By Patti Koning
Algae has great promise as a potential source of biofuel — it grows easily and abundantly and can be rich in triacylglycerols, a lipid that can be extracted and converted into fuel. But after 30 years of research, little is known about the mechanism that produces triacylglycerols, the missing link that could take us from algae to “oilgae.”
CATCH AND RELEASE FOR ALGAE — Using laser trapping Raman spectroscopy, Huawen Wu and Seema Singh (both 8634) are able to analyze lipid accumulation in a single algae cell without changing or harming it, something not possible with conventional methods. This technique will greatly accelerate research into candidates for algal biofuel production. (Photo by Randy Wong)
“Certain algae are known to be really good candidates because of their lipid accumulation,” says Blake Simmons (8630). “One of the biggest problems is a lack of fundamental understanding about how these algae do what they do, what environments they do it in, and how we can manipulate them to make them more attractive from a biofuel production standpoint.”
Sandia researchers may have found the key to solving that problem with a new method of analyzing algae at the cellular level. Over the past two years, Seema Singh (8634) has led a Laboratory Directed Research and Development (LDRD) project to develop a real-time in situ method for lipid profiling using laser-trapping Raman spectroscopy (LTRS). While the LDRD project is studying microalgae, the method is universal and can be used for several applications in the field of lipidomics.
“We believe our method has the potential to transform the field of algal biofuel research,” says Seema. “Now we can analyze a single cell’s chemical fingerprint, not just once, but over and over and examine how different factors affect lipid production. The overarching goal is multifactorial correlation of biotic and abiotic factors to algal growth and lipid accumulation.”
The method combines two existing technologies — laser trapping and Raman spectroscopy — that have been widely used for many years. The novelty of Seema’s approach is bringing the two together to study algae.
After months of work to select the best algae candidates, the team began with Raman spectroscopy as a way to develop chemical fingerprints of single-cell algae. “The ‘ah-ha’ moment came when we were saying it would be great if we could just observe these events on a single-cell basis,” Blake says. “Then we realized we could, that the technologies had been demonstrated separately, but that no one had combined these techniques for analyzing lipids in microalgal cells before.”
Seema brought postdoc Huawen Wu (8634) onto the project because of his experience using LTRS to study lipids in human cells. Algae, he says, are comparatively easier because their lipids are more abundant.
The laser immobilizes a single cell in an optical trap; once immobilized, the cell is interrogated with a Raman spectroscopy system. “You can see what lipids are present, the degree of unsaturation, and even the relative quantitation of the absolute lipids present. Then you let the cell go,” Blake explains. “It’s basically a catch-and-release program we developed for algae.”
That catch-and-release aspect is important because other methods can significantly perturb and even destroy the algae in the process of analysis. Two methods are commonly used to study algae today — extraction and fluorescence labeling.
In the extraction method, researchers grow and harvest algae, and then extract the lipids to analyze them. And then start over. The process is slow, involves tricky chemicals and laborious fractionation, and ultimately doesn’t yield much specific information unless you use very expensive mass spectrometry equipment.
“It’s an Edisonian, empirical approach that doesn’t easily get down into the important details of lipid composition in the algae,” says Blake.
The fluorescence labeling method doesn’t fare much better. “It’s a brute force method,” says Blake. “The dye doesn’t like being in water, so it partitions to the lipid within the algae. You can then use that signal as the dye is taken up by the cell to get a rough metric of how much lipid is there. You still don’t know what kind of lipid or how much, and it doesn’t work for all cells. And it’s toxic to the algae.”
To exacerbate the problem, algae themselves are rich in fluorescent pigments and create interference. Sometimes they are just too bright to be seen clearly under a fluorescent microscope.
In contrast, the LTRS method is fast, yielding a complete analysis within a few seconds, and insensitive to culture conditions. A researcher can repeat the analysis over and over within the same sample while adjusting parameters.
“You can grow a culture, test it at one temperature and again at an increased temperature, or even as the culture is getting warmer,” says Seema. “We’re able to track these changes, get a handle on the fundamental biological process of lipid production, and establish a constitutive relationship between growth conditions and lipid yield and profile.”
