOs) from oleaginous yeast cultivated on biodiesel-derived glycerol and subsequently proceed to the enzymatic synthesis of high-value biosurfactant-type molecules in an aqueous medium, with SCOs implicated as acyl donors (ADs). Indeed, the initial screening of five non-conventional oleaginous yeasts revealed that the most important lipid producer was the microorganism Cryptococcus curvatus ATCC 20509. SCO production was optimised according to the nature of the nitrogen source and the initial concentration of glycerol (Glyc0) employed in the medium. Lipids up to 50% w/w in dry cell weight (DCW) (SCOmax = 6.1 g/L) occurred at Glyc0 ≈ 70 g/L (C/N ≈ 80 moles/moles). Thereafter, lipids were recovered and were subsequently used as ADs in the N-acylation reaction catalysed by aminoacylases produced from Streptomyces ambofaciens ATCC 23877 under aqueous conditions, while Candida antarctica lipase B (CALB) was used as a reference enzyme. Aminoacylases revealed excellent activity towards the synthesis of acyl-lysine only when free fatty acids (FAs) were used as the AD, and the rare regioselectivity in the α-amino group, which has a great impact on the preservation of the functional side chains of any amino acids or peptides. Aminoacylases presented higher α-oleoyl-lysine productivity and final titer (8.3 g/L) with hydrolysed SCO than with hydrolysed vegetable oil. The substrate specificity of both enzymes towards the three main FAs found in SCO was studied, and a new parameter was defined, viz., Specificity factor (Sf), which expresses the relative substrate specificity of an enzyme towards a FA present in a FA mixture. The Sf value of aminoacylases was the highest with palmitic acid in all cases tested, ranging from 2.0 to 3.0, while that of CALB was with linoleic acid (0.9–1.5). To the best of our knowledge, this is the first time that a microbial oil has been successfully used as AD for biosurfactant synthesis. This bio-refinery approach illustrates the concept of a state-of-the-art combination of enzyme and microbial technology to produce high-value biosurfactants through environmentally friendly and economically sound processes.

[1]  S. Papanikolaou,et al.  Biotechnological Conversions of Mizithra Second Cheese Whey by Wild-Type Non-Conventional Yeast Strains: Production of Yeast Cell Biomass, Single-Cell Oil and Polysaccharides , 2022, Applied Sciences.

[2]  Min Li,et al.  Production, Biosynthesis, and Commercial Applications of Fatty Acids From Oleaginous Fungi , 2022, Frontiers in Nutrition.

[3]  Yuanda Song,et al.  Microbes: A Hidden Treasure of Polyunsaturated Fatty Acids , 2022, Frontiers in Nutrition.

[4]  S. Papanikolaou,et al.  Sustainable arabitol production by a newly isolated Debaryomyces prosopidis strain cultivated on biodiesel-derived glycerol , 2022, Carbon Resources Conversion.

[5]  M. Sauer,et al.  Utilizing yeasts for the conversion of renewable feedstocks to sugar alcohols - A review. , 2021, Bioresource technology.

[6]  S. Papanikolaou,et al.  Biotechnological valorization of biodiesel-derived glycerol: Trials with the non-conventional yeasts Yarrowia lipolytica and Rhodosporidium sp , 2021 .

[7]  S. Papanikolaou,et al.  Production of added-value microbial metabolites during growth of yeast strains on media composed of biodiesel-derived crude glycerol and glycerol/xylose blends. , 2020, FEMS microbiology letters.

[8]  S. Delaunay,et al.  N-acylation of L-amino acids in aqueous media: Evaluation of the catalytic performances of Streptomyces ambofaciens aminoacylases. , 2020, Enzyme and microbial technology.

[9]  A. Koutinas,et al.  Bioprocess development for the production of novel oleogels from soybean and microbial oils. , 2019, Food research international.

[10]  S. Papanikolaou,et al.  Sources of microbial oils with emphasis to Mortierella (Umbelopsis) isabellina fungus , 2019, World journal of microbiology & biotechnology.

[11]  S. Delaunay,et al.  Molecular rules for selectivity in lipase-catalysed acylation of lysine , 2018, Process Biochemistry.

[12]  S. Delaunay,et al.  An aminoacylase activity from Streptomyces ambofaciens catalyzes the acylation of lysine on α‐position and peptides on N‐terminal position , 2018, Engineering in life sciences.

[13]  W. Rymowicz,et al.  Recent advances in biological production of erythritol , 2018, Critical reviews in biotechnology.

[14]  John M. Woodley,et al.  Role of Biocatalysis in Sustainable Chemistry. , 2017, Chemical reviews.

[15]  A. Elazzazy,et al.  Microbial oils as food additives: recent approaches for improving microbial oil production and its polyunsaturated fatty acid content. , 2016, Current opinion in biotechnology.

[16]  S. Papanikolaou,et al.  Oleaginous yeast Cryptococcus curvatus exhibits interplay between biosynthesis of intracellular sugars and lipids , 2015 .

