Oleaginous Microorganisms Research Articles

Purification and characterization of isocitrate dehydrogenase from Mortierella alpina

Isocitrate dehydrogenase (IDH) in oleaginous microorganisms plays an essential role in fatty acid synthesis. We expressed and purified an IDH isozyme from the oleaginous fungus Mortierella alpina (MaIDH4) and identified its enzymatic properties. MaIDH4 is NADP+-dependent and has no detectable activity with NAD+. It exerts its maximum activity when Mn2+ is used as a cofactor, followed by Mg2+. The optimum temperatures was 60 °C and 65 °C, and the optimum pH was 8.5 and 9.5, in the presence of Mn2+ and Mg2+, respectively. The catalytic efficiency of MaIDH4 for NADP+ was 16.33 (Mn2+) and 3.86 (Mg2+) μM−1s−1, while that for isocitrate was 3.06 (Mn2+) and 0.65 (Mg2+) μM−1s−1. Characterization of the enzymatic properties of MaIDH4 provides a basis for further understanding lipid synthesis in M. alpina.

Overexpression of the Mitochondrial Malic Enzyme Genes (malC and malD) Improved the Lipid Accumulation in Mucor circinelloides WJ11

Mucor circinelloides serves as a model organism to investigate the lipid metabolism in oleaginous microorganisms. It is considered as an important producer of γ-linolenic acid (GLA) that has vital medicinal benefits. In this study, we used WJ11, a high lipid-producing strain of M. circinelloides (36% w/w lipid, cell dry weight, CDW), to examine the role in lipid accumulation of two mitochondrial malic enzyme (ME) genes malC and malD. The homologous overexpression of both malC and malD genes enhanced the total lipid content of WJ11 by 41.16 and 32.34%, respectively. In parallel, the total content of GLA was enhanced by 16.73 and 46.76% in malC and malD overexpressing strains, respectively, because of the elevation of total lipid content. The fact that GLA content was enhanced more in the strain with lower lipid content increase and vice versa, indicated that engineering of mitochondrial MEs altered the fatty acid profile. Our results reveal that mitochondrial ME plays an important role in lipid metabolism and suggest that future approaches may involve simultaneous overexpression of distinct ME genes to boost lipid accumulation even further.

Bioconversion of agro-industrial by-products into microbial lipids and bioemulsifier by Absidia cylindrospora var. cylindrospora UCP 1301

Large amounts of agro-industrial by-products and wastes are generated in Brazil annually. Often, they are underutilized and improperly disposed, causing environmental pollution. In this context, reuse them as raw material to obtain high-added value products, becomes a promising strategy. Hence, this study aimed to investigate the biotechnological potential of the Mucoralean fungus Absidia cylindrospora UCP 1301 for production of oleaginous biomass and bioemulsifier (BE), using agro-industrial by-products as low-cost substrates. Firstly, the morphological identification of the strain was performed for confirmation. Then, fermentations were carried out, according to a 2³ full-factorial design (FFD) in order to evaluate the influence of substrates concentrations on production of biomass, total lipids and BE. The fungus was identified as Absidia cylindrospora var. cylindrospora and the results showed it as an oleaginous microorganism, since it accumulated lipids above 20 % of dry weight. The highest biomass production (11.38 g/L) was found in condition 3 of the FFD, while the highest lipid yield (32.22%) was reached in assay 6. Moreover, this strain was confirmed as a promising BE-producing microorganism, because of it achieved EI24 values above 50% in all conditions of the FFD. The highest EI24 with motor oil (93.3%) was obtained in condition 7 and this emulsion remained stable after 150 days of incubation. Statistical analysis showed that the three agro-industrial by-products had a significant influence on the production of biomass, lipids and BE, confirming the suitability of unconventional substrates for obtaining both microbial oil and emulsifier, which makes this bioprocess attractive for several industries.

