Algae Biofuels

Research Overview

The algae biotechnology efforts of the Curtis Lab started in earnest in ~2005 to assist the first major algal biofuels company, GreenFuel Technologies, including having several of their earliest employees undertake training at Penn State as they setup laboratories in Boston. This work also included producing 100 liters of innoculum for the MIT cogeneration powerplant demonstration. In 2007-08, Dr. Curtis went on sabbatical, devoting half of it as a technical advisor to Green Fuels Technologies startup effort in Boston.

Since then, CurtisLab has similarly continued applying chemical engineering principles to develop methods of producing algae biofuels emphasizing economic feasibility. This includes:

  • Demonstration of ultra high-density ( > 20g/L), continuous growth vis-a-vis Wayne's Freshwater Algae Media (WFAM) and a low-cost Trickle Screen Reactor (patent pending), which resulted in the Chappell Lab & CurtisLab's NSF-CBET highlight

  • Subsequent NSF-SBIR (Small Business Innovation Research) grant for Dr. Curtis' short-lived Calyx Bioprocess Technology LLC for scale-up

  • Comparison of algal biofuels candidates Botryococcus braunii and Chlorella vulgaris via growth kinetics and bomb calorimetry to substantiate B. braunii's superiority as a biofuels candidate

  • Feed-forward pH control strategies to reduce operational costs and optimize growth

  • Genetic engineering of B. braunii's botryococcene-synthesizing genes into alternative hosts

  • Novel method for maintaining laboratory-scale axenic algal cultures Isolation and genetic characterization of a novel ectosymbiont

  • Curation of an open-source wiki page, botryococcus.org, to centralize information relevant to B. braunii

  • Tandem production of high-value protein and biofuel in microalgae

Recent Publications

  1. Wang, J., Rosov, T., Wensel, P., McGowen, J., Curtis, W.R. (2016) A preliminary implementation of metabolic-based pH control to reduce CO2 usage in outdoor flat-panel photobioreactor cultivation of Nannochloropsis oceanica microalgae. Algal Research, 18: 288-295. DOI:10.1016/j.algal.2016.07.001

  2. Wang, Jun, Curtis, Wayne R. (2015) Proton stochiometric imbalance during algae photosynthetic growth on various nitrogen sources: Toward metabolic pH control. Journal of Applied Phycology. DOI: 10.1007/s10811-015-0551-3

  3. Khatri W, Hendrix R, Niehaus T, Chappell J, Curtis WR (2013) Hydrocarbon Production in High Density Botryococcus braunii Race B Continuous Culture, Biotechnology & Bioengineering, 111 (3), 493-503 doi:10.1002/bit.25126

  4. Myers JA, Curtis BS, Curtis WR (2013) Improving accuracy of cell and chromophore concentration measurements using optical density. BMC Biophysics 2013, 6:4, 1-16.

  5. Scherholz ML, Curtis WR. (2013) Achieving pH control in microalgal cultures through fed-batch addition of stoichiometrically-balanced growth media. BMC Biotechnol. 13(1):39, 1-16.

  6. Curtis WR, Curtis MS (2014) Biomass-2-Energy, Chapter 3, In: Systems Engineering for Clean and Renewable Energy Manufacturing in Europe and Asia, NSF sponsored Report, WTEC. (Full Report on Web; http://wtec.org/SEEM/)

Other papers, presentations, and patents

  1. Curtis WR, Wang J, Khatri W, Tuerk A. A Decade of algae bioprocess engineering: the neglected importance of operational strategy and control. American Chemical Society (ACS) National meeting, Aug 16-20, 2015. Boston, MA.

  2. Curtis WR, Wang J. Metabolism based feed forward pH control of high-density microalgae cultures. DOE-BIOMASS 2014, July 29-30, Washington, DC.

  3. Wang J, Geveke B, Johnson R, Curtis WR. Feed-Forward Control of Dynamic High-Density Microalgae Cultures Using Model Based Predictive pH Control and a Novel Biomass Sensor. AIChE Annual meeting, November 3-8, 2013. San Francisco, CA.

  4. Link D, Tuerk A. Overall Energy Considerations for Algae Species Comparison and Selection in Algae-to-Fuels Processes. Alternative Energy NOW Conference, Lake Buena Vista, FL, February 23, 2011.

