High-value Lipids Research Articles

Accumulation of high-value lipids in single-cell microorganisms: a mechanistic approach and future perspectives.

In recent years attention has been focused on the utilization of microorganisms as alternatives for industrial and nutritional applications. Considerable research has been devoted to techniques for growth, extraction, and purification of high-value lipids for their use as biofuels and biosurfactants as well as high-value metabolites for nutrition and health. These successes argue that the elucidation of the mechanisms underlying the microbial biosynthesis of such molecules, which are far from being completely understood, now will yield spectacular opportunities for industrial scale biomolecular production. There are important additional questions to be solved to optimize the processing strategies to take advantage of the assets of microbial lipids. The present review describes the current state of knowledge regarding lipid biosynthesis, accumulation, and transport mechanisms present in single-cell organisms, specifically yeasts, microalgae, bacteria, and archaea. Similarities and differences in biochemical pathways and strategies of different microorganisms provide a diverse toolset to the expansion of biotechnologies for lipid production. This paper is intended to inspire a generation of lipid scientists to insights that will drive the biotechnologies of microbial production as uniquely enabling players of lipid biotherapeutics, biofuels, biomaterials, and other opportunity areas into the 21st century.

More than biofuels – Potential uses of microalgae as sources of high‐value lipids

AbstractCultivation of microalgae has potential in producing a feedstock for biofuels and, in addition, high‐value lipid bioproducts such as long‐chain (⪈C20, LC) omega‐3 oils, carotenoid pigments and squalene, as well as other non‐lipid materials including exopolysaccharide. The high‐value LC omega‐3 oils have been conventionally sourced from fish. This paper provides a perspective on our recent study of the heterotrophic growth of thraustochytrids for their wider biotechnological potential, including as a source of a range of higher value lipids in addition to biofuels.

Wax ester profiling of seed oil by nano-electrospray ionization tandem mass spectrometry

BackgroundWax esters are highly hydrophobic neutral lipids that are major constituents of the cutin and suberin layer. Moreover they have favorable properties as a commodity for industrial applications. Through transgenic expression of wax ester biosynthetic genes in oilseed crops, it is possible to achieve high level accumulation of defined wax ester compositions within the seed oil to provide a sustainable source for such high value lipids. The fatty alcohol moiety of the wax esters is formed from plant-endogenous acyl-CoAs by the action of fatty acyl reductases (FAR). In a second step the fatty alcohol is condensed with acyl-CoA by a wax synthase (WS) to form a wax ester. In order to evaluate the specificity of wax ester biosynthesis, analytical methods are needed that provide detailed wax ester profiles from complex lipid extracts.ResultsWe present a direct infusion ESI-tandem MS method that allows the semi-quantitative determination of wax ester compositions from complex lipid mixtures covering 784 even chain molecular species. The definition of calibration prototype groups that combine wax esters according to their fragmentation behavior enables fast quantitative analysis by applying multiple reaction monitoring. This provides a tool to analyze wax layer composition or determine whether seeds accumulate a desired wax ester profile. Besides the profiling method, we provide general information on wax ester analysis by the systematic definition of wax ester prototypes according to their collision-induced dissociation spectra. We applied the developed method for wax ester profiling of the well characterized jojoba seed oil and compared the profile with wax ester-accumulating Arabidopsis thaliana expressing the wax ester biosynthetic genes MaFAR and ScWS.ConclusionsWe developed a fast profiling method for wax ester analysis on the molecular species level. This method is suitable to screen large numbers of transgenic plants as well as other wax ester samples like cuticular lipid extracts to gain an overview on the molecular species composition. We confirm previous results from APCI-MS and GC-MS analysis, which showed that fragmentation patterns are highly dependent on the double bond distribution between the fatty alcohol and the fatty acid part of the wax ester.

Supercritical Carbon Dioxide Extraction of Corn Distiller’s Dried Grains with Solubles: Experiments and Mathematical Modeling

Corn distiller's dried grains with solubles (DDGS) is a byproduct of the ethanol industry and has potential as a source of valuable compounds. In this study, corn DDGS was extracted using supercritical carbon dioxide (SC-CO(2)) at 50-70 °C, 34.5-49.6 MPa, and constant CO(2) flow rate of 1 L/min (measured at ambient conditions). The highest yield of total lipids (9.2%, w/w) was obtained at 49.6 MPa/70 °C. Apparent solubility of corn DDGS lipids ranged between 0.010 kg/kg CO(2) at 34.5 MPa/50 °C and 0.026 kg/kg CO(2) at 49.6 MPa/70 °C. The extract contained 107 mg/kg carotenoids, 1538 mg/kg tocochromanols, and 15904 mg/kg phytosterols at 49.6 MPa/70 °C. The Sovova model and Chrastil model were successfully used to describe the extraction of total lipids and apparent solubility of total and minor lipids, respectively. The study revealed that DDGS is a good inexpensive source of lipids and valuable minor lipid components and that SC-CO(2) extraction can be used as a "green" process to add value to corn DDGS by recovering such high-value lipids.