Eco-friendly production of nutraceutical and cosmeceutical fatty acids by oleaginous fungus Lichtheimia hyalospora UCP 1266 using renewable substrates

Polyunsaturated fatty acids (PUFAs) are essential for human functions and can be used in nutraceuticals; however, traditional sources are environmentally unsustainable. A promising strategy to reduce the production costs of PUFAs is the use of economical agro-industrial sources as substrates in culture media for oleaginous microorganisms. In this study, we investigated the potential of the fungus Lichtheimia hyalospora UCP 1266 (isolated from the Caatinga soil, Brazil) in the production of PUFAs, through the metabolic conversion of alternative substrates crude glycerol (CG) and corn steep liquor (CSL). The cultivation of L. hyalospora UCP 1266 in medium containing glucose (Synthetic Medium for Mucorales - SMM) yielded 2.1 g/L of biomass and 0.6 g/L of total lipids. However, assay 4 of the 22 full-factorial design, with CG 8% and CSL 8%, provided higher yields of biomass (15.5 g/L) and total lipids (12.8 g/L). The lipids produced in this medium presented a composition of saturated, monounsaturated and polyunsaturated fatty acids (SFAs=35.13%, MUFAs=46.09% and PUFAs=18.14%). The microorganism proved to be promising in the production of omega 6 (linoleic acid, C18:2), γ-linolenic acid (C18:3), omega 3 (α-linolenic acid, C18:3) and omega 9 (oleic acid, C18:1). The results indicated L. hyalospora UCP 1266 as an oleaginous fungus with great potential for use in the development of nutraceutical ingredients for food or applied in cosmeceuticals and/or nutricosmetics.

Impact of brewery wastewater inhibitors in pure and mixed cultures of the yeast Rhodosporidium toruloides NCYC 921 and the microalga Tetradesmus obliquus ACOI 204/07

Brewery wastewater (BWW) is an appealing low-cost substrate for the production of single cell oils by oleaginous microorganisms. However, it may contain inhibitor compounds that may affect the microbial metabolism. This work investigated, for the first time, the presence of potential inhibitor compounds in primary brewery wastewater (PBWW) and secondary brewery wastewater (SBWW) for the pure and mixed cultivation of the yeast Rhodosporidium toruloides NCYC 921 and the microalga Tetradesmus obliquus ACOI 204/07. Three organic acids (OrgAc) were identified in the brewery effluents (acetic, propionic and butyric acids). Yeast and microalga pure and mixed cultivations were performed in PBWW and SBWW in order to understand the behaviour of the microorganisms, individually and together. Flow cytometry (FC) was used to monitor each microbial population during the mixed cultivations, and to study the yeast and microalga cell viability throughout all cultivations. The yeast cells in pure cultures grown in both effluents were severely affected by the OrgAc presence confirmed by the cell stress results obtained by FC. However, in the mixed cultures, the yeast cells were able to develop, and the levels of stress conditions were considerably lower. Only in microalga pure and mixed cultures efficient OrgAc removal was observed.

Optimization of cis-9-Heptadecenoic Acid Production from the Oleaginous Yeast Yarrowia lipolytica

Odd-chain fatty acids (OCFA) have been studied for their therapeutic and nutritional properties, as well as for their potential use in the chemical industry for the production of biofuel. Genetic modification strategies have demonstrated an improved production of OCFA by oleaginous microorganisms. In this study, the production of OCFA-enriched lipids by fermentation using a genetically engineered Yarrowia lipolytica strain was investigated. The major fatty acid produced by this strain was the cis-9-heptadecenoic acid (C17:1). Its biosynthesis was optimized using a design of experiment strategy involving a central composite design. The optimal responses maximizing the cell density (optical density at 600 nm) and the C17:1 content (%) in lipids were found using 52.4 g/L sucrose, 26.9 g/L glycerol, 10.4 g/L sodium acetate, 5 g/L sodium propionate, and 4 g/L yeast extract. Under these conditions, in a 5 L scale bioreactor, the respective contents of lipids and C17:1 in culture medium were 2.52 ± 0.05 and 0.82 ± 0.01 g/L after 96 h fermentation. The results obtained in this work pave the way toward the process upscale of C17:1 and encourage its industrial production.