  5. Johnson, Ryan, Khatri, W., Curtis, W. (Calyx Bioprocess Technology, LLC., Port Royal, PA) SBIR Phase I: Hydro-ThermAlgae, Photobioreactor for Minimal Water Loss. Final Proposal [not accepted] Arlington, VA: National Science Foundation; 2012 Sep. IIP-Small Business Phase I.

  6. Amalie Tuerk*, Wayne R. Curtis. Algae-Based Hydrocarbon Production. USDA Research Center, Philadelphia, PA, April 2010.

  7. Curtis WR. Trickle-film Bioreactor for Growth of Photo-hetrotrophic anaerobes for Membrane protein production and growth of photosynthetic algae. Patent US 14/729,692. Applied: 2009.

  8. Algae-Based Hydrocarbon Production Waqas Khatri, Steve Gabauer, Joseph Chappell, Wayne Curtis. AIChE Annual Meeting, Philadelphia, PA, Nov. 18 2008

  9. Genetic Engineering Hydrocarbon Production in Algae Stephen Gabauer, Thomas D. Niehaus, Waqas Khatri, Joseph Chappell, Wayne Curtis. AIChE Annual Meeting, Philadelphia, PA, Nov 19 2008

Theses

  1. Curtis WR, Wang J, Khatri W, Tuerk A. A Decade of algae bioprocess engineering: the neglected importance of operational strategy and control. American Chemical Society (ACS) National meeting, Aug 16-20, 2015. Boston, MA.

  2. Curtis WR, Wang J. Metabolism based feed forward pH control of high-density microalgae cultures. DOE-BIOMASS 2014, July 29-30, Washington, DC.

  3. Wang J, Geveke B, Johnson R, Curtis WR. Feed-Forward Control of Dynamic High-Density Microalgae Cultures Using Model Based Predictive pH Control and a Novel Biomass Sensor. AIChE Annual meeting, November 3-8, 2013. San Francisco, CA.

  4. Link D, Tuerk A. Overall Energy Considerations for Algae Species Comparison and Selection in Algae-to-Fuels Processes. Alternative Energy NOW Conference, Lake Buena Vista, FL, February 23, 2011.

  5. Johnson, Ryan, Khatri, W., Curtis, W. (Calyx Bioprocess Technology, LLC., Port Royal, PA) SBIR Phase I: Hydro-ThermAlgae, Photobioreactor for Minimal Water Loss. Final Proposal [not accepted] Arlington, VA: National Science Foundation; 2012 Sep. IIP-Small Business Phase I.

  6. Amalie Tuerk*, Wayne R. Curtis. Algae-Based Hydrocarbon Production. USDA Research Center, Philadelphia, PA, April 2010.

  7. Curtis WR. Trickle-film Bioreactor for Growth of Photo-hetrotrophic anaerobes for Membrane protein production and growth of photosynthetic algae. Patent US 14/729,692. Applied: 2009.

  8. Algae-Based Hydrocarbon Production Waqas Khatri, Steve Gabauer, Joseph Chappell, Wayne Curtis. AIChE Annual Meeting, Philadelphia, PA, Nov. 18 2008

  9. Genetic Engineering Hydrocarbon Production in Algae Stephen Gabauer, Thomas D. Niehaus, Waqas Khatri, Joseph Chappell, Wayne Curtis. AIChE Annual Meeting, Philadelphia, PA, Nov 19 2008

Photobioreactor (PBR) Design and Operational Strategies

Trickle Film Reactor for Pilot-Scale Production

Many processes and photobioreactor designs in the algae community have obvious or hidden limitations that have significant impact when evaluated from a biochemical engineering stance (i.e. simple mass and energy balance principles). Alternatively, CurtisLab's Trickle Film Reactor (TFR) considers the entire algae biomass production process to address the inherent limitations of light physics, algae physiology, and economics, thereby achieving high-density growth at minimal costs, both fiscal and energetic, and limited only by light. Specifically, the TFR achieves maximum productivity via a turbulent-mixed film at minimum thickness in conjunction with light-limited, high-density algae growth. Furthermore, it facilitates low-cost downstream dewatering and reduced ‘hydraulic load’ costs throughout the process.