This method also allows researchers to quickly amass hundreds of data points, which then enable the development of algorithms to conduct a ratiometric analysis of lipid, chain length, and degree of unsaturation.
“At the outset, we wanted to demonstrate the ability to grab a cell and understand its total profile. Then we got greedy and began wondering if we could use this method to predict melting temperature,” says Seema. “Now we can use this calibration plot for predictions. You bring me any algae, or any other cell for that matter, and we can probe it and make a prediction on its melting temperature within plus or minus 1 degree. This allows us to gain insight into a particular algae’s suitability for a biofuels application.”
Because of Sandia’s unique experience with Raman spectroscopy for microalgae, the corporation is now partnering with BaySpec to commercialize a pond-deployable algal analyzer.
The project received a phase one National Science Foundation Small Business Innovation Research grant and Seema is hopeful it will be funded for phase two.
This work was published in the March 2011 issue (vol. 108, 3809–3814) of the Proceedings of the National Academy of Sciences USA. The research also was recently highlighted by the Nature Publishing Group and the LIPID MAPS consortium in the March 23, 2011, article, “Techniques: Lipids probed by Raman spectroscopy in living cells.”
“One beauty of this method is it isn’t limited to algae — you can use it to study virtually anything,” Seema says. “If we can develop something that’s easy to use and get it into the hands of researchers in academia, national labs, and industry, there is so much that can be discovered. The potential, I think, is limitless.” - Patti Koning
By Mike Janes
Fuel cells were used in the space shuttle as one component of the electrical power system, so perhaps it was appropriate that a hydrogen fuel cell-powered mobile lighting system could be seen on the grounds of the Kennedy Space Center as the space shuttle Atlantis launched into orbit last month for the 135th and final mission of the NASA Space Shuttle Program.
The lighting system, sponsored by the DOE’s Office of Energy Efficiency and Renewable Energy (EERE) in conjunction with Boeing Co., and developed by
Sandia and several industry partners, was deployed to the site of the final space shuttle launch and observed by visitors, shuttle astronauts, and members of the international media.
The unit provided lighting in the international press area, and its auxiliary power was used to conveniently recharge the camera battery packs for a number of photographers at the event. The NASA deployment was the latest in a series of high-profile test sites where the lighting system has been utilized.
The hydrogen fuel cell-powered mobile lighting system is a clean, quiet, and efficient alternative to traditional technologies commonly powered by diesel-fueled generators. The system features a fuel cell running on pure hydrogen, resulting in zero-emission electrical power. The fuel cell produces electricity for an advanced, power-saving Light Emitting Plasma™ (LEP) lighting system and additional auxiliary power up to 2.5 kW, which allows additional equipment (such as power tools, public address systems, or security metal detectors) to be powered by the unit at the same time the system is providing illumination.
Current mobile lighting typically uses diesel generators that produce greenhouse gases such as carbon dioxide and nitrogen oxides, which produce pollutants and create smog and soot, making them environmentally objectionable. In addition, diesel units are noisy and can create a safety hazard when construction personnel are distracted and cannot hear oncoming traffic.
Sandia researchers estimate that a single hydrogen fuel cell-powered lighting system would offset 900 gallons of diesel fuel per year and completely eliminate soot and nitrogen-oxide and carbon-dioxide emissions, allowing the system to be used indoors in contrast to current diesel technology.
“This hydrogen fuel cell-powered mobile lighting system has the very real potential to drastically reduce dependence on diesel-fueled mobile lighting across the United States and abroad,” said Lennie Klebanoff (8367), Sandia’s project lead.
The prototype system has been tested in a variety of environments and has primarily focused on the entertainment, transportation, and airport sectors. In addition to NASA (which also used the system during the space shuttle Endeavor launch) customers who have provided test sites include the California Department of Transportation, the 2010 Academy Awards® ceremony, the 2011 Golden Globe Awards, the 2011 Screen Actors Guild Awards, and the 2011 Grammy Awards. Boeing, the San Francisco International Airport, and Paramount Pictures will soon be deploying units as well.