The relationship between amino acid and lipid metabolism in oleaginous eukaryotic microorganism.

Amino acids are the building blocks of protein, promoting the balance between growth and lipid synthesis. However, the accumulation of microbial lipids involves multiple pathways, which requires the analysis of the global cellular metabolic network in which amino acid metabolism is involved. This review illustrates the dependence patterns of intracellular amino acids and lipids of oleaginous eukaryotic microorganisms in different environments and points out the contribution of amino acid metabolic precursors to the de novo synthesis of fatty acids. We emphasized the key role of amino acid metabolism in lipid remodeling and autophagy behavior and highlighted the regulatory effects of amino acids and their secondary metabolites as signal factors for microbial lipid synthesis. The application prospects of omics technology and genetic engineering technology in the field of microbial lipids are described. KEY POINTS: • Overview of microbial lipid synthesis mediated by amino acid metabolism • Insight into metabolic mechanisms founding multiple regulatory networks is provided • Description of microbial lipid homeostasis mediated by amino acid excitation signal.

New insights into phenotypic heterogeneity for the distinct lipid accumulation of Schizochytrium sp. H016

BackgroundSchizochytrium sp. is a marine heterotrophic protist and an important sustainable resource for high value-added docosahexaenoic acid in the future. The production of different phenotypes during the continuous subculture of Schizochytrium sp. results in a serious reduction in lipid yield and complicates the used of this strain in scientific research and industrial production. Hence, obtaining an improved understanding of the phenotypic differences and molecular mechanisms underlying the cell-to-cell heterogeneity of Schizochytrium sp. is necessary.ResultsAfter continuous culture passage, Schizochytrium sp. H016 differentiated into two subpopulations with different morphologies and showed decreased capacity for lipid production. The presence of cell subpopulations with degraded lipid droplets led to a substantial decrease in overall lipid yield. Here, a rapid screening strategy based on fluorescence-activated cell sorting was proposed to classify and isolate subpopulations quickly in accordance with their lipid-producing capability. The final biomass and lipid yield of the subpopulation with high cell lipid content (i.e., H016-H) were 38.83 and 17.22 g/L, respectively, which were 2.07- and 5.38-fold higher than those of the subpopulation with low lipid content (i.e., H016-L), respectively. Subsequently, time‑resolved transcriptome analysis was performed to elucidate the mechanism of phenotypic heterogeneity in different subpopulations. Results showed that the expression of genes related to the cell cycle and lipid degradation was significantly upregulated in H016-L, whereas the metabolic pathways related to fatty acid synthesis and glyceride accumulation were remarkably upregulated in H016-H.ConclusionThis study innovatively used flow cytometry combined with transcriptome technology to provide new insights into the phenotypic heterogeneity of different cell subpopulations of Schizochytrium sp. Furthermore, these results lay a strong foundation for guiding the breeding of oleaginous microorganisms with high lipid contents.

Overexpression of Shinorhizobium meliloti flavohemoglobin improves cell growth and fatty acid biosynthesis in oleaginous fungus Mucor circinelloides.

Oxygen availability is a limiting factor for lipid biosynthesis in eukaryotic microorganisms. Two bacterial hemoglobins from Vitreoscilla sp. (VHb) and Shinorhizobium meliloti (SHb), which deliver oxygen to the respiratory chain to produce more ATP, were introduced into Mucor circinelloides to alleviate oxygen limitation, thereby improving cell growth and fatty acid production. The VHb and SHb genes were integrated into the M. circinelloides MU402 genome by homologous recombination. VHb and SHb protein expression was verified by carbon monoxide difference spectrum analysis. The biomass was increased by ~ 50% in the strain expressing SHb compared with VHb. The total fatty acid (TFA) content of the strain expressing SHb reached 15.7% of the dry cell weight (~ 40% higher than that of the control strain) during flask cultivation. The biomass and TFA content were markedly increased (12.1g/L and 21.1% dry cell weight, respectively) in strains expressing SHb than strains expressing VHb during fermenter cultivation. VHb and SHb expression also increased the proportion of polyunsaturated fatty acids. Overexpressed bacterial hemoglobins, especially SHb, increased cell growth and TFA content in M. circinelloides at low and high aeration, suggesting that SHb improves fatty acid production more effectively than VHb in oleaginous microorganisms.