Schematic of the trickle-film photobioreactor, illustrating the overall flow pattern for liquid and gas (A), a close up of the trickling film depicting the short path length for light penetration as well as mixing induced in the falling film due to drag (B), and an idealized view of how the diagonal screen increases resistance to the fluid flow path and induces vorticity to the flowing algae culture (C).

Algae is grown as a suspension that trickles down a screen to induce turbulent mixing in a simple geometry that promotes efficient photon use by avoiding a surface that can foul between the algae and the light, thereby supporting densities of 30+g/L biomass—correlating the highest oil productivities found in literature irrespective of the intrinsic growth rate among various strains of algae. We have demonstrated that CO2 fertilization can be accomplished in the evaporative water make-up saturated with nearly pure CO2. This avoids prohibitive costs of heat removal and gas compression on agricultural scales. This approach also avoids the capital expense and safety/permitting issues of light blocking ‘green-house’ enclosures and use of existing scrubbing technologies to provide concentrated CO2.

Dynamic Feeding & Feed-Forward pH control

Critical to algae biofuels are operational costs associated with controlling pH, which inherently affects algal health and growth. Algae pH is affected by both carbon and nitrogen utilization, and significantly by the source of nitrogen. For example, preferntial consumption of ammonium (NH4) results in a dissociation into ammonia (NH3) and proton (H+) which lowers pH while consumption of nitrate dissociates to form nitrite (NO2) and a hydroxyl group (OH-), in turn raising pH. Unsophisticated methods of buffering that are heavily relied upon in the world of algae biofuels include dissolving CO2 to buffer these pH changes, however, they are intensively wasteful by ignoring Henry's Law for the dissolution of gasses to waste ~90% of CO2 at $50/ton and thereby antithetically contributing to GHG emissions. In revealing these significant obstacles, CurtisLab developed an OD sensor (patent pending).

Laboratory scale pH monitoring and dynamic feeding implementation

To improve this problem, CO2 was appropriately minimized to a volume appropriate for photosynthetic growth. Meanwhile, a dynamic feeding strategy was used to counteract algae's preferential consumption of ammonium---to the point of gluttonously killing itself. Finally, feed-forward process control, whereby proactive predictive modeling rather than a lagging reactive response informs the control strategy, was implemented to regulate pH. Specifically, growth predictions based on light cycles dictated the respective dosing of acids and bases to maintain pH.

This was initially demonstrated at the laboratory scale, and to date has been scaled-up for the single cell, saltwater algae, Nanochloropsis, at Arizona State's (ASU) AzCATI, the Arizona Center for Algae Technology and Innovation. Now graduated, the Ph.D. who pursued this work, Jun Wang, continues to collaborate with CurtisLab and AzCATI to improve feed-forward modeling and process control in addition to scaling and automating this process for other alga including but not limited to Hamaetococcus, Chlorella vulgaris, Galdieria sulphuria, Botryococcus braunii, Poryphridium purpureum, and Scenedesmus dimorphous, and Chlamydomonas reinhardii.

'Alginator' for Bench-Scale Microalgae Characterization Based on Photoefficiency Metrics

In response to the Trickle Film (TF) reactor's success in demonstrating performance of microalgae biofuels candidates under commercially relevant conditions at pilot-scale, in turn contradicting the utility of intrinsic growth rate as a valuable performance metric, the 'Alginator' bioreactor was designed as a bench-scale reactor to evaluate microalgae biofuels' performance using photoefficiency. The emphasis on efficiency of solar-to-biomass conversion is central to the Alginator's design and relevance to commercially-revelant conditions of continuous operation and high-density, light-limited growth.

Left to Right: Visiting Scholar Ramya Mohandass (SRM University, India), Ben Geveke, Lucas Nugent, and (Michael) Grant Gill in summer 2016 with Alginator. (Note: this is one of the last laboratory photos taken in 232 Fenske Laboratory.)

