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.

Lipid Productivity and Biosynthesis Genes Response of Indigenous Chlorella sp. T4 Strain under Different Nitrogen and Phosphorus Load

Microalgae can synthesize and accumulate high neutral lipids upon exposure to abiotic stress such as nutrient starvation or limitation. In this study, indigenous microalgae Chlorella sp. T4 was cultivated in nitrogen and phosphorus under both limiting and replete conditions. Growth, lipid yield, fatty acid profiles and biosynthetic gene expression levels were determined to ascertain cell’s response under these conditions. An impaired cell growth was observed under nitrogen limiting condition, evident by the lowest biomass yield (0.58±0.03 g L−1) as revealed by low quantum efficiency of photosystem II (Fv/Fm) value and chlorophyll a content. An increase in lipid content yield was observed under nitrogen and phosphorus limiting conditions as compared to the control. Nutrient limiting conditions produced fatty acid methyl ester that is suitable for biodiesel production compared to the control (BG-11). Gene expression analysis using real time q-PCR for photosynthesis (rbcL) and lipid biosynthesis (accD, KAS-1, ω-6 FAD, ω-3 FAD) genes revealed different expression levels under both limiting and replete conditions. Under nutrient limiting conditions, increase in the expression of accD, KAS-1, ω-6 FAD and ω-3 FAD genes was observed, whereas a decrease in rbcL gene expression level was noted. A significant correlation could be drawn between the expression levels of the biosynthetic genes and growth rate, biomass yield, physiological response, lipid yield and fatty acid composition. These results provide an insight into the physiological response and gene expression level under different nutrient levels, which could be harnessed for future genetic engineering of Chlorella sp. T4 for improved lipid production.

Strategies for Improvement of Lipid Production by Yeast Trichosporon oleaginosus from Lignocellulosic Biomass

Microbial lipids have similar fatty acid composition to plant oils, and therefore, are considered as an alternative feedstock for biodiesel production. Oleaginous yeasts accumulate considerable amounts of lipids intracellularly during growth on low-cost renewable feedstocks such as lignocellulosic biomass. In this study, we cultivated yeast Trichosporon oleaginosus on hydrolysate of alkaline pretreated corn cobs. Different process configurations were evaluated and compared, including separate hydrolysis and fermentation (SHF) with cellulase recycle and simultaneous saccharification and fermentation (SSF) in batch and fed-batch mode. At low enzyme loading, the highest lipid concentration of 26.74 g L−1 was reached in fed-batch SSF fed with 2.5% (g g−1) substrate. Batch SHF was conducted for four rounds with recycling the cellulase adsorbed on unhydrolyzed lignocellulosic biomass. Thirty percent of cellulase saving was achieved for rounds 2–4 without compromising productivity and lipid yield. The addition of Tween 80 to lignocellulosic slurry improved the hydrolysis rate of structural carbohydrates in pretreated lignocellulosic biomass. Furthermore, supplementing the growth medium with Tween 80 improved lipid yield and productivity without affecting yeast growth. Oleaginous yeast T. oleaginosus is a promising strain for the sustainable and efficient production of lipids from renewable lignocellulosic feedstock.

Optimisation of Healthy-Lipid Content and Oxidative Stability during Oil Extraction from Squid (Illex argentinus) Viscera by Green Processing

Green extraction was applied to Argentinean shortfin squid (Illex argentinus) viscera, consisting of a wet pressing method including a drying step, mechanic pressing, centrifugation of the resulting slurry, and oil collection. To maximise the oil yield and ω3 fatty acid content and to minimise the oil damage degree, a response surface methodology (RSM) design was developed focused on the drying temperature (45–85 °C) and time (30–90 min). In general, an increase of the drying time and temperature provided an increase in the lipid yield recovery from the viscera. The strongest drying conditions showed a higher recovery than 50% when compared with the traditional chemical method. The docosahexaenoic and eicosapentaenoic acid contents in the extracted oil revealed scarce dependence on drying conditions, showing valuable ranges (149.2–166.5 and 88.7–102.4 g·kg−1 oil, respectively). Furthermore, the values of free fatty acids, peroxides, conjugated dienes, and ω3/ω6 ratio did not show extensive differences by comparing oils obtained from the different drying conditions. Contrary, a polyene index (PI) decrease was detected with increasing drying time and temperature. The RSM analysis indicated that optimised drying time (41.3 min) and temperature (85 °C) conditions would lead to 74.73 g·kg−1 (oil yield), 1.87 (PI), and 6.72 (peroxide value) scores, with a 0.67 desirability value.