Production of single cell oil by Yarrowia lipolytica JCM 2320 using detoxified desiccated coconut residue hydrolysate.

Nowadays, the replacement of petro-diesel with biodiesel has raised the concern among the community for the utilization of improper feedstocks and the cost involved. However, these issues can be solved by producing single cell oil (SCO) from lignocellulosic biomass hydrolysates by oleaginous microorganisms. This study introduced Yarrowia lipolytica JCM 2320 with a desiccated coconut residue (DCR) hydrolysate (obtained from the 2% dilute sulphuric acid pretreatment) as a carbon source in generating SCO. However, common inhibitors formed during acid pretreatment of biomass such as five-hydroxymethylfurfural (HMF), furfural, acetic acid and levulinic acid resulting from the sugar degradations may have detrimental effects towards the fermentation process. To visualize the effect of inhibitors on Y. lipolytica, an inhibitory study was conducted by adding 0.5–5.0 g/L of potential inhibitors to the YPD (yeast, peptone and D-glucose) medium. It was found that the presence of furfural at 0.5 g/L would increase the lag phase, which beyond that was detrimental to Y. lipolytica. Furthermore, increasing the five-hydroxymethylfurfural (HMF) concentration would increase the lag phase of Y. lipolytica, whereas, for acetic acid and levulinic acid, it showed a negligible effect. Detoxification was hence conducted to remove the potential inhibitors from the DCR hydrolysate prior its utilization in the fermentation. To examine the possibility of using adsorption resins for the detoxification of DCR hydrolysate, five different resins were tested (Amberlite® XAD-4, Amberlite® XAD-7, Amberlite® IR 120, Amberlite® IRA 96 and Amberlite® IRA 402) with five different concentrations of 1%, 3%, 5%, 10% and 15% (w/v), respectively. At resin concentration of 10%, Amberlite® XAD-4 recorded the highest SCO yield, 2.90 ± 0.02 g/L, whereas the control and the conventional overliming detoxification method, recorded only 1.29 ± 0.01 g/L and 1.27 ± 0.02 g/L SCO accumulation, respectively. Moreover, the fatty acid profile of the oil produced was rich in oleic acid (33.60%), linoleic acid (9.90%), and palmitic acid (14.90%), which indicates the potential as a good biodiesel raw material.

Lipid production by oleaginous microorganisms using food wastes: a review

Food wastes are rich in nutrients and can be used for producing useful chemicals through biotransformation. Some oleaginous microorganisms can use food wastes to produce lipids and high value-added metabolites such as polyunsaturated fatty acids, squalene, and carotenoids. This not only reduces the production cost, but also improves the economic value of the products, thus has large potential for commercial production. This review summarized the advances in food waste treatment, with a focus on the lipid production by oleaginous microorganisms using food wastes. Moreover, challenges and future directions were prospected with the aim to provide a useful reference for related researchers.

