Fremyella diplosiphon is an ideal third-generation biofuel source due to its ability to produce transesterified lipids. While nanofer 25s zero-valent iron nanoparticles (nZVIs) improve lipid production, an imbalance between reactive oxygen species (ROS) and cellular defense can be catastrophic to the organism. In the present study, the effect of ascorbic acid on nZVI and UV-induced stress in F. diplosiphon strain B481-SD was investigated, and lipid profiles in the combination regimen of nZVIs and ascorbic acid compared. Comparison of F. diplosiphon growth in BG11 media amended with 2, 4, 6, 8, and 10 mM ascorbic acid indicated 6 mM to be optimal for the growth of B481-SD. Further, growth in 6 mM ascorbic acid combined with 3.2 mg/L nZVIs was significantly higher when compared to the combination regimen of 12.8 and 51.2 mg/L of nZVIs and 6 mM ascorbic acid. The reversal effect of UV-B radiation for 30 min and 1 h indicated that ascorbic acid restored B481-SD growth. Transesterified lipids characterized by gas chromatography–mass spectrometry indicated C16 hexadecanoate to be the most abundant fatty acid methyl ester in the combination regimen of 6 mM ascorbic acid and 12.8 mg/L nZVI-treated F. diplosiphon. These findings were supported by microscopic observations in which cellular degradation was observed in B481-SD cells treated with 6 mM ascorbic acid and 12.8 mg/L nZVIs. Our results indicate that ascorbic acid counteracts the damaging effect of oxidative stress produced by nZVIs.
more »
« less
Nitrogen Deprivation in Fremyella diplosiphon Augments Lipid Production without Affecting Growth
Metabolic products such as lipids and proteins produced in cyanobacteria represent an
excellent source of biomass and do not compete with agricultural land use unlike soybean and
corn. Given their potential use as novel materials for biodiesel production, we aimed to explore
the e ect of cultivation period and nitrogen concentration on the growth rate and lipid content of
Fremyella diplosiphon, a model cyanobacterium. In this study, F. diplosiphon grown in BG11/HEPES
medium supplemented with 1.5 g L1 sodium nitrate (NaNO3) for 7, 10, 15, and 20 days were compared
to the untreated control in media amended with 0.25, 0.5, and 1.0 g L1 NaNO3. Cultures were
inoculated in liquid media and grown under continuous fluorescent light in an orbital incubator shaker, and extracted lipids subjected to gravimetric analysis and gas chromatography-mass spectroscopy to determine the best culture conditions for lipid production. Our results demonstrated that a reduction in nitrogen concentration had no significant effect on the growth rate across all cultivation periods; however, the accumulation of total lipid content was significantly influenced by nitrogen concentration. A maximum lipid production (40%) with no reduction in growth was observed in 10-day old cultures in a BG11/HEPES medium supplemented with 1.0 g L1 NaNO3. Fatty acid methyl ester composition of transesterified lipids demonstrated high amounts of methyl palmitate (50–70%) followed by methyl octadecenoate (17–30%) in the accumulated lipids at all treatments. Trace quantities of methyl dodecanoate, methyl hexadecanoate, methyl octadecanoate, and methyl octadecadienoate (1–8%) were also observed in all tested samples, indicating that nitrogen deprivation in culture media increases lipid production without affecting growth.
more »
« less
- Award ID(s):
- 1900966
- PAR ID:
- 10248770
- Date Published:
- Journal Name:
- Energies
- ISSN:
- 1996-1073
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
More Like this
-
-
more » « less
Abstract With the dramatic decrease in fossil fuel stocks and their detrimental effects on the environment, renewable energy sources have gained imminent importance in the mitigation of emissions. As lipid-enriched energy stocks, cyanobacteria are the leading group of microorganisms contributing to the advent of a new energy era. In the present study, the impact of Nanofer 25 s nanoscale zero-valent iron nanoparticles (nZVIs) and ampicillin on lipid production and cellular structural changes in
Fremyella diplosiphon strain B481-SD were investigated. Total lipid abundance, fatty acid methyl ester (FAME) compositions, and alkene production as detected by high-resolution two-dimensional gas chromatography with time-of-flight mass spectrometry (GC × GC/TOF–MS) was significantly higher (p < 0.05) in the individual application of 0.8 mg/L ampicillin, 3.2 mg/L nZVIs, and a combined regimen of 0.8 mg/L ampicillin and 3.2 mg/L nZVIs compared to the untreated control. In addition, we identified significant increases (p < 0.05) in monounsaturated fatty acids (MUFAs) inF. diplosiphon treated with the combination regimen compared to the untreated control, 0.8 mg/L of ampicillin, and 3.2 mg/L of nZVIs. Furthermore, individual treatment with 0.8 mg/L ampicillin and the combination regimen (0.8 mg/L ampicillin + 3.2 mg/L nZVIs) significantly increased (p < 0.05) Nile red fluorescence compared to the untreated control, indicating neutral membrane lipids to be the main target of ampicillin added treatments. Transmission electron microscopy studies revealed the presence of single-layered thylakoid membranes in the untreated control, while complex stacked membranes of 5–8 layers were visualized in ampicillin and nZVI-treatedF. diplosiphon . Our results indicate that nZVIs in combination with ampicillin significantly enhanced total lipids, essential FAMEs, and alkenes inF. diplosiphon . These findings offer a promising approach to augment the potential of using the strain as a large-scale biofuel agent. -