In addition to the DOE’s sponsorship and Sandia’s design and technical management role, the industry partners on the project include Boeing, Multiquip Inc., Altergy Systems, Luxim Corp., Lumenworks Inc., Stray Light Optical Technologies, Golden State Energy, and Ovonic Hydrogen Solutions. The California Fuel Cell Partnership has provided support on hydrogen fuel for several deployments. Multiquip is implementing a manufacturing and commercialization plan for the system. -- Mike Janes
By Mike Janes
When funding became available for the NNSA labs to develop climate-change solutions, a number of researchers at Sandia put their collective heads together and agreed that a mobile facility — one that could be transported to various locations on an as-needed basis — would be a timely and useful contribution.
So that’s what they built.
“What we proposed to do was build a mobile facility that would measure greenhouse gases and other species associated with those gases so they could be traced and identified,” says Hope Michelsen (8353), a combustion and atmospheric chemist and one of the lead researchers on the project.
In addition to pinpointing the chemicals’ location information, the idea behind the mobile facility is to learn whether the gases are biogenic (coming from plant sources) or anthropogenic (coming from man-made sources). This is important when officials look at ways to mitigate emission impacts in their communities, regions, states, or even nations.
“Elected officials who have enacted new policies to help reduce unwanted greenhouse gas emissions could conceivably find a mobile facility to be of great use,” Hope says. “To figure out whether emissions reduction policies are effective, we need a way to measure emissions by emissions sector, such as power generation or transportation. We currently don’t have the tools in place to do these types of measurements, so we hope our idea can be part of the solution.”
The mobile system, which has already been deployed once to the Atmospheric Radiation Measurement (ARM) facility in Oklahoma, consists of two moving van-sized trucks, each equipped with instrumentation and equipment. Included are instruments that measure greenhouse gases, such as carbon dioxide (CO2) and methane (CH4), and species co-emitted with greenhouse gases, such as sulfur dioxide (SO2), nitrogen oxide (NOx), carbon monoxide (CO), ozone (O3), and other traditional pollutants.
While the instruments are all commercially available, another current project funded internally at Sandia aims to build an instrument that measures black carbon soot, a devastating warming agent formed through the incomplete combustion of fossil fuels, biofuels, and biomass.
Distinguishing emission sources
Atop each truck sits an antennae-like mast that sucks in air, sends it down into the truck, and distributes it to the various instruments, including a mass spectrometer that sorts out hydrocarbons and helps to distinguish between emission sources, which can range from traffic to pine trees.
A piece of equipment provided by Lawrence Livermore National Laboratory (LLNL) captures samples in flasks for analysis at LLNL’s Center for Accelerator Mass Spectrometry to measure the radiocarbon (14C) fractionation of CO2 (14C is a radioactive isotope of carbon).
Because 14C is severely depleted in fossil fuels, it is a powerful indicator of man-made CO2 sources. Some of these samples were also sent to the National Oceanic and Atmospheric Administration and to the University of California, Davis for further trace gas and isotopic analysis.
The ARM location in Oklahoma, says researcher Ray Bambha (8128), was selected because of its solid history as a climate research site. Ray served as the principal investigator for the field experiment, alongside several other Sandians serving in key roles. Collaborators from both LLNL and Los Alamos National Laboratory joined the Sandia team for portions of the Oklahoma deployment.
‘Uncertainty quantification’ capability
The pilot deployment, Ray says, was successful in that the system collected large quantities of data, which is still being analyzed. It allowed the team to test instruments that hadn’t been used previously, and it helped them to understand the atmospheric community’s need for an “uncertainty quantification” capability — a method of assigning a confidence level to an estimate — and tracer measurements, which provide a more effective method for identifying the source of certain emissions.
Hope says Sandia is building a team of researchers that can take the next step with the system and begin to use wind information and inverse modeling to more accurately identify emission sources.
In the short-term, Hope says, program development efforts are well under way in hopes of securing follow-on funding and other test deployments. The long-term vision for the program calls for a full network of mobile facilities that could be deployed strategically in select regions, states, or cities to enable the capturing of a broad spectrum of emissions and related information. -- Mike Janes