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Microbial Lipids: Production, Characterization and Applications

A special issue of International Journal of Molecular Sciences (ISSN 1422-0067). This special issue belongs to the section "Molecular Microbiology".

Deadline for manuscript submissions: closed (28 February 2023) | Viewed by 15505

Special Issue Editors


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Guest Editor
INRA INA-PG CNRS, Centre de Biotechnologie Agro-Industrielle, F-78850 Thiverval-Grignon, France
Interests: lipid accumulation; microbial genetics; metabolic engineering; biotechnology of yeasts; algal engineering
Special Issues, Collections and Topics in MDPI journals

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Guest Editor
Colin Ratledge Center for Microbial Lipids, School of Agriculture Engineering and Food Sciences, Shandong University of Technology, 266 Xincun Rd., Zibo, Shandong, China
Interests: lipid biochemistry; functional lipid biosynthesis

Special Issue Information

Dear Colleagues,

Oleaginous microorganisms possess the ability to accumulate lipids in excess of 20% of their dry biomass. Under nutrient limitations, oleaginous microorganisms, such as fungi (molds and yeasts), microalgae, and to some extent bacteria, produce single cell oils (SCOs) with varying fatty acid profiles, making them highly suitable for industrial applications. SCOs have been proven to be highly beneficial and lucrative biotechnological products, especially polyunsaturated fatty acids (ω-3 and ω-6) similar to that of plant oils with the advantages of short cultivation times and high product purity. Researches are invited focussing on molecular studies of lipid accumulation in oleaginous microorganisms, metabolic regulation of single desired fatty acids with high cell densities, cellular mechanism of lipid accumulation under varying culture conditions, fermentation strategies, and genetic modifications for enhancing lipid yield and valuable fatty acid content. Moreover, for the extraction of lipids at the cellular level, a multitude of cell disruption and extraction methods could also directly influence subsequent downstream operations. The optimal strategies have to be elucidated with each production strain separately.  In addition, applications of microbial lipids for the health and physical wellbeing of humans in food industries or the bio-based economy need to be explored. Papers related to molecular sciences of lipid accumulation accompanied with biotechnology, metabolic engineering, fermentation engineering, and processing of microbial lipids will be considered for this Special Issue.

Dr. Jean-Marc Nicaud
Prof. Dr. Yuanda Song
Guest Editors

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Keywords

  • molecular lipogenesis
  • oleaginous microorganism
  • single cell oils
  • polyunsaturated fatty acids
  • fermentation
  • biofuels
  • microalgae
  • lipid accumulation

Published Papers (7 papers)