In humans, dietary polyunsaturated fatty acids (PUFAs) are involved in therapeutic processes such as prevention and treatment of cardiovascular diseases, neuropsychiatric disorders, and dementia. We examined the physiology, PUFA accumulation and glycerol lipid biosynthesis in the marine microalga Nannochloropsis salina in response to constant suboptimal temperature (<20 °C). As expected, N. salina exhibited significantly reduced growth rate and photosynthetic activity compared to optimal cultivation temperature. Total fatty acid contents were not significantly elevated at reduced temperatures. Cultures grown at 5 °C had the highest quantity of eicosapentanoic acid (EPA) (C20:5n3) and the lowest growth rate. Additionally, we monitored broadband lipid composition to model the occurrence of metabolic alteration and remodeling for various lipid pools. We focused on triacylglycerol (TAG) with elevated PUFA content. TAGs with EPA at all three acyl positions were higher at a cultivation temperature of 15 °C. Furthermore, monogalactosyldiacylglycerol and digalactosyldiacylglycerol, which are polar lipids associated with chloroplast membranes, decreased with reduced cultivation temperatures. Moreover, gene expression analysis of key genes involved in Kennedy pathway for de novo TAG biosynthesis revealed bimodal variations in transcript level amongst the temperature treatments. Collectively, these results show that Nannochloropsis salina is a promising source of PUFA containing lipids.more » « less
-
Dr. Krishna Ganesh (Ed.)Nanoscale zero-valent iron nanoparticles (nZVIs) are known to boost biomass production and lipid yield in Fremyella diplosiphon, a model biodiesel-producing cyanobacterium. However, the impact of nZVI-induced reactive oxygen species (ROS) in F. diplosiphon has not been evaluated. In the present study, ROS in F. diplosiphon strains (B481-WT and B481-SD) generated in response to nZVI-induced oxidative stress were quantified and the enzymatic response determined. Lipid peroxidation as a measure of oxidative stress revealed significantly higher malondialdehyde content (p < 0.01) in both strains treated with 3.2, 12.8, and 51.2 mg L–1 nZVIs compared to untreated control. In addition, ROS in all nZVI-treated cultures treated with 1.6–25.6 mg L–1 nZVIs was significantly higher than the untreated control as determined by the 2′,7′-dichlorodihydrofluorescein diacetate fluorometric probe. Immunodetection using densitometric analysis of iron superoxide dismutase (SOD) revealed significantly higher SOD levels in both strains treated with nZVIs at 51.2 mg L–1. In addition, we observed significantly higher (p < 0.001) SOD levels in the B481-SD strain treated with 6.4 mg L−1 nZVIs compared to 3.2 mg L–1 nZVIs. Validation using transmission electron microscopy equipped with energy-dispersive X-ray spectroscopy (EDS) revealed adsorption of nZVIs with a strong iron peak in both B481-WT and B481-SD strains. While the EDS spectra showed strong signals for iron at 4 and 12 days after treatment, a significant decrease in peak intensity was observed at 20 days. Future efforts will be aimed at studying transduction mechanisms that cause metabolic and epigenetic alterations in response to nZVIs in F. diplosiphon.more » « less
-
more » « less
Abstract Engineered cyanobacterium
Synechococcus elongatus can use light and CO2to produce sucrose, making it a promising candidate for use in co-cultures with heterotrophic workhorses. However, this process is challenged by the mutual stresses generated from the multispecies microbial culture. Here we demonstrate an ecosystem whereS. elongatus is freely grown in a photo-bioreactor (PBR) containing an engineered heterotrophic workhorse (either β-carotene-producingYarrowia lipolytica or indigoidine-producingPseudomonas putida ) encapsulated in calcium-alginate hydrogel beads. The encapsulation prevents growth interference, allowing the cyanobacterial culture to produce high sucrose concentrations enabling the production of indigoidine and β-carotene in the heterotroph. Our experimental PBRs yielded an indigoidine titer of 7.5 g/L hydrogel and a β-carotene titer of 1.3 g/L hydrogel, amounts 15–22-fold higher than in a comparable co-culture without encapsulation. Moreover,13C-metabolite analysis and protein overexpression tests indicated that the hydrogel beads provided a favorable microenvironment where the cell metabolism inside the hydrogel was comparable to that in a free culture. Finally, the heterotroph-containing hydrogels were easily harvested and dissolved by EDTA for product recovery, while the cyanobacterial culture itself could be reused for the next batch of immobilized heterotrophs. This co-cultivation and hydrogel encapsulation system is a successful demonstration of bioprocess optimization under photobioreactor conditions.
- Free Publicly Accessible Full Text
-
Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.3390/en13215769
