Production of single cell oil by Lipomyces starkeyi from waste plant oil generated by the palm oil mill industry.

Only a few reports available about the assimilation of hydrophobic or oil-based feedstock as carbon sources by Lipomyces starkeyi. In this study, the ability of L. starkeyi to efficiently utilize free fatty acids (FFAs) and real biomass like palm acid oil (PAO) as well as crude palm kernel oil (CPKO) for growth and lipid production was investigated. PAO, CPKO, and FFAs were evaluated as sole carbon sources or in the mixed medium containing glucose. L. starkeyi was able to grow on the medium supplemented with PAO and FFAs, which contained long-chain length FAs and accumulated lipids up to 35% (w/w) of its dry cell weight. The highest lipid content and lipid concentration were achieved at 50% (w/w) and 10.1 g/L, respectively, when L. starkeyi was cultured in nitrogen-limited mineral medium (-NMM) supplemented with PAO emulsion. Hydrophobic substrate like PAO could be served as promising carbon source for L. starkeyi.

Obtaining hemicellulosic hydrolysate from sugarcane bagasse for microbial oil production by Lipomyces starkeyi.

OBJECTIVE: The extraction of the hemicellulose fraction of sugarcane bagasse (SCB) by acid hydrolysis was evaluated in an autoclave and a Parr reactor aiming the application of the hydrolysate as a carbon source for lipid production by Lipomyces starkeyi. RESULTS: The hydrolysis that resulted in the highest sugar concentration was obtained by treatment in the Parr reactor (HHR) at 1.5% (m/v) H2SO4 and 120 °C for 20 min, reaching a hemicellulose conversion of approximately 82%. The adaptation of the yeast to the hydrolysate provided good fermentability and no lag phase. The fermentation of hemicellulose-derived sugars (HHR) by L. starkeyi resulted in a 27.8% (w/w) lipid content and YP/S of 0.16 g/l.h. Increasing the inoculum size increased the lipid content by approximately 61%, reaching 44.8% (w/w). CONCLUSION: The hemicellulose hydrolysate from SCB is a potential substrate for L. starkeyi to produce lipids for biodiesel synthesis based on the biorefinery concept.

Lipid production by Lipomyces starkeyi using sap squeezed from felled old oil palm trunks.

The ability of oleaginous yeast Lipomyces starkeyi to efficiently produce lipids when cultivated on sap extracted from felled oil palm trunk (OPT) as a novel inexpensive renewable carbon source was evaluated. OPT sap was found to contain approximately 98 g/L glucose and 32 g/L fructose. Batch fermentations were performed using three different OPT sap medium conditions: regular sap, enriched sap, and enriched sap at pH 5.0. Under all sap medium conditions, the cell biomass and lipid production achieved were approximately 30 g/L and 60% (w/w), respectively. L. starkeyi tolerated acidified medium (initial pH ≈ 3) and produced considerable amounts of ethanol as well as xylitol as by-products. The fatty acid profile of L. starkeyi was remarkably similar to that of palm oil, one of the most common vegetable oil feedstock used in biodiesel production with oleic acid as the major fatty acid followed by palmitic, stearic and linoleic acids.

The algal trophic mode affects the interaction and oil production of a synergistic microalga-yeast consortium.

The use of non-food feedstocks to produce renewable microbial resources can limit our dependence on fossil fuels and lower CO2 emissions. Since microalgae display a virtuous CO2 and O2 exchange with heterotrophs, the microalga Chlamydomonas reinhardtii was combined with the oleaginous yeast Lipomyces starkeyi, known for their production of oil, base material for biodiesel. The coupled growth was shown to be synergistic for biomass and lipid production. The species were truly symbiotic since synergistic growth occurred even when the alga cannot use the organic carbon in the feedstock and in absence of air, thus depending entirely on CO2-O2 exchange. Since addition of acetate as the algal carbon source lowered the performance of the consortium, the microbial system design should take into account algal mixotrophy. The mixed biomass was found be suitable for biodiesel production, and whereas lipid production increased in the consortium, yields should be improved in future studies.

Lipomyces starkeyi: an emerging cell factory for production of lipids, oleochemicals and biotechnology applications.

Oils and oleochemicals produced by microbial cells offer an attractive alternative to petroleum and food-crop derived oils for the production of transport fuel and oleochemicals. An emerging candidate for industrial single cell oil production is the oleaginous yeast Lipomyces starkeyi. This yeast is capable of accumulating storage lipids to concentrations greater than 60% of the dry cell weight. From the perspective of industrial biotechnology L. starkeyi is an excellent chassis for single-cell oil and oleochemical production as it can use a wide variety of carbon and nitrogen sources as feedstock. The strain has been used to produce lipids from hexose and pentose sugars derived from cellulosic hydrolysates as well as crude glycerol and even sewage sludge. L. starkeyi also produces glucanhydrolases that have a variety of industrial applications and displays potential to be employed for bioremediation. Despite its excellent properties for biotechnology applications, adoption of L. starkeyi as an industrial chassis has been hindered by the difficulty of genetically manipulating the strain. This review will highlight the industrial potential of L. starkeyi as a chassis for the production of lipids, oleochemicals and other biochemicals. Additionally, we consider progress and challenges in engineering this organism for industrial applications.