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Research

19 pages, 3851 KiB  
Article
Simultaneous Lipid and Carotenoid Production via Rhodotorula paludigena CM33 Using Crude Glycerol as the Main Substrate: Pilot-Scale Experiments
by Cheeranan Sriphuttha, Pailin Boontawan, Pasama Boonyanan, Mariena Ketudat-Cairns and Apichat Boontawan
Int. J. Mol. Sci. 2023, 24(24), 17192; https://doi.org/10.3390/ijms242417192 - 6 Dec 2023
Cited by 3 | Viewed by 1226
Abstract
Rhodotorula paludigena CM33 is an oleaginous yeast that has been demonstrated to accumulate substantial quantities of intracellular lipids and carotenoids. In this study, crude glycerol, a by-product of biodiesel production, was used as a carbon source to enhance the accumulation of lipids and [...] Read more.
Rhodotorula paludigena CM33 is an oleaginous yeast that has been demonstrated to accumulate substantial quantities of intracellular lipids and carotenoids. In this study, crude glycerol, a by-product of biodiesel production, was used as a carbon source to enhance the accumulation of lipids and carotenoids in the cells. The culture conditions were first optimized using response surface methodology, which revealed that the carotenoid concentration and lipid content improved when the concentration of crude glycerol was 40 g/L. Different fermentation conditions were also investigated: batch, repeated-batch, and fed-batch conditions in a 500 L fermenter. For fed-batch fermentation, the maximum concentrations of biomass, lipids, and carotenoids obtained were 46.32 g/L, 37.65%, and 713.80 mg/L, respectively. A chemical-free carotenoid extraction method was also optimized using high-pressure homogenization and a microfluidizer device. The carotenoids were found to be mostly beta-carotene, which was confirmed by HPLC (high pressure liquid chromatography), LC-MS (liquid chromatography-mass spectrometry), and NMR (nuclear magnetic resonance). The results of this study indicate that crude glycerol can be used as a substrate to produce carotenoids, resulting in enhanced value of this biodiesel by-product. Full article
(This article belongs to the Special Issue Microbial Lipids: Production, Characterization and Applications)
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16 pages, 8098 KiB  
Article
Lipid Membrane Remodeling by the Micellar Aggregation of Long-Chain Unsaturated Fatty Acids for Sustainable Antimicrobial Strategies
by Sungmin Shin, Hyunhyuk Tae, Soohyun Park and Nam-Joon Cho
Int. J. Mol. Sci. 2023, 24(11), 9639; https://doi.org/10.3390/ijms24119639 - 1 Jun 2023
Cited by 3 | Viewed by 1789
Abstract
Antimicrobial fatty acids derived from natural sources and renewable feedstocks are promising surface-active substances with a wide range of applications. Their ability to target bacterial membrane in multiple mechanisms offers a promising antimicrobial approach for combating bacterial infections and preventing the development of [...] Read more.
Antimicrobial fatty acids derived from natural sources and renewable feedstocks are promising surface-active substances with a wide range of applications. Their ability to target bacterial membrane in multiple mechanisms offers a promising antimicrobial approach for combating bacterial infections and preventing the development of drug-resistant strains, and it provides a sustainable strategy that aligns with growing environmental awareness compared to their synthetic counterparts. However, the interaction and destabilization of bacterial cell membranes by these amphiphilic compounds are not yet fully understood. Here, we investigated the concentration-dependent and time-dependent membrane interaction between long-chain unsaturated fatty acids—linolenic acid (LNA, C18:3), linoleic (LLA, C18:2), and oleic acid (OA, C18:1)—and the supported lipid bilayers (SLBs) using quartz crystal microbalance-dissipation (QCM-D) and fluorescence microscopy. We first determined the critical micelle concentration (CMC) of each compound using a fluorescence spectrophotometer and monitored the membrane interaction in real time following fatty acid treatment, whereby all micellar fatty acids elicited membrane-active behavior primarily above their respective CMC values. Specifically, LNA and LLA, which have higher degrees of unsaturation and CMC values of 160 µM and 60 µM, respectively, caused significant changes in the membrane with net |Δf| shifts of 23.2 ± 0.8 Hz and 21.4 ± 0.6 Hz and ΔD shifts of 5.2 ± 0.5 × 10−6 and 7.4 ± 0.5 × 10−6. On the other hand, OA, with the lowest unsaturation degree and CMC value of 20 µM, produced relatively less membrane change with a net |Δf| shift of 14.6 ± 2.2 Hz and ΔD shift of 8.8 ± 0.2 × 10−6. Both LNA and LLA required higher concentrations than OA to initiate membrane remodeling as their CMC values increased with the degree of unsaturation. Upon incubating with fluorescence-labeled model membranes, the fatty acids induced tubular morphological changes at concentrations above CMC. Taken together, our findings highlight the critical role of self-aggregation properties and the degree of unsaturated bonds in unsaturated long-chain fatty acids upon modulating membrane destabilization, suggesting potential applications in developing sustainable and effective antimicrobial strategies. Full article