Isolation, molecular identification of lipid-producing Rhodotorula diobovata: optimization of lipid accumulation for biodiesel production

BackgroundThe increased demand for oil and fats to satisfy the ever-increasing human needs has enhanced the research in this field. Single-cell oils or microbial lipids produced by oleaginous microorganisms are being utilized as an alternative to traditional oil sources. Oleaginous yeasts can accumulate lipids above 20% of their biomass when they are grown under controlled conditions. ResultsIn the present study, sixty-five yeasts were isolated from different sources. Using Sudan Black B staining technique, five yeast isolates were selected. Under nitrogen-limited cultivation conditions, the Co1 isolate was the best lipid accumulation potential of 39.79%. Isolate (Co1) was characterized morphologically and identified using the ribosomal DNA internal transcribed spacers regions (rDNA-ITS) from their genomic DNA. The sequence alignment revealed a 99.2% similarity with Rhodotorula diobovata. Under the optimized conditions, Rhodotorula diobovata accumulated lipids up to 45.85% on a dry biomass basis. R. diobovata, when grown on different raw materials, accumulated lipid up to 46.68% on sugar beet molasses medium, and the lipid had a high degree of monounsaturated fatty acids which gives biodiesel better quality. ConclusionsThe data suggest that the potent oleaginous yeast, R. diobovata, together with the use of cheap feedstock raw materials such as sugar beet molasses, can be considered as a promising feedstock for biodiesel production.

Autophagy Improves ARA-Rich TAG Accumulation in Mortierella alpina by Regulating Resource Allocation.

ABSTRACTThe present study was designed to explore the possibility of improving lipid production in oleaginous filamentous fungus Mortierella alpina based on an autophagy regulation strategy. According to multiomics information, vacuolate-centered macroautophagy was identified as the main type of autophagy in M. alpina under nitrogen-limited conditions. Mutation of autophagy-related gene MAatg8 led to impaired fatty acid synthesis, while overexpression of both MAatg8 and phosphatidylserine decarboxylases (MApsd2) showed promoting effects on fatty acid synthesis. MAatg8 overexpression strain with external supply of ethanolamine significantly increased arachidonic acid (ARA)-rich triacylglycerol (TAG) and biomass synthesis in M. alpina, and the final fatty acid content increased by approximately 110% compared with that in the wild-type strain. Metabolomics and lipidomics analyses revealed that cell autophagy enhanced the recycling of preformed carbon, nitrogen, and lipid in mycelium, and the released carbon skeleton and energy were contributed to the accumulation of TAG in M. alpina. This study suggests that regulation of autophagy-related MAatg8-phosphatidylethanolamine (MAatg8-PE) conjugation system could be a promising strategy for attaining higher lipid production and biomass growth. The mechanism of autophagy in regulating nitrogen limitation-induced lipid accumulation elucidated in this study provides a reference for development of autophagy-based strategies for improving nutrient use efficiency and high value-added lipid production by oleaginous microorganism.IMPORTANCE Studies have indicated that functional oil accumulation occurs in oleaginous microorganisms under nitrogen limitation. However, until now, large-scale application of nitrogen-deficiency strategies was limited by low biomass. Therefore, the identification of the critical nodes of nitrogen deficiency-induced lipid accumulation is urgently needed to further guide functional oil production. The significance of our research is in uncovering the function of cell autophagy in the ARA-rich TAG accumulation of oleaginous fungus M. alpina and demonstrating the feasibility of improving lipid production based on an autophagy regulation strategy at the molecular and omics levels. Our study proves that regulation of cell autophagy through the MAatg8-PE conjugation system-related gene overexpression or exogenous supply of ethanolamine would be an efficient strategy to increase and maintain biomass productivity when high TAG content is obtained under nitrogen deficiency, which could be useful for the development of new strategies that will achieve more biomass and maximal lipid productivity.

Expression Profile of Selected Genes Involved in Storage Lipid Synthesis in a Model Oleaginous Yeast Species Yarrowia lipolytica.

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.