Genome, Functional Gene Annotation, and Nuclear Transformation of the Heterokont Oleaginous Alga Nannochloropsis oceanica CCMP1779

Unicellular marine algae have promise for providing sustainable and scalable biofuel feedstocks, although no single species has emerged as a preferred organism. Moreover, adequate molecular and genetic resources prerequisite for the rational engineering of marine algal feedstocks are lacking for most candidate species. Heterokonts of the genus Nannochloropsis naturally have high cellular oil content and are already in use for industrial production of high-value lipid products. First success in applying reverse genetics by targeted gene replacement makes Nannochloropsis oceanica an attractive model to investigate the cell and molecular biology and biochemistry of this fascinating organism group. Here we present the assembly of the 28.7 Mb genome of N. oceanica CCMP1779. RNA sequencing data from nitrogen-replete and nitrogen-depleted growth conditions support a total of 11,973 genes, of which in addition to automatic annotation some were manually inspected to predict the biochemical repertoire for this organism. Among others, more than 100 genes putatively related to lipid metabolism, 114 predicted transcription factors, and 109 transcriptional regulators were annotated. Comparison of the N. oceanica CCMP1779 gene repertoire with the recently published N. gaditana genome identified 2,649 genes likely specific to N. oceanica CCMP1779. Many of these N. oceanica–specific genes have putative orthologs in other species or are supported by transcriptional evidence. However, because similarity-based annotations are limited, functions of most of these species-specific genes remain unknown. Aside from the genome sequence and its analysis, protocols for the transformation of N. oceanica CCMP1779 are provided. The availability of genomic and transcriptomic data for Nannochloropsis oceanica CCMP1779, along with efficient transformation protocols, provides a blueprint for future detailed gene functional analysis and genetic engineering of Nannochloropsis species by a growing academic community focused on this genus.

A Lipid Droplet Protein of Nannochloropsis with Functions Partially Analogous to Plant Oleosins

As our understanding of the dynamics of lipid droplets (LDs) in animal, plant, and fungal cells is rapidly evolving, still little is known about the formation and turnover of these organelles in microalgae. Yet with the growing importance of algal feedstock for the production of biofuels and high-value lipids, there is a need to understand the mechanisms of LD dynamics in microalgae. Thus, we investigated the proteins associated with LDs of the emerging heterokont model alga Nannochloropsis sp. and discovered an abundant hydrophobic lipid droplet surface protein (LDSP) with unique primary sequence but structural similarities to other LD proteins. LDSP abundance in Nannochloropsis cells closely tracked the amount of triacylglycerols during conditions of oil accumulation and degradation. Functional characterization of LDSP in an Arabidopsis (Arabidopsis thaliana) OLEOSIN1-deficient mutant allowed a separation of its physical and structural properties in its interaction with LDs from its physiological or biochemical activities. Although LDSP presence in Arabidopsis predictably affected LD size, it could not reverse the physiological impact of OLEOSIN deficiency on triacylglycerol hydrolysis during germination.

Addition of an N-terminal epitope tag significantly increases the activity of plant fatty acid desaturases expressed in yeast cells

Saccharomyces cerevisiae shows great potential for development of bioreactor systems geared toward the production of high-value lipids such as polyunsaturated omega-3 fatty acids, the yields of which are largely dependent on the activity of ectopically expressed enzymes. Here, we show that the addition of an N-terminal epitope tag sequence (either Myc or hemagglutinin) to oleate desaturase (FAD2) or omega-3 linoleate desaturase (FAD3) enzymes from plants, which catalyze consecutive reactions in the production of long chain omega-3 fatty acids, significantly increases their activity up to fourfold when expressed in yeast cells. Quantitative protein blotting using an antibody specific for native FAD2 revealed that the steady-state amount of the epitope-tagged FAD2 protein was also approximately fourfold higher than that of its untagged counterpart, demonstrating a direct relationship between the epitope tag-induced increase in enzyme amount and fatty acid product formation. Protein half-life and RNA blotting experiments indicated that the half-lives and mRNA content of the tagged and untagged FAD2 proteins were essentially the same, suggesting that the epitope tags increased protein abundance by improving translational efficiency. Taken together, these results indicate that the addition of an epitope tag sequence to a plant fatty acid desaturase (FAD) not only provides a useful means for protein immunodetection using highly specific, commercially available antibodies, but that it also significantly increases FAD activity and the production of polyunsaturated fatty acids in yeast cells.