(This article belongs to the Special Issue Microbial Lipids: Production, Characterization and Applications)
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17 pages, 1718 KiB  
Article
Yarrowia lipolytica as a Platform for Punicic Acid Production
by Veronika Urbanikova, Young-Kyoung Park, Daniela Krajciova, Mehdi Tachekort, Milan Certik, Ioana Grigoras, Roman Holic, Jean-Marc Nicaud and Peter Gajdos
Int. J. Mol. Sci. 2023, 24(10), 8823; https://doi.org/10.3390/ijms24108823 - 16 May 2023
Cited by 3 | Viewed by 1589
Abstract
Punicic acid (PuA) is a polyunsaturated fatty acid with significant medical, biological, and nutraceutical properties. The primary source of punicic acid is the pomegranate seed oil obtained from fruits of trees that are mainly cultivated in subtropical and tropical climates. To establish sustainable [...] Read more.
Punicic acid (PuA) is a polyunsaturated fatty acid with significant medical, biological, and nutraceutical properties. The primary source of punicic acid is the pomegranate seed oil obtained from fruits of trees that are mainly cultivated in subtropical and tropical climates. To establish sustainable production of PuA, various recombinant microorganisms and plants have been explored as platforms with limited efficiencies. In this study, the oleaginous yeast Yarrowia lipolytica was employed as a host for PuA production. First, growth and lipid accumulation of Y. lipolytica were evaluated in medium supplemented with pomegranate seed oil, resulting in the accumulation of lipids up to 31.2%, consisting of 22% PuA esterified in the fraction of glycerolipids. In addition, lipid-engineered Y. lipolytica strains, transformed with the bifunctional fatty acid conjugase/desaturase from Punica granatum (PgFADX), showed the ability to accumulate PuA de novo. PuA was detected in both polar and neutral lipid fractions, especially in phosphatidylcholine and triacylglycerols. Promoter optimization for PgFADX expression resulted in improved accumulation of PuA from 0.9 to 1.8 mg/g of dry cell weight. The best-producing strain expressing PgFADX under the control of a strong erythritol-inducible promoter produced 36.6 mg/L PuA. These results demonstrate that the yeast Y. lipolytica is a promising host for PuA production. Full article
(This article belongs to the Special Issue Microbial Lipids: Production, Characterization and Applications)
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21 pages, 2017 KiB  
Article
Yeast Lipid Produced through Glycerol Conversions and Its Use for Enzymatic Synthesis of Amino Acid-Based Biosurfactants
by Dimitris Karayannis, Seraphim Papanikolaou, Christos Vatistas, Cédric Paris and Isabelle Chevalot
Int. J. Mol. Sci. 2023, 24(1), 714; https://doi.org/10.3390/ijms24010714 - 31 Dec 2022
Cited by 11 | Viewed by 2073
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 [...] Read more.
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. Full article
(This article belongs to the Special Issue Microbial Lipids: Production, Characterization and Applications)
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14 pages, 5865 KiB  
Article
The High Plasticity of Nonpathogenic Mycobacterium brumae Induces Rapid Changes in Its Lipid Profile during Pellicle Maturation: The Potential of This Bacterium as a Versatile Cell Factory for Lipid Compounds of Therapeutic Interest
by Víctor Campo-Pérez, Sandra Guallar-Garrido, Marina Luquin, Alejandro Sánchez-Chardi and Esther Julián
Int. J. Mol. Sci. 2022, 23(21), 13609; https://doi.org/10.3390/ijms232113609 - 6 Nov 2022
Cited by 1 | Viewed by 1639
Abstract
The immunomodulatory potential of mycobacteria to be used for therapeutic purposes varies by species and culture conditions and is closely related to mycobacterial lipid composition. Although the lipids present in the mycobacterial cell wall are relevant, lipids are mainly stored in intracellular lipid [...] Read more.
The immunomodulatory potential of mycobacteria to be used for therapeutic purposes varies by species and culture conditions and is closely related to mycobacterial lipid composition. Although the lipids present in the mycobacterial cell wall are relevant, lipids are mainly stored in intracellular lipid inclusions (ILIs), which have emerged as a crucial structure in understanding mycobacteria-host interaction. Little is known about ILI ultrastructure, production, and composition in nonpathogenic species. In this study, we compared the lipid profiles of the nonpathogenic immunomodulatory agent Mycobacterium brumae during pellicle maturation under different culture conditions with qualitative and quantitative approaches by using high-resolution imaging and biochemical and composition analyses to understand ILI dynamics. The results showed wax esters, mainly in early stages of development, and acylglycerols in mature ILI composition, revealing changes in dynamics, amount, and morphometry, depending on pellicle maturation and the culture media used. Low-glycerol cultures induced ILIs with lower molecular weights which were smaller in size in comparison with the ILIs produced in glycerol-enriched media. The data also indicate the simple metabolic plasticity of lipid synthesis in M. brumae, as well as its high versatility in generating different lipid profiles. These findings provide an interesting way to enhance the production of key lipid structures via the simple modulation of cell culture conditions. Full article
