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

Abstract

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.

Growth Response of Non-Conventional Yeasts on Sugar-Rich Media: Part 1: High Production of Lipid by Lipomyces starkeyi and Citric Acid by Yarrowia lipolytica

“…The production and subsequent utilization of palm oil (this oil presents enormous applications in the sectors of food and biodiesel production) is one of the leading contributors to tropical deforestation, resulting in habitat destruction and increased CO 2 emissions [ 31 ], therefore potential production in large-scale operations of microbial alternatives of this fatty material (like the lipids of R. toruloides NRRL Y-27012 and L. starkeyi DSM 70296) would have much to offer to the resolution to the above-mentioned remarkable problems. On the other hand, all yeast lipids produced (see Table 2 ) contained significant quantities of the FA Δ9 C18:1, rendering them as ideal candidates for the synthesis of 2nd generation biodiesel [ 6 , 9 ] and implying the potential of their subsequent utilization as a starting material in chemo-enzymatic syntheses [ 32 ], specifically if microbial lipids are produced when low-cost compounds are implicating as starting materials of the bioprocesses. Poly-unsaturated cellular FAs were not detected in high concentrations.…”

“…Intra-cellular polysaccharides theoretically are accumulated inside the yeast cells through similar biochemical mechanisms with these of cellular lipids (nitrogen limitation can, in addition to lipid accumulation, also trigger biosynthesis and the production of endopolysaccharides [ 30 , 33 , 34 ], therefore the non-negligible accumulation of polysaccharides at the early growth steps seems as a non-expected kinetic and physiological result. Nevertheless, it must be pointed out that for several species of oleaginous yeasts (mostly belonging to R. toruloides and C. curvatus ) cultivated under conditions enabling the de novo lipid accumulation process (i.e., batch sugar- or glycerol-based nitrogen-limited ones), similar physiological behavior with increased concentrations of endopolysaccharides at the early growth phases has been reported, despite the nitrogen presence into the medium [ 12 , 14 , 32 , 35 ]. Moreover, the conversion yield of total DCW produced per unit of glucose consumed was constant throughout the culture presenting quite high values (Y X/Glc = 0.39 g/g) ( Figure 1 c), while the conversion yield of lipid produced per unit of glucose consumed was constant with a Y L/Glc value being =0.14 g/g ( Figure 1 d).…”

New trends in microbial lipid-based biorefinery for fermentative bioenergy production from lignocellulosic biomass

“…Meanwhile, optimizing oxygen and nitrogen concentrations enhances aerobic microbial lipid productivity. Including glycerol and specific amino acid substrates significantly boosts fungal and yeast lipid production yield (Karayannis et al, 2023).…”

Screening of New Industrially Important Bacterial Strains for 1,3-Propanediol, 2,3-Butanediol and Ethanol Production through Biodiesel-Derived Glycerol Fermentations

“…Among others, PDO is used for polymer production, such as polytrimethylene terephthalate (PTT), while BDO is converted after dehydration to methyl-ethyl ketone (MEK) and can be used as a fuel additive. There are several kinds of polymers, surfactants and flavoring agents deriving from glycerol valorization that can be produced following subsequent microbial and enzymatic or chemical processes [ 12 , 13 , 14 , 15 ].…”