A Novel Analysis Utilizing Invasive Fermented Macroalgae

Ariana KimSaint Andrew’s PrioryHonolulu, Hawaii

Acres of beaches globally are suffering

from macroalgal infestation - millions

spent on clearing projects

Invasive macroalgae fermented,

analyzed for compounds,

utilized

● Nitrate, Nitrite, Phosphorus: Important source for plant and animal life

● Ammonia & Total Kjeldahl Nitrogen: Nutrient for plant growth, dietary requirement for all organisms

● Lignin, Cellulose, Hemicellulose: Inorganic substance binding cells, fibers, and vessels (constitutes wood and lignified elements)

● Neutral Detergent Fiber, Acid Detergent Fiber: Animal feed analysis

Green Algae1. Green: Avraınvıllea amadelpha

- No research done regarding the utilization of Avraınvıllea am.

- → Project unveils how this invasive species can be utilized

Hawaii Department of Land and Natural Resources

Red Algae1. Red: Eucheuma spınosum

- Extremely detrimental to color reefs (ex: Kaneohe Bay) → Attaches to substrate and coral

- Causes inflammation of coral reefs → irritation through algal abrasion and over growth

- Creates thick algal mats that carpet coral → stops photosynthesis → coral polyps die

Eyes of the Reef Hawaii

Hypotheses

1. Fermenting the macroalgae will decrease the presence of nitrates and nitrites for ideal fertilizer and animal feed usage

2. Avraınvıllea am. will have a large concentration of lignin, Eucheuma sp. will have a small concentration

3. Fermentation will lead to the breakdown of the cell wall

FermentationThe presence of ammonia will increase due to hydrolysis, occurring at the beginning of fermentation.

Fiber AnalysisAvraınvıllea am. will have a large concentration of lignin due to its tough structure. Eucheuma sp. will have a smaller concentration due to its pliability and weak structure.

MicroscopyFermentation will visibly degradate the cell wall through SEM imaging. The cell walls should have an uneven, ragged texture. Non-fermented algae will show to have a smooth cell wall, non-damaged.

Hypotheses Support

Materials & ProceduresPart One Algal collection and prepping: Algae was collected from Kaneohe Bay

and Hawaii Kai beaches. They were then blended and stored in Ziploc bags.

Part Two Fermentation: Samples were placed in serum bottles along with cow manure and aquaponics water. Nitrogen was inserted in order to displace the oxygen. The bottles were kept in an incubator at 60°C.

Part Three Component analyses: Ammonia, nitrate, nitrite, COD, phosphorus, and fibers.

Part Four Microscopy: A fluorescence microscope and a scanning electron microscope (SEM) were used to observe the species.

Data & Results

Fermented Eucheuma sp. with water

Fermented Eucheuma sp. plain

Non-fermented Eucheuma sp.

Fermented Avrainvillea am. with water

Fermented Avrainvillea am. with water

Fermented Avrainvillea am. plain

Fermented Avrainvillea am. plain

Non-fermented Avrainvillea am.

Statistical AnalysesSet One: Comparison of post-fermentation components between Avrainvillea am. and Eucheuma sp

Set Two: Comparison of difference of components from pre- to post-fermentation between Avrainvillea am. and Eucheuma sp

Set Three: Comparison of Avrainvillea am. and Eucheuma sp. for pre- to post-fermentation TKN between water and no water

Set Four: Comparison between am. and Eucheuma sp. for post-fermentation TKN for water and no water

Set One Set Two

Ammonia: Significant

Ammonia: Significant

Nitrate: Significant

Nitrate: Significant

Nitrite: Insignificant

Nitrite: Insignificant

COD: Insignificant

COD: Significant

Phosphorus: Significant

Set Three Set Four

Av: Water to no water: Significant

Water: Significant

Av: Pre to post fermentation diff: Significant

No water: Significant

Euch: Water to no water: Insignificant

Euch: Pre to post difference: Insignificant

Discussion

FermentationAvraınvıllea am.

● Nitrates decreased significantly● Aquaponics water positively

impacted TKN results● Insignificant increase in nitrites● High concentration of ammonia● Comparatively low concentrations of

phosphorous● Not ideal for plant growth

Conclusion: Avraınvıllea should not be used as a fertilizer

Eucheuma sp.

● Nitrates decreased with minimal significance

● Aquaponics water negatively impacted TKN results

● Insignificant increase in nitrites● Drastic increase in ammonia

○ Can be toxic for plant growth● Phosphorus content ideal for fertilizer

usage

Conclusion: Eucheuma has great potential as a fertilizer

SEM Imaging and Fiber AnalysisAvraınvıllea am.

● SEM Images: Showed breakdown of cell walls after fermentation

● Fiber Analysis: High concentrations of lignin, natural detergent fibers, and acid detergent fibers○ Comparatively high

concentration of cellulose

Conclusion: Avraınvıllea should be utilized for its lignin and cellulose content

Eucheuma sp.

● SEM Images: Showed breakdown of cell walls

● Less breakdown occurred for samples with aquaponics water

● Fiber Analysis: No presence of lignin○ Low concentration of natural

detergent fibers and acid detergent fibers

○ Low concentration of cellulose

Conclusion: Eucheuma has low contents of detergent fibers → Utilized as animal feed

Constituents Scot Pine Spruce Eucalyptus Silver Burch Avrainvillea

Cellulose (%) 40.0 39.5 45.0 41.0 23.18

Hemicellulose

(%)

28.5 30.6 19.2 32.4 4.47

Lignin (%) 27.7 27.5 31.3 22.0 22.83

Comparison of Fibers Between Tree and Algal Cells

Further ApplicationsAvraınvıllea am.