(This article belongs to the Special Issue Microbial Lipids: Production, Characterization and Applications)
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13 pages, 1451 KiB  
Article
Improving Lipid Production of Yarrowia lipolytica by the Aldehyde Dehydrogenase-Mediated Furfural Detoxification
by Jiwon Kim, Hyeoncheol Francis Son, Sungmin Hwang, Gyeongtaek Gong, Ja Kyong Ko, Youngsoon Um, Sung Ok Han and Sun-Mi Lee
Int. J. Mol. Sci. 2022, 23(9), 4761; https://doi.org/10.3390/ijms23094761 - 26 Apr 2022
Cited by 10 | Viewed by 2634
Abstract
Yarrowia lipolytica, the non-conventional yeast capable of high lipogenesis, is a microbial chassis for producing lipid-based biofuels and chemicals from renewable resources such as lignocellulosic biomass. However, the low tolerance of Y. lipolytica against furfural, a major inhibitory furan aldehyde derived from [...] Read more.
Yarrowia lipolytica, the non-conventional yeast capable of high lipogenesis, is a microbial chassis for producing lipid-based biofuels and chemicals from renewable resources such as lignocellulosic biomass. However, the low tolerance of Y. lipolytica against furfural, a major inhibitory furan aldehyde derived from the pretreatment processes of lignocellulosic biomass, has restricted the efficient conversion of lignocellulosic hydrolysates. In this study, the furfural tolerance of Y. lipolytica has been improved by supporting its endogenous detoxification mechanism. Specifically, the endogenous genes encoding the aldehyde dehydrogenase family proteins were overexpressed in Y. lipolytica to support the conversion of furfural to furoic acid. Among them, YALI0E15400p (FALDH2) has shown the highest conversion rate of furfural to furoic acid and resulted in two-fold increased cell growth and lipid production in the presence of 0.4 g/L of furfural. To our knowledge, this is the first report to identify the native furfural detoxification mechanism and increase furfural resistance through rational engineering in Y. lipolytica. Overall, these results will improve the potential of Y. lipolytica to produce lipids and other value-added chemicals from a carbon-neutral feedstock of lignocellulosic biomass. Full article
(This article belongs to the Special Issue Microbial Lipids: Production, Characterization and Applications)
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15 pages, 6062 KiB  
Article
Expression Profile of Selected Genes Involved in Storage Lipid Synthesis in a Model Oleaginous Yeast Species Yarrowia lipolytica
by Agata Fabiszewska, Magdalena Paplińska-Goryca, Paulina Misiukiewicz-Stępień, Małgorzata Wołoszynowska, Dorota Nowak and Bartłomiej Zieniuk
Int. J. Mol. Sci. 2022, 23(3), 1041; https://doi.org/10.3390/ijms23031041 - 18 Jan 2022
Cited by 8 | Viewed by 2890
Abstract
Yarrowia lipolytica yeast is a model species of the group of oleaginous microorganisms capable of intracellular lipids accumulation in an amount exceeding 20% of the dry mass. Single cell oil biosynthesis can follow one of two biochemical pathways—de novo accumulation of cellular lipids [...] Read more.
Yarrowia lipolytica yeast is a model species of the group of oleaginous microorganisms capable of intracellular lipids accumulation in an amount exceeding 20% of the dry mass. Single cell oil biosynthesis can follow one of two biochemical pathways—de novo accumulation of cellular lipids in medium containing non-lipid carbon sources (including saccharides, glycerol) and ex novo microbial oil synthesis which involves fatty acids uptake from the environment. The mRNA expression of selected genes of de novo and ex novo lipid synthesis pathways was analyzed and correlated with the phenotypically observed features. It was proved that the accumulation yield of storage lipids via ex novo pathway was to some extent dependent on the limitation of the nitrogen source in the medium. It was also proposed that the synthesis of intracellular lipids in lipid-rich medium proceeded mainly via ex novo pathway, although the activity of genes encoding the enzymes of the de novo pathway were not completely inhibited at the stage of transcription by fatty acids present in the medium (e.g., ATP-citrate lyase). Molecular markers of two biosynthesis routes has been outlined and a hypothetical connection point between de novo and ex novo route were indicated. Full article
(This article belongs to the Special Issue Microbial Lipids: Production, Characterization and Applications)
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