Extraction of Amaranth seed oil by supercritical carbon dioxide

The present work studied various aspects of extracting high value lipids from a model seed material ( Amaranth cruentus) using supercritical carbon dioxide as solvent. Experimentally, it investigated the effects of process temperature, pressure, solvent flow rate, scale of extraction, and sample pre-treatment on the extraction rate and yield. It was found that the rate of extraction was function of the solvent flow rate, while the yield depended on the flow rate, pre-treatment, temperature and pressure. The solubility of Amaranth seed oil (regarded as a pseudo-component) was found to increase with temperature at high pressures and decrease with increasing temperature at lower pressures. Mathematical modelling of the solubility and the extraction process are discussed.

Production of lipid compounds in the yeast Saccharomyces cerevisiae.

This review describes progress using the yeast Saccharomyces cerevisiae as a model organism for the fast and efficient analysis of genes and enzyme activities involved in the lipid biosynthetic pathways of several donor organisms. Furthermore, we assess the impact of baker's yeast on the production of novel, high-value lipid compounds. Yeast can be genetically modified to produce selected substances in relatively high amounts. A major advantage in choosing yeast as an object for metabolic engineering is the fact that the lipid pathways in this organism have been described in detail and are well characterized. We focus on the de novo production of three major families of lipid products. These are: (1) sterols, providing some previously known and some novel applications as examples of the lipid pathway enhancement that occurs naturally in yeast, (2) the reconstitution of the biosynthetic pathway of steroid hormones and (3) the biosynthesis of polyunsaturated fatty acids, leading to the biosynthesis of different omega-3 and omega-6 fatty acids which do not occur naturally in yeast. We utilize the current knowledge and point out perspectives and problems for future biotechnological applications in the field of lipid compounds.

Phosphatidylinositol-3-kinase pathway is stimulated by fungal lipid containing cocoa butter equivalents (CBE) in FMet-Leu-Phe (FMLP)- and phorbolester (PMA)-challenged human neutrophils.

It is well established that fungi produce fats and oils, although a commercial utilization of these processes with rare exception could not yet be achieved. The reason hereto lied primarily on the high costs of production. Recently, a novel biotechnological process for the production of fungal lipids from Mucor circinelloides by using acetic acid (a cheap carbon source) at sub-toxical levels as the starting substrate has been described (1). The products obtained were oil- and butter-like compounds depending upon the dissolved oxygen tension maintained (Figure 1). The analyses of the lipid demonstrated a γ-linolenic acid (GLA)-rich lipid (so-called fungal oil) and a stearic acid-rich lipid (so-called fungal butter) (2). This first report of low cost production of high value lipids thus raised a caveat for more consideration.KeywordsCocoa ButterHuman NeutrophilDissolve Oxygen TensionCocoa Butter EquivalentMucor CircinelloidesThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Hydroxy long-chain fatty acids in fungi

Hydroxy long-chain fatty acids occur widely in animals and plants and have important physiological activities in these eukaryotes. There are indications that these compounds are also common and important in fungi. The occurrence of hydroxy-polyunsaturated fatty acids (hydroxy-PUFAs) is of biotechnological importance, because these compounds are potentially high-value lipid products with medical applications. This review pays particular attention to the production of hydroxy-PUFAs by yeasts and other fungi. Hydroxy-PUFAs derived from lipoxygenase activity appear to be present in most fungi, while hydroxy-PUFAs from cyclooxygenase activity (i.e. prostaglandins) have mainly been implicated in the Oomycota and in yeasts from the genus Dipodascopsis. The occurrence of other hydroxy long-chain fatty acids in fungi is also discussed briefly; these include hydroxy fatty acids that are generally associated with cytochrome P-450 monooxygenase activity (i.e. terminal and sub-terminal hydroxy acids and diols derived from the corresponding epoxides) as well as 2-hydroxy-fatty acids and 3-hydroxy-fatty acids.