In principle, the Alginator is a flat-panel, external loop airlift photobioreactor that is designed to be low-cost and bench-scale to make the technology to characterize algae or, more specifically to 'bioprospect' for a biofuels production candidate, accessible for standard laboratories. The critical design elements that enable measurement of photoefficiency is the Alginator's flat-panel design, narrow path length, and defined light path & spectra. Together, these elements define the energy inputs and outputs for generating an energy balance and calculating photoefficiency. The airlift design with CO2 supplementation also promotes sufficient mixing and carbon mass transfer to sustain high-density photoautotrophic growth as shown by the dense green culture below.

Genetic Engineering for Fuels Production

Our collaborators at Chappell Lab elucidated the two biosynthetic pathways in Botryococcus braunii, the Mevalonic Acid (MVA) and 2-C-methyl-D-erythritol 4-phosphate (MEP) pathways. As part of CurtisLab's work in metabolic engineering, we engineered the botryoccene-synthesizing genes into a bacterial host as part of an ARPA-e Electrofuels grant.

Tandem Protein Production

In 2014, the Department of Energy's Bioenergy Technologies Office (DOE BETO) announced its aim to reduce algal biofuels price from its ~$8 gge (gas gallon equivalent) to $3 gge by 2030 with an intermediate goal of achieving $5 gge by 2019. To that end, algal biofuels are being pushed as a dual production platform in which the cost of producing large amounts of algal biofuels is offset by a co-produced high-value product, and significantly, one which maintains its production value at the associated production levels. Parlaying our work in using plants as a protein production platform, our ongoing work similarly aims to produce a high-value protein with the potential to be as a bioplastic derivative. CurtisLab is currently investigating the optimal platform and protein expression system.

To that end, we have previously collaborated with the Demirel Lab (PSU, Materials Sciences) to scale-up production of a potentially high-value protein for their material characterization. Further we have ongoing Penn State collaborations with the Bryant Lab (Biochemistry & Molecular Biology) as well as Phil Savage's Lab (Chemical Engineering). Future work may also include transplastomic engineering.

Establishing & Maintaining Axenic Botryococcus braunii

Akin to our TSReactor work, we also grew Botryococcus braunii semi-continuously at high-density at laboratory scale. A surprising struggle we found was in isolating Botryococcus braunii to establish axenic (i.e. B. braunii and ONLY B. braunii) cultures---in fact, taking almost a year-and-a-half to isolate it! This led to (1) a method of maintaining B. braunii to reduce contamination and (2) genomic study of the novel contaminating ectosymbiont.

Novel Axenic Maintenance Method

The video below details the SOP for maintaining a co-culture of axenic phototrophically-grown B. braunii and heterotrophically-grown plant roots for optimal gas exchange, respectively via photosynthesis and aerobic respiration. Ongoing work aims to quantify growth of numerous algae and plant root species, in turn, enabling optimized pairings of algae and plants.

Meet the Algae Biofuel Team

Curtis Lab Researchers

  • Lisa Grady (graduate)

  • Waqas Khatri (graduate)

  • Amalie Tuerk (graduate)

  • Jun Wang (graduate)

  • Ryan Benoit (UG)

  • Chris Colona (UG)

  • Erik Curtis (UG)

  • Ben Geveke (UG)

  • Michael Grant Gill (UG)

  • Nate Hamaker (UG)

  • Robert Hendrix (UG)

  • Patrick Hillery (UG)

  • Katie Legenski (UG)

  • Erica Lennox (UG)

  • Taylor Maher (UG)

  • David Martino (UG)

  • Mainara Muhl (UG)

  • Bill Muzika (UG)

  • Adam Nebzydoski (UG)

  • Marcia Rodrigues (UG)

  • Megerle Scherholz (UG)

  • Eva Mei Shouse (UG)

  • Steve Tran (UG)

  • Amalie Tuerk (UG)

  • Haonan Xu (UG)

  • Justin Yoo (UG)

  • Joy Yuan (UG)

  • Ryan Johnson (technician)

Chappell Lab Researchers

  • Tom Niehaus (graduate)

  • Steve Gaubauer (graduate)

  • Erik Nybo (graduate)

  • Scott Kinison (technician)

DOE Arizona Testbed Public-Private Partnership

  • John McGowen (director)

  • Pierre Wensel (technician)

  • Theresa Rosov (technician)

Collaborators

  • Joe Chappell

  • Phillip Savage

  • Randy van der Waal

  • Don Bryant

  • Istvan Albert