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Highland Heights, OH, United States

Tominaga H.,Mie University | Coury D.A.,Mie University | Coury D.A.,Phycal, Llc | Amano H.,Mie University | Kakinuma M.,Mie University

Synthesis and accumulation of molecular chaperones are universal responses found in all cellular organisms when exposed to a variety of unfavorable conditions. Heat shock protein 70 (Hsp70), which is one of the major classes of molecular chaperones, plays a particularly important role in cellular stress responses, and the Hsp70 system is the most intensely studied in higher plants and algae. Therefore, we isolated and characterized a cDNA clone encoding Hsp70 from a sterile strain of Ulva pertusa (Ulvales, Chlorophyta). The sterile U. pertusa Hsp70 (UpHsp70) cDNA consisted of 2,272 nucleotides and had an open reading frame encoding a polypeptide of 663 amino acid (AA) residues with a molecular mass of 71.7 kDa. Amino acid alignment and phylogenetic analysis of Hsp70s from other organisms showed that UpHsp70 was more similar to cytoplasmic Hsp70s from green algae and higher plants (≥75%) than to those from other algae and microorganisms. Southern blot analysis indicated that the sterile U. pertusa genome had at least four cytoplasmic Hsp70-encoding genes. UpHsp70 mRNA levels were significantly affected by diurnal changes, rapidly increased by high-temperature stress, and gradually increased by exposure to copper, cadmium, and lead. These results suggest that UpHsp70 plays particularly important roles in adaptation to high-temperature conditions and diurnal changes, and is potentially involved in tolerance to heavy metal toxicity. © 2010 Springer Science+Business Media, LLC. Source

Agency: NSF | Branch: Standard Grant | Program: | Phase: | Award Amount: 50.00K | Year: 2010

This Small Business Innovation Research Phase I project develops a system for the biosecure production of algal oils in open raceways while utilizing genetically modified algae. Through production of transgenic algae containing specific gene regulatory sequences, this research develops algal strains that conditionally express an essential chloroplast-encoded gene only in the presence of the specific inducer compound. Thus, transgenic algal strains would survive only in the presence of the inducer compound (which will be added to the ponds in our production facility). On escape from the production facility, these algae will stop production of the essential target protein and be unable to reproduce in the environment. This biosecure production system will allow the use of genetically modified algae in open pond production systems without fear of impacting the surrounding systems.

The broader impact of the proposed research will be to enable the use of low cost, open-pond culturing systems for the production of biofuels using microalgae genetically manipulated for maximum oil and biomass production. This will be accomplished in a biosecure manner through the development of molecular biological strategies that prevent the genetically manipulated algal strains from reproducing once they are removed from the production ponds. The use of molecular genetics to both increase the production of oil and ensure biosecure use of the transgenic algae (unable to replicate in the surrounding environment) could enable cost effective production of biofuels using algal systems.

Agency: NSF | Branch: Standard Grant | Program: | Phase: | Award Amount: 180.00K | Year: 2010

This SBIR Phase I project will develop a system for magnetic separation from growth media and cell disruption of algae to improve the efficiency of harvesting algae for bio-fuels production. The use of magnetic technologies has the potential to improve the energy efficiency of removing algae from growth media. The use of a magnetic separation technology introduces an energy efficient means by which algae can be dewatered and converted into bio-fuels in an integrated bio-refinery. Using algal strains modified to overproduce ferritin (a magnetically susceptible bio-particle) enables the removal of algae from contaminating biomass as well.

The broader/commercial impact of the project will be magnetic harvesting and dewatering of algal product which could lead to higher efficiencies of oil production. This will enable the use of low cost open pond culturing systems for the production of biofuels using microalgae. The development of technologies using magnetically responsive algae (either
direct, or through association with magnetically responsive particles) to either separate them from growth media and contaminants or to induce cell lysis and aide in oil extraction. The project targets the two highest cost factors in algal oil production, namely dewatering and extraction. As such, advances in these two areas can drive algal based biofuels technology to be
cost competitive with petroleum sources.

