Studying the effect of different irradiances of light on the triacylglycerol yield in the marine diatom Thalassiosira pseudonanaMegan Snyder and Sam Huynh

Background InformationT. pseudonana is classified as

phytoplankton (photosynthesizing marine organisms)◦Diatom (microscopic, unicellular,

photosynthetic algae with a silica shell called a frustule)

Bacillariophyceae class (largest group of biomass producers)100,000+ species

High oil content (21-31%) by dry weightEntire genome is sequencedOil helps it float on the water to

photosynthesizehttp://www.diark.org/img/species_pict/Thalassiosira_pseudonana_CCMP1335

Oil from diatom is a potential alternative fuel

Oil composed of lipids◦Lipid: biological compound insoluble in water

but soluble in organic solventsTriacylglycerol (specific lipid) acts

primarily as energy storage in diatoms (three fatty acids attached to one glycerol)

Triaclyglcerol is a non-polar, or neutral lipid◦Polar lipids have a hydrophilic and a

hydrophobic end; neutrals do not. ◦Polar lipids located in cell walls

Triacylglycerol Transesterificationprocess to refine triacylglycerol; how does

it become biodiesel? Oil is purified and reacted with methanol or

ethanol in the presence of a catalyst (such as KOH potassium hydroxide or NaOH sodium hydroxide); the triacylglycerol is transformed to form esters and glycerol – remaining esters are biodiesel

Triacylglycerol Chemical Formula

Glycerol 3 Fatty Acids http://www.oliveoilsource.com/page/chemical-characteristics

Nile red (9-diethylamino-5H-benzo[α]phenoxazine-5-one) is a dye that collects in the neutral lipids of living cells◦Fluoresces under a certain wavelength,

then emits another wavelength◦The intensity of the fluorescence is

directly correlated to lipid content

Purpose: increase the triacylglycerol content in the diatom using different light irradiances

Importance: increased triacylglycerol yields per diatom can make the process of making bio-fuel from algae more efficient and economically feasible

Irradiance: measure of light that reaches the product surface

Why measure in irradiance and not intensity? Photosynthesis depends on the amount of light reaching the surface of the plant, not the intensity of the source

ProcedureDiatoms will grow a plastic container

under different irradiances of fluorescent light (test one irradiance at a time)◦Sides of the container will be covered with

aluminum foil to block out excess light◦In previous experiments, irradiances of

light have affected photosynthetic processes in T. pseudonana

◦Light on a 16 hour-on, 8 hour-off cycle, maintained by a light timer

Grown in f/2 agar, a solution that provides nutrients for marine algae

pH measured periodically with a Vernier probe to make sure CO2 levels stay constant

Container inside Styrofoam box with ice packs to keep temperature constant at about 53°F

Nile red stain will be applied to a few sample diatoms after six days

Data CollectionUse UV microscope equipped with a camera

◦Takes pictures of the cells fluorescingUse Image J program to analyze level of

fluorescence

http://accu-scope.net/3032_EPI_318CU.gif

http://www.chemistrytimes.com/Images/green_alga_with_nile_red.jpg

Inverted UV microscope

Green Algae stained with Nile red- Triacylglycerol fluoresces yellow-gold, phospholipids fluoresce red

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http://chemed.chem.wisc.edu/chempaths/GenChem-Textbook/Polar-Lipids-1019.html Chemical Education Digital Library (2007 b). Nonpolar Lipids. Retrieved from:

http://chemed.chem.wisc.edu/chempaths/GenChem-Textbook/Nonpolar-Lipids-1018.html

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