Biodiesel production from lignocellulosic biomass using Yarrowia lipolytica

• Sugarcane bagasse hydrolysates was used as a bio resource for lipid production. • Acid, alkaline and ultra-sonication techniques were attempted in pretreatment of Sugarcane bagasse. • 16.39 g/l of biomass concentration were achieved with Sugarcane bagasse hydrolysates. • In-situ transesterification carried out with the catalyst K2CO3 yields 80% biodiesel. Depletion of hydrocarbons is forcing to find alternative resources to meet the energy demand of the growing population. Microbial biodiesel as a fuel can act as a cheaper and eco-friendly alternative to fossil fuel. Single-cell oil (SCO) consisting of carbon, hydrogen and oxygen grown over the lignocellulosic biomass using oleaginous microorganisms are triacylglycerols which can be converted to biodiesel, with physicochemical properties similar to conventional diesel. However, several cost-effective pretreatment methods are required to utilize lignocellulosic biomasses. The current research study investigates the SCO yield (and biodiesel characteristics) obtained from sugarcane bagasse hydrolysate through various pretreatment techniques. The pretreatment with 4% v/v H 2 SO 4 at 25 min of ultra-sonication provided the best depolymerisation results (based on the glucose concentration). Yarrowia lipolytica was inoculated into the hydrolysates, allowed to grow at 25 °C, pH of 6.5 and rapid mixing for six days yielded biomass of 16.39 g/l. Biodiesel was extracted from the biomass via in-situ and ex-situ transesterification. In-situ transesterification carried out with the catalyst K 2 CO 3 yields 80% biodiesel. In comparison, 63% were achieved with ex-situ transesterification, where lipid extraction was carried out as a first step and transesterified further in the presence of catalyst KOH to obtain biodiesel. The obtained fatty acid methyl esters (FAME) was subjected to FTIR analysis, and the observed physicochemical properties were within the international standards.

Metabolic Engineering and Genome Editing Strategies for Enhanced Lipid Production in Oleaginous Microorganisms

Metabolic Engineering and Genome Editing Strategies for Enhanced Lipid Production in Oleaginous Microorganisms

Isolation of oleaginous fungi from natural sources, and fatty acid profile of Cunninghamella echinulata

Oleaginous microorganisms accumulate 20% or more of their dry weight as lipid. In the present study, 247 fungal isolates were obtained from different types of soil, marine water, and food, and screened for their ability to accumulate considerable amounts of lipid. A primary screening with Nile red staining was followed by sulfo-phosphovanillin reaction. The most potent isolate was identified employing both morphological and molecular methods as the Zygomycetous fungus, Cunninghamella echinulata AUMC14396. The lipid of this fungus was extracted, transesterified, and analyzed via GC-MS to elucidate the fatty acid (FA) composition. The fatty acid profile revealed the presence of various fatty acids with different concentrations. Of the total fatty acids, saturated FAs comprised 44.49%, while monounsaturated FAs were 19.91%, and polyunsaturated fatty acids (PUFAs) were 35.61%. The most predominant species was linoleic acid (13.79%) followed by stearic (12.78%) and oleic (12.50%) acids. The polyunsaturated fatty acid, gamma linolenic acid was present in a considerable amount (8.02%). Other important PUFAs were also found in the fatty acid profile such as arachidonic acid, eicosapentaenoic acid, and docosahexaenoic acid. The results of this study were discussed in respect to the important applications in the context of production of pharmaceuticals and nutraceuticals.

Comparison of Ex-Situ and In-Situ Transesterification for the Production of Microbial Biodiesel

