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Preliminary Analysis of the Source and Destination of Carbon in Phaeodactlyum tricornutum under Nitrogen Deprivation. Steven Baldassano. AMOPs in Biofuels. AMOP = Aquatic Microbial Oxygenic Photoautotroph Includes cyanobacteria, algae, diatoms High neutral lipid yield (20-50% of dry weight) - PowerPoint PPT Presentation
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Steven Baldassano
AMOPs in BiofuelsAMOP = Aquatic Microbial Oxygenic
PhotoautotrophIncludes cyanobacteria, algae, diatomsHigh neutral lipid yield (20-50% of dry weight)
Potentially more useful for biofuels than current sources such as corn grain ethanolMore efficient conversion of solar energyUse of little or no landEasier conversion of lipids to useful biofuelsUseful byproducts such as protein for animal feed
PurposeDetermination of the effects of nitrogen
deprivation on Phaeodactylum tricornutumInability to create proteins will change carbon
compositionMeasurement of carbohydrate/protein/lipid
composition
Use of C13 labeling to determine source and distribution of carbon
Experimental Setup
•Cultures of Phaeodactylum tricornutum grown for 4 days in F/2 media with sufficient N and C12
•On day 4, cells were pelleted and resuspended in C13 labeled bicarbonate in sealed flasks
•3 in F/2 media•3 in F/2 media without NO3-
•N2 bubbling used to evacuate flasks
•Assays performed on day 4 (t=0 timepoint) and day 7 (t=3 days after resuspension).
MethodspHCell count by microscopyOptical density at 625 nmChlorophyll contentβ-1,3-glucan (carbohydrate storage)
C13 content by mass spectroscopyMembrane carbohydratesProtein contentLipid Content
Fractionation and C13 content using LCMSAsh weight percent
ResultsN- cells became much larger and more ovoidSignificantly lower cell count in N- samples
due to arrested cell divisionDecreased chlorophyll content in N- samples
Decrease in pH relative to N+ samples
Optical density readings correlate well with cell count and dry weight for N+ samples only
ResultsN-, ash-free dry weight %
N+, ash-free dry weight %
ResultsOverall lipid increase after resuspension:
Ongoing ResearchCompletion of lipid content analysis
FractionationC13 labeling
Mass spectroscopy for carbohydrate C13
content
C13 content will provide information as to the source of carbon for glucan and lipid increaserearrangement vs de novo synthesis
SummaryNitrogen deprivation resulted in changes in
compositionIncrease in cell size and massIncrease in glucan contentIncrease in lipid content (per cell and dry weight)Slight decrease in overall lipids per mlDecrease in membrane carbohydrate contentDecrease in protein content
Source of carbon for lipid and glucan increase to be determined soon