Characterization of oil-producing yeast Lipomyces starkeyi on glycerol carbon source based on metabolomics and 13C-labeling.

Lipomyces starkeyi is an oil-producing yeast that can produce triacylglycerol (TAG) from glycerol as a carbon source. The TAG was mainly produced after nitrogen depletion alongside reduced cell proliferation. To obtain clues for enhancing the TAG production, cell metabolism during the TAG-producing phase was characterized by metabolomics with 13C labeling. The turnover analysis showed that the time constants of intermediates from glycerol to pyruvate (Pyr) were large, whereas those of tricarboxylic acid (TCA) cycle intermediates were much smaller than that of Pyr. Surprisingly, the time constants of intermediates in gluconeogenesis and the pentose phosphate (PP) pathway were large, suggesting that a large amount of the uptaken glycerol was metabolized via the PP pathway. To synthesize fatty acids that make up TAG from acetyl-CoA (AcCoA), 14 molecules of nicotinamide adenine dinucleotide phosphate (NADPH) per C16 fatty acid molecule are required. Because the oxidative PP pathway generates NADPH, this pathway would contribute to supply NADPH for fatty acid synthesis. To confirm that the oxidative PP pathway can supply the NADPH required for TAG production, flux analysis was conducted based on the measured specific rates and mass balances. Flux analysis revealed that the NADPH necessary for TAG production was supplied by metabolizing 48.2% of the uptaken glycerol through gluconeogenesis and the PP pathway. This result was consistent with the result of the 13C-labeling experiment. Furthermore, comparison of the actual flux distribution with the ideal flux distribution for TAG production suggested that it is necessary to flow more dihydroxyacetonephosphate (DHAP) through gluconeogenesis to improve TAG yield.

Effect of inoculum size on single-cell oil production from glucose and xylose using oleaginous yeast Lipomyces starkeyi.

Oleaginous microbes can convert substrates such as carbon dioxide, sugars, and organic acids to single-cell oils (SCOs). Among the oleaginous microorganisms, Lipomyces starkeyi is a particularly well-suited host given its impressive native abilities, including the capability to utilize a wide variety of carbon sources. In this work, the potential of L. starkeyi NBRC10381 to produce SCOs in a synthetically nitrogen-limited mineral medium (-NMM) was investigated by differing the inoculum size using glucose and/or xylose as a carbon source. Fermentation using glucose and xylose as mixed carbon sources generated the highest production of biomass at 40.8 g/L, and achieved a lipid content of 84.9% (w/w). When either glucose or xylose was used separately, the totals for achieved lipid content were 79.6% (w/w) and 85.1% (w/w), respectively. However, biomass production was higher for glucose than for xylose (30.3 vs. 28.7 g/L, respectively). This study describes the first simultaneous achievement of higher levels of cell mass and lipid production using glucose and/or xylose as the carbon sources in different inoculum sizes.

Potential biodegradation of crude petroleum oil by newly isolated halotolerant microbial strains from polluted Red Sea area.

Two microbial isolates from oil polluted Red Sea water in Egypt, designated as RS-Y1 and RS-F3, were found capable of degrading Belayim mix (BX) crude oil. Strains RS-Y1 and RS-F3 were assigned to the genera Lipomyces tetrasporus and Paecilomyces variotii based on their morphological and physiological characteristics. Both isolates were compared for the biodegradation of crude petroleum-oil hydrocarbons in basal salt medium supplemented with 5% (w/v) of BX-crude oil. Gas chromatography profile showed that the biodegradation of total petroleum hydrocarbons (TPHs) inoculated with L. tetrasporus (68.3%) and P. variotii (58.15%) along with their consortium (66%) significantly reduced TPHs levels as compared to the control after 30days. L. tetrasporus (44.5%) was more effective than P. variotii strain (32.89%) in reducing the unresolved complex mixtures (UCM) content from the medium. Both isolates exhibited a strong growth over a wide range of salinity (5-45g/L NaCl).

Production of single cell oil from Lipomyces starkeyi ATCC 56304 using biorefinery by-products.