Agency: NSF | Branch: Standard Grant | Program: | Phase: | Award Amount: 495.76K | Year: 2012

This Small Business Innovation Research Phase II project develops novel technologies for separation and concentration of intrinsically magnetically susceptible algae for production of biofuels and biochemicals. Phase II builds on the feasibility demonstrated in Phase I using a model alga. During Phase II, an algal strain used for production of renewable biofuel feedstock will be utilized. Novel transformation vectors and tools developed for a production strain, Auxenochlorella protothecoides, will be used to make the algae magnetically susceptible. These traits provided an advantage vs. wild-type strains in growth in low iron medium for the model alga. Phase II will test modified algal strains at lab- and subpilot-scale to determine their performance in growth, and competition with wild-type and weedy algal strains. Additionally, strains will be tested for their ability to be separated or harvested magnetically. This separation will be modeled to determine cost efficacy for primary or secondary dewatering. The specificity of this separation will also be evaluated in relation to downstream use in a heterotrophic bioreactor. The OSU collaboration allows use of these strains in novel rare earth magnetic separators. The endpoint will be novel technologies to improve the overall cost structure for the production of algae-derived biofuels and biochemicals.

The broader impact/commercial potential of this Phase 2 research project will be to provide improvements in the economics of producing renewable biofuels using algae as the production system. It directly addresses one of the major issues with algal biofuels, cost effective dewatering. It also provides a potential selective advantage of the modified strains by improving its ability to compete for iron in an open environment (such as open raceways or photobioreactors). The nation has a critical need to improve its energy security and reduce its dependence on fossil fuels. This research will help address both of these needs. The overall purpose of this research project is business related and focused on commercialization of this technology through integration in a biofuel production process. This research project focuses on a high cost portion of the production process, dewatering, as well as a critical unit process, the heterotrophic bioreactor. The collaboration with OSU and the Cleveland Clinic will result in training of students in this area. The company plans to publish the results of this project once proper control of the intellectual property generated is accomplished.

Agency: Department of Energy | Branch: | Program: SBIR | Phase: Phase II | Award Amount: 1.00M | Year: 2011

Extraction is a major cost driver in the production of algal oil. Reducing extraction cost is critical for the economic viability of algal biofuels. This project uses solid phase, hydrophobized silica resin to improve extraction of algal lipids. Process streams containing lipid and other hydrophobic compounds are treated to recover lipids and Corp) for extraction of the algal lipid produced after the Heteroboost process. Hydrophobized resins were evaluated with model triacylglycerides (TAGs) and fatty acids (FFA) to efficacy and selectivity of adsorption of TAGs and FFAs. Resins were then tested in complex mixtures of algal lipids, with or without algal cell debris or biomass. The inability to separate silica from cell debris cleanly ruled out use in these complex mixtures. However, the resins effectively removed algal TAGs and FFAs from clarified aqueous process liquids generated in Phycals aqueous extraction process. Additionally, this efficacy was maintained for 10 adsorption/desorption cycles. Phase II will focus on recovery of hydrophobic compounds from two process streams generated by Phycals aqueous extraction process. The ability to capture algal oil that is currently lost directly improves production cost. Recovering polar lipids that are not currently captured in aqueous extraction will provide additional cost savings. The project will focus on validation of the adsorption/desorption process at the subpilot, documentation of any higher value hydrophobic compounds recovered from this process, scale up of the process to the pilot plant, and develop a cost analysis/model to allow transition to the commercial plants to be constructed in Hawaii. The deliverables will be an optimized reactor design for adsorption/desorption on hydrophobized resins, optimized silica resin, optimized process at subpilot-scale, and field testing of the optimized reactor/methods at pilot-scale. Commercial Applications & amp; Other Benefits: Phycal is constructing a pilot-scale algal oil production facility on the Island of Oahu, Hawaii. This facility is designed to incorporate our existing production methods, but also to be available for testing improvements to these methods that will lower the cost of the algal biofuels that will be produced. This project could have a significant and positive impact on the cost of production of the algal oil by reducing the loss of lipid in process wastestreams and capturing of other hydrophobic compounds that have value either as an additional fuel feedstock or higher value product.

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