[17]  A. Elazzazy,et al.  Microalgal lipids biochemistry and biotechnological perspectives. , 2014, Biotechnology advances.

[18]  R. Mechoulam,et al.  N‐Acyl amino acids and their impact on biological processes , 2014, BioFactors.

[19]  V. Tyagi,et al.  Synthesis, Properties, and Applications of Amino Acids Based Surfactants: A Review , 2013 .

[20]  Seraphim Papanikolaou,et al.  Lipids of oleaginous yeasts. Part I: Biochemistry of single cell oil production. , 2011 .

[21]  Tomislav Ivanković,et al.  Surfactants in the Environment , 2010, Arhiv za higijenu rada i toksikologiju.

[22]  K. Imamura,et al.  Efficient Nepsilon-lauroyl-L-lysine production by recombinant epsilon-lysine acylase from Streptomyces mobaraensis. , 2009, Journal of biotechnology.

[23]  I. Alric,et al.  Synthesis and Properties of Lipoamino Acid–Fatty Acid Mixtures: Influence of the Amphiphilic Structure , 2009 .

[24]  L. Pérez,et al.  Cationic surfactants from lysine: synthesis, micellization and biological evaluation. , 2009, European journal of medicinal chemistry.

[25]  I. Chevalot,et al.  Enzymatic acylation of polar dipeptides: Influence of reaction media and molecular environment of functional groups , 2009 .

[26]  F. Blanchard,et al.  Chemo-selectivity of the N,O-enzymatic acylation in organic media and in ionic liquids , 2008 .

[27]  H. Khandelia,et al.  The impact of peptides on lipid membranes. , 2008, Biochimica et biophysica acta.

[28]  Z. Cohen,et al.  Microbial and algal oils: Do they have a future for biodiesel or as commodity oils? , 2008 .

[29]  S. Papanikolaou,et al.  Biotechnological valorisation of raw glycerol discharged after bio-diesel (fatty acid methyl esters) manufacturing process: Production of 1,3-propanediol, citric acid and single cell oil , 2008 .

[30]  Zhaofu Fei,et al.  Cytotoxicity of ionic liquids and precursor compounds towards human cell line HeLa , 2007 .

[31]  M. Babizhayev,et al.  Biological activities of the natural imidazole-containing peptidomimetics n-acetylcarnosine, carcinine and L-carnosine in ophthalmic and skin care products. , 2006, Life sciences.

[32]  S. Adachi,et al.  A novel acylase from Streptomyces mobaraensis that efficiently catalyzes hydrolysis/synthesis of capsaicins as well as N-acyl-L-amino acids and N-acyl-peptides. , 2006, Journal of agricultural and food chemistry.

[33]  S. Adachi,et al.  Lipase-catalyzed synthesis of O-lauroyl l-serinamide and O-lauroyl l-threoninamide , 2005 .

[34]  M. Basri,et al.  Response surface methodological study on lipase-catalyzed synthesis of amino acid surfactants , 2004 .

[35]  T. Fujita,et al.  Stability of Acyl Derivatives of Insulin in the Small Intestine: Relative Importance of Insulin Association Characteristics in Aqueous Solution , 1994, Pharmaceutical Research.

[36]  M. Infante,et al.  Amino acid-based surfactants , 2004 .

[37]  S. Papanikolaou,et al.  Yarrowia lipolytica as a potential producer of citric acid from raw glycerol , 2002, Journal of applied microbiology.

[38]  P. Clapés,et al.  Amino Acid-based Surfactants: Enzymatic Synthesis, Properties and Potential Applications , 2002 .

[39]  G. Nychas,et al.  Characterization of Pseudomonas spp. Associated with Spoilage of Gilt-Head Sea Bream Stored under Various Conditions , 2002, Applied and Environmental Microbiology.

[40]  A. Kiener,et al.  Industrial biocatalysis today and tomorrow , 2001, Nature.

[41]  P. Bahadur,et al.  A comparative study on the surface activity and micellar behaviour of some N-acylamino acid based surfactants , 1998 .

[42]  S. Handy,et al.  Towards the Ideal Synthesis , 1998 .

[43]  J. Kato,et al.  n-acylation of β-amino alcohol by acyl migration following enzyme-catalyzed esterification , 1996 .

[44]  G. Eggink,et al.  High-cell-density cultivation of the lipid accumulating yeast Cryptococcus curvatus using glycerol as a carbon source , 1996, Applied Microbiology and Biotechnology.

[45]  Alexander M. Klibanov,et al.  Enzyme-catalyzed processes in organic solvents. , 1985 .

[46]  C. T. Evans,et al.  Influence of Nitrogen Metabolism on Lipid Accumulation by Rhodosporidium toruloides CBS 14 , 1984 .

[47]  M. M. Bradford A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. , 1976, Analytical biochemistry.