Microbial biodiesel is converted from microbial lipids via transesterification process. Most microbial biodiesel studies are focusing on the use of microalgal lipids as feedstock. Apart from using microalgae for lipid biosynthesis, lipids can also be extracted from other oleaginous microorganisms like fungi and yeast. However, there are gaps in the studies of lipid production from filamentous fungi, especially in-situ transesterification process. The aim of this project is to compare in-situ with the ex-situ transesterification of fungal biomass from Aspergillus oryzae. In ex-situ transesterification, two methods of lipid extraction, the Soxhlet extraction and the Bligh and Dyer extraction, were performed. For in-situ transesterification, two methods using different catalysts were investigated. Base-catalyzed in-situ transesterification of fungal biomass resulted on the highest Fatty Acid Methyl Esters (FAME) yield. The base-catalyzed in-situ transesterification was further optimized via Central Composite Design (CCD) of Response Surface Methodology (RSM). The parameters investigated were the catalyst loading, methanol to biomass ratio and reaction time. The optimization showed that the highest FAME yield was at 25.1% (w/w) with 10 minutes reaction time, 5% catalyst and 360:1 of the ratio of the methanol to biomass. Based on Analysis of Variance (ANOVA), the model was found to be significant according to the value of “Prob >F” of 0.0028. Copyright © 2021 by Authors, Published by BCREC Group. This is an open access article under the CC BY-SA License (https://creativecommons.org/licenses/by-sa/4.0).

Strategy to increase the lipid stability of the microbial oil produced by Umbelopsis isabellina for food purposes: Use of microencapsulation by external ionic gelation

Oleaginous microorganisms, including the fungus Umbelopsis isabellina, have emerged as a biotechnological alternative to obtain polyunsaturated fatty acid-rich oils, which are strongly linked to energy purposes (biofuel) than the food industry. Considering the composition of microbial oil and its use by the food industry, it is necessary to investigate strategies that increase its lipid stability. Ergo, this pioneering study aimed to microencapsulate the oil produced by Umbelopsis isabellina and evaluate its oxidative stability throughout the storage period against factors such as temperature and luminosity. The microbial oil was microencapsulated through the external ionic gelation technique, producing an encapsulation efficiency of 80% and proving to be a suitable method because it maintained oil composition. Combining microencapsulation and refrigerated storage led to the best effects on storage time, increasing the evaluated lipid stability through the peroxide values and conjugated diene formation. Moreover, saturated and monounsaturated fatty acid content increased, and polyunsaturated fatty acid content decreased during storage for both the free and microencapsulated oil, regardless of storage temperature, although microencapsulation reduced the changes. The results primarily demonstrate how microencapsulation prolongs the oxidative stability and unsaturated fatty acid content of the microbial oil by reducing its reactions to external environmental factors, thus facilitating its use in the food industry.

Phosphorus and Nitrogen Limitation as a Part of the Strategy to Stimulate Microbial Lipid Biosynthesis

Microbial lipids called a sustainable alternative to traditional vegetable oils invariably capture the attention of researchers. In this study, the effect of limiting inorganic phosphorus (KH2PO4) and nitrogen ((NH4)2SO4) sources in lipid-rich culture medium on the efficiency of cellular lipid biosynthesis by Y. lipolytica yeast has been investigated. In batch cultures, the carbon source was rapeseed waste post-frying oil (50 g/dm3). A significant relationship between the concentration of KH2PO4 and the amount of lipids accumulated has been revealed. In the shake-flask cultures, storage lipid yield was correlated with lower doses of phosphorus source in the medium. In bioreactor culture in mineral medium with (g/dm3) 3.0 KH2PO4 and 3.0 (NH4)2SO4, the cellular lipid yield was 47.5% (w/w). Simultaneous limitation of both phosphorus and nitrogen sources promoted lipid accumulation in cells, but at the same time created unfavorable conditions for biomass growth (0.78 gd.m./dm3). Increased phosphorus availability with limited cellular access to nitrogen resulted in higher biomass yields (7.45 gd.m./dm3) than phosphorus limitation in a nitrogen-rich medium (4.56 gd.m./dm3), with comparable lipid yields (30% and 32%). Regardless of the medium composition, the yeast preferentially accumulated oleic and linoleic acids as well as linolenic acid up to 8.89%. Further, it is crucial to determine the correlation between N/P molar ratios, biomass growth and efficient lipid accumulation. In particular, considering the contribution of phosphorus as a component of coenzymes in many metabolic pathways, including lipid biosynthesis and respiration processes, its importance as a factor in the cultivation of the oleaginous microorganisms was highlighted.