Analyzing and optimizing the carbon utilization and lipid yield at different light intensities for Scenedesmus arcuatus

Microalgae, the photosynthetic microorganism growing abundantly in marine and aquatic ecosystems, are potential source for biological sequestration of CO2. The carbon uptake differs in the presence of other nutrients, light intensity etc. The biomass yield of Scenedesmus arcuatus var capitatus was studied based on the Face Centred Central Composite design (FCCD) of Response Surface Methodology (RSM) for nitrate, phosphate and carbonate under different conditions (laboratory, room and sunlight conditions). Various pre-treatments (osmotic shock, autoclaving, microwave and ultrasonication) were employed to find the best method for maximum lipid yield. The biomass yield reached a maximum of 1 g/L under sunlight conditions of nitrate concentration 500 ppm and carbonate 2000 ppm. The laboratory conditions resulted in a biomass yield of 0.59 g/L at 500 ppm nitrate, 1000 ppm carbonate and 250 ppm phosphate. Under room conditions, the yield was very low (0.11 g/L). Osmotic shock resulted in higher lipid yield than the other pre-treatment methods. The ability of Scenedesmus arcuatus to uptake high carbon under sunlight conditions and to adapt to high light intensity and fluctuations in light intensity concludes that this species is suitable for large-scale open pond cultivation for CO2 sequestration and production of metabolites.

Optimization of a novel lipid extraction process from microalgae

Previous study found that the solvent extraction efficiency of lipid in microalgae could be greatly improved by washing algae cells before the second time extraction. Based on the "organic solvents–water–organic solvents" method, this research further studied the effect of four solvent systems (acetone, chloroform/methanol, chloroform/methanol/water, dichloromethane/methanol), two types of water treatment (vortex and ultrasonic), three water treatment time gradient (0 s, 30 s, 120 s) on the lipid extraction at three different microalgae growth stages (3rd day, 5th day, 9th day). The results show that the combination of water treatment type, treatment time and solvent is very important to the efficiency of lipid extraction. The total lipid extracted was generally increased by 10–30% after water treatment. Especially under the condition of 120 s vortex water treatment with dichloromethane/methanol as extraction solvent, the total lipid extracted increased by 61.14%. In addition, microalgae cells at different culture stages had different sensitivity to water treatment. In this study, under the combination of chloroform/methanol/water as extraction solvent and vortex water treatment for 120 s, the highest lipid yield was obtained on the ninth day of cell culture, which accounts 47.88% of the cell dry weight (478 mg/g cell dry weight). The changes of cell morphology and structure after water treatment were studied by scanning electron microscope, and it was found that water treatment could seriously destroy the cell membrane damaged by solvent, thus promoting the release of lipids. This study further optimizes the "solvent–water–solvent" lipid extraction method, which neither produces impurities nor damages the lipid quality, and can reduce the amount of organic solvent applied in the classical lipid extraction method with the same lipid yield, so it has a broad application prospect.

Lipid yields of algae dried in an enhanced solar chimney

This work focuses on the lipid yield of algae dried using an enhanced solar chimney dryer with the aim towards high quality biodiesel production and low energy consumption. Jaworski’s medium was chosen for the cultivation in this research project as it can provide enough nutrients to green algae of type Chlorella. Sp. Centrifugation is an effective method to harvest the algae from its medium prior to a drying process. In this project, the methods used for drying were oven drying, open-sun drying and an enhanced solar chimney drying. The moisture content was determined where the average moisture content were 82.5% for oven drying method, 81.6% for open sun drying and 82.2% for solar chimney drying. Methods were found to affect differently the algae properties in terms of the lipid yield and mineral content. The lipid yields were 23.7%, for oven drying, 20.6% for open sun drying and 24.4% for enhanced solar chimney drying. While the oven drying was found to be the fastest way to dry the algae, the solar chimney drying proved best in energy saving while producing the same amount or more algae lipid within reasonable drying times.