● Extract lignin from cell wall → Creation of carbon nanofibers

● Currently no existing methods for algal lignin extraction

Conclusion: Developed a basis for algal lignin extraction. Avraınvıllea has roughly the same amount of lignin as trees with less hemicellulose and cellulose → Easier to separate constituents → Can also be used for ethanol production, current methods exist

Eucheuma sp.

● Temperature decreased during fermentation → Decreased presence of ammonia and minimized decrease of nitrates

● Fermented Eucheuma sp. found to be a superior fertilizer in comparison to current popular additives

Conclusion: Eucheuma has great potential as a fertilizer → Low concentration of natural and acid detergent fibers → Also ideal as animal feed

Non-fermented Eucheuma sp. Compost

Miracle Grow Only top soilFermented Eucheuma sp.

Preliminary Cost Analysis: Green Algae● Carbon nanotube current cost:

~$50/1 gram● Popular source for creation of

carbon nanotubes: Graphite○ Graphite has to be mined →

Bad for environment → Better to utilize algae■ Remaining algal carcass →

Fertilizer

Meijo Nano Carbon Company

200xStrength of steel

12xLighter than steel

5xElectrical conductivity

15xThermal conductivity

Increasing global market for carbon nanotubes

Preliminary Cost Analysis: Red Algae

● Anaerobic Digester: $240,000 - $290,000

● Gas by-product used to power generator → Cost payback made in 3 ~ 5 years○ According to the U.S.

Environmental Protection Agency’s Biogas Recovery in the Agriculture Sector, anaerobic digester revenue is ~$50,000 yearly

○ Algae had high methane yield → high biogas yield

American Biogas Council

ConclusionLorem ipsum dolor sit amet, consectetur adipiscing elit, sed do eiusmod tempor incididunt ut labore et dolore magna aliqua. Ut enim ad minim veniam, quis nostrud exercitation ullamco laboris nisi ut aliquip.

Acknowledgments● Dr. Samir Khanal, University of Hawaii at Manoa - Mentor● Sreelakshmi Kutty, Saint Andrew’s Priory - Partner● Sumeth Wongkiew, University of Hawaii at Manoa - PhD Student● Ai Hirasuna, Saint Andrew’s Priory - Teacher● Dr. Tina Weatherby, University of Hawaii at Manoa - SEM● University of Hawaii at Manoa - Grant money

Works Cited "1. INTRODUCTION." HANDBOOK ON EUCHEUMA SEAWEED. N.p., n.d. Web. 06 Nov. 2014.

"Avrainvillea Amadelpha." Avrainvillea Amadelpha. N.p., n.d. Web. 06 Nov. 2014.

"Energy: Algae Biofuels - European Commission." Energy: Algae Biofuels - European Commission. N.p., n.d. Web. 06 Nov. 2014.

"Gracilaria Parvispora." Gracilaria Parvispora. N.p., n.d. Web. 06 Nov. 2014.

"PART IVCOUNTRY EXPERIENCE REPORTS." Report On The Training Course On Gracilaria Algae Manila, Philippines 1–30 April 1981. N.p., n.d. Web. 07 Nov. 2014.

"Propagating Gracilaria for Fun and Food!" Reef Builders The Reef and Marine Aquarium Blog Propagating Gracilaria for Fun and Food Comments. N.p., n.d. Web. 06 Nov. 2014.

"Rice Edu Nitrates." Nitrates. N.p., n.d. Web. 07 Nov. 2014.

"The Fluorescence Microscope." The Fluorescence Microscope. N.p., n.d. Web. 07 Nov. 2014.

Works Cited"Ulva Fasciata Delile 1813." Ulva Fasciata Delile 1813. N.p., n.d. Web. 06 Nov. 2014.

The Editors of Encyclopædia Britannica. "Nitrogen Cycle (biochemistry)."Encyclopedia Britannica Online. Encyclopedia Britannica, n.d. Web. 07 Nov. 2014. "Cell Respiration: Fermentation." Cell Respiration: Fermentation. N.p., n.d. Web. 06 Jan. 2015.

"What Is Nitrate and Why Is It Important?" What Is Nitrate and Why Is It Important? N.p., n.d. Web. 08 Jan. 2015.

"What Is Nitrate & Nitrite?" What Is Nitrate & Nitrite? N.p., n.d. Web. 08 Jan. 2015.

"Nitrogen (N) and Water." Nitrogen (N) and Water. N.p., n.d. Web. 09 Jan. 2015.

"About Lignin." About Lignin. N.p., n.d. Web. 09 Jan. 2015.

"Scanning Electron Microscopy (SEM)." Scanning Electron Microscopy (SEM). N.p., n.d. Web. 12 Jan. 2015.

PROS CONSGreen Algae Green Algae

Lignin extraction: Reduces need for trees → Other source of carbon is graphite → needto be mined → Bad for environment

Lignin extraction: Never been done before → Limited research → Difficult to create method → Trial and error

Ethanol: Needed to help us transition to a world without fossil fuels

Ethanol: Hard to get cellulose, not enoughcellulose to create lots of ethanol

Red Algae Red Algae

Fertilizer: Fermented algae better than commercial products → Reduces need for artificial fertilizers

- Can make methane for energy concurrently- Not limited to cow manure, can use sewage

as well → Instead of dumping human sewage into ocean, incorporate as fertilizer

Fertilizer: Costly start up, although pay off consumes initial start up after 3-5 years

Animal Feed: Reduces land and waterfootprint → Land and water can be usedto grow food for people

Animal Feed: Algae has salt content, needs to be washed with fresh water