Single cell oil (SCO) is a valuable noncrop-based renewable oil source. Hemicellulose derived sugars can be utilized to produce SCO using the oleaginous yeast Lipomyces starkeyi ATCC 56304. Bran by-products were tested as hemicellulose-rich feedstocks for the production of SCO. Whole and destarched corn and wheat bran hydrolysates were produced using hydrothermal and dilute sulfuric acid (0%, 0.5%, 1.0%, v/v) pretreatment along with enzymatic hydrolysis. Whole bran hydrolysates produced from hydrothermal pretreatment generated the highest average oil yields of 126.7 and 124.3 mg oil/g sugar for both wheat and corn bran, respectively. 1.0% acid pretreatment was effective for the destarched bran generating a hemicellulose hydrolysis efficiency of 94% and 84% for wheat and corn bran, respectively, resulting in the highest oil yield of 70.7 mg oil/g sugar. The results indicate pretreated corn and wheat bran hydrolysates can serve as viable feedstocks for oleaginous yeast SCO bioconversion.

Bioconversion of corncob acid hydrolysate into microbial oil by the oleaginous yeast Lipomyces starkeyi.

For the first time, corncob acid hydrolysate was used for microbial oil production by the oleaginous yeast Lipomyces starkeyi. After hydrolysis by dilute sulfuric acid, corncob could turn into an acid hydrolysate with a sugar concentration of about 42.3 g/L. Detoxified by overliming and absorption with activated carbon, the corncob hydrolysate could be used by L. starkeyi efficiently that a total biomass of 17.2 g/L with a lipid content of 47.0 % (corresponding to a lipid yield of 8.1 g/L) and a lipid coefficient of 20.9 could be obtained after cultivation on the corncob hydrolysate for 8 days. Therefore, L. starkeyi is a promising strain for microbial oil production from lignocellulosic biomass. Glucose and xylose were used by L. starkeyi simultaneously during lipid fermentation while arabinose could not be utilized by it. Besides, the lipid composition of L. starkeyi was similar to that of vegetable oils; thus, it is a promising feedstock for biodiesel production.

Phylogenetic and biochemical characterization of the oil-producing yeast Lipomyces starkeyi.

Lipomyces starkeyi is an oleaginous yeast, and has been classified in four distinct groups, i.e., sensu stricto and custers α, ß, and γ. Recently, L. starkeyi clusters α, ß, and γ were recognized independent species, Lipomyces mesembrius, Lipomyces doorenjongii, and Lipomyces kockii, respectively. In this study, we investigated phylogenetic relationships within L. starkeyi, including 18 Japanese wild strains, and its related species, based on internal transcribed spacer sequences and evaluated biochemical characters which reflected the phylogenetic tree. Phylogenetic analysis showed that most of Japanese wild strains formed one clade and this clade is more closely related to L. starkeyi s.s. clade including one Japanese wild strain than other clades. Only three Japanese wild strains were genetically distinct from L. starkeyi. Lipomyces mesembrius and L. doorenjongii shared one clade, while L. kockii was genetically distinct from the other three species. Strains in L. starkeyi s.s. clade converted six sugars, D-glucose, D-xylose, L-arabinose, D-galactose, D-mannose, and D-cellobiose to produce high total lipid yields. The Japanese wild strains in subclades B, C, and D converted D-glucose, D-galactose, and D-mannose to produce high total lipid yields. Lipomyces mesembrius was divided into two subclades. Lipomyces mesembrius CBS 7737 converted D-xylose, L-arabinose, D-galactose, and D-cellobiose, while the other L. mesembrius strains did not. Lipomyces doorenjongii converted all the sugars except D-cellobiose. In comparison to L. starkeyi, L. mesembrius, and L. doorenjongii, L. kockii produced higher total lipid yields from D-glucose, D-galactose, and D-mannose. The type of sugar converted depended on the subclade classification elucidated in this study.

Microbial conversion of olive oil mill wastewaters into lipids suitable for biodiesel production.

Lipomyces starkey were able to survive and proliferate in the presence of olive oil mill wastewaters (OMW), a medium difficult to process by biological treatments, due to the antimicrobial activities of their phenolic components. The microorganisms were grown in the presence of undiluted OMW, without external organic supplements, producing a significant reduction of both the total organic carbon (TOC) and the total phenols content. The OMW treated by L. starkey showed a significant increase of the germination index. The preliminary dilution of OMW enhanced the reduction of polluting components of OMW, leading to a complete TOC removal, as well as to lower levels of residual phenols. The activities of extracellular lipases and esterases significantly increased in the course of the OMW fermentation. A significant increase in lipid yield was observed in L. starkey in the course of the OMW treatment, particularly enhanced when the feedstock was preliminarily diluted. The fatty acid distribution showed a prevalence of oleic acid, demonstrating the potential of L. starkeyi as a source of lipids to be used as a feedstock for the synthesis of II generation biodiesel.