Berkeley Lab researchers led by Hoi-Ying Holman have invented a technology that overcomes challenges in determining the chemical composition of complex biological systems, such as tissues, biofilms and bacterial colonies. Spatially resolved Ambient Infrared Laser Ablation Mass Spectrometry (AIRLAB-MS) uses an IR microscope with an infinity-corrected reflective objective and a continuous-flow solvent probe coupled to a mass spectrometer. Laser ablation of water that is naturally present in biological samples ejects sample materials into a plume of fine droplets, which can be ionized by intersection with an electrospray plume or captured in solvent for ionization by electrospray. AIRLAB-MS enables transfer of material from sample to electrospray ionization emitter at a significantly higher efficiency (~50%) than values reported for similar techniques. Sample preparation is not required. The goal of measuring and imaging the chemical composition of biological samples under native conditions, and with minimal modification/preparation, is crucial to understanding processes such as cell differentiation, photosynthesis, and metabolism. To analyze biological samples with low concentrations of some molecular species, high transfer efficiency is important. However, existing techniques report either low transfer efficiency and significant sample losses or high spatial resolution but ambiguous chemical information. AIRLAB-MS overcomes these problems, reporting a significant transfer efficiency of ~50%.

Researchers at JBEI have metabolically engineered both bacteria (E. coli) and yeast (S. cerevisiae) to produce a chemical precursor, bisabolene, that readily converts to bisabolane when saturated with hydrogen gas under pressure. With continuous yield improvements, biosynthetic bisabolane could become a renewable diesel fuel alternative offering comparable energy density and superior cold weather performance to standard D2 diesel fuel. Although bisabolane had not previously been considered as a biofuel, testing revealed that its performance rating, or Derived Cetane Number (DCN), was 41.9, which is within the 40-55 range of standard diesel fuel. In addition, the analysis showed its “cloud point,” an important marker for cold weather performance, was -78°C, better than diesel’s -35°C, and vastly superior to commercial biodiesel’s -3°C. In sufficient quantities, bisabolane could initially serve as a cold weather additive to diesel and biodiesel formulations and, with higher yields, could substitute for these fuels. (In order to acquire enough material for cetane testing, the JBEI researchers isolated bisabolane from commercially available mixtures of the precursor chemical, bisabolene.) The JBEI team also developed a screening process to identify plant genes involved in the natural production of the precursor chemical, bisabolene, which is produced in small quantities by spruce and fir trees. Using synthetic biology techniques, JBEI scientists inserted a gene from the Grand Fir tree into E. coli bacteria, as well as in a yeast strain widely used for fermentation of ethanol. With further genetic adjustments designed to optimize the production of bisabolene, the yield of that chemical from E. coli has been increased tenfold. Berkeley Lab scientists have thus identified an organic chemical with enormous potential as a diesel oil substitute and developed a robust synthetic biology platform to produce it — and future engineered biofuels — microbially.

Researchers at the Joint BioEnergy Institute (JBEI) led by Blake Simmons and Seema Singh have developed a cost effective pretreatment technology for lignocellulosic processing utilizing ammonium based ionic liquids (ILs) that perform efficiently in large amounts of water under low temperatures. This development is essential in the effort to use sustainable processes for the production of chemicals, materials, and fuels from alternative renewable sources such as biomass. The concept of inducing labile biomass deconstruction with reduced energy and minimal IL loading inputs for enzymatic saccharification is found in the Lab approach. Pretreatment of tetrabutyl-ammonium hydroxide (TBAH) containing 60% water is shown to dissolve 10 wt% of switchgrass under mild conditions at 50°C, with a 93±1.3% glucose yield after 3 hours. This indicates that the aqueous TBAH pretreatment significantly improves the accessibility of the cellulose present in the cell-wall matrix for creating fermentable glucose through enzymatic hydrolysis. The present day pretreatments of biomass include dilute acid, ammonia, and ILs, which are achieved with relatively severe conditions (~120-200°C) followed by a drastic variation of heating from 40-80°C for the enzymatic saccharification to yield around 60-90% sugar, depending on the selection of pretreatment methods. However, the ammonium based ILs pretreat biomass as efficiently as [C2C1Im][OAc], the current highest performing IL, as well as having the advantage of allowing pretreatment to be performed at low temperature (cutting 80% steam requirement) and in less than half the IL quantity (in presence of 60% water), outperforming current pretreatments methods with regards to glucose yield. In addition, these ammonium based ILs are affordable and biocompatible.

Geometry of a single-well survey. Fine grid spacing is used around the low velocity borehole.

Berkeley Lab’s AnisWave2D software is computationally-efficient, fully anisotropic finite-difference code for modeling seismic wave propagation in complex media like that surrounding hydrocarbon reservoirs. Most hydrocarbon reservoirs are overlain by low seismic velocity and anisotropic sediments whose effects on the wavefield must be considered in order to successfully image deeper structures. Conventional uniform-grid finite-difference (FD) schemes running on a single processor cannot tackle large-scale anisotropic models with realistic velocity structure. AnisWave’s use of variable grid spacing improves the efficiency of the FD method by avoiding the oversampling of high velocity zones that occurs in uniform grid methods. This enables more efficient modeling of materials with strong velocity contrasts, such as boreholes, fractures and soft marine sediments. The use of variable-gridding combined with parallelization and optimization of the code greatly increases the complexity of the problems that can be tackled, without sacrificing accuracy.

Researchers at Berkeley Lab led by Gao Liu have developed an ant-nest electrode structure in lithium sulfur (Li-S) batteries that provides large area storage capability and interconnected pathways to channel ion conductivity within the electrodes. The Berkeley Lab technology yields high sulfur loading, strong cycling stability, improved conductivity, and very efficient ion transport in a cost efficient, streamlined and elegant design for Li-S batteries. A porous structure was produced using a polymer binder, conductive carbon nanotube framework and salt micro-particles as a sacrificial additive. Resulting electrodes attained a maximum specific capacity of 1,123.5 mAh/g at c/10 and 908.5 mAh/g at c/3 rates for up to 100 cycles. Additionally, the design allows for a high sulfur ratio of 80% to 85% and high sulfur loading of 3 mg/cm2. Past Li-S batteries have fallen short for commercial use due to low energy density and poor cycling stability due to the dissolution of the polysulfide in the electrolyte during charging and discharging. The Berkeley Lab ant-nest electrode structure overcomes this limitation to meet future commercial demands with high electrochemical performance and high sulfur utilization.

David Chen, Ping-Chi Benjamin Chen, and Doug Chan have demonstrated that autophosphorylation of DNA-PKcs is an important event in the repair of DNA double strand breaks (DSBs) by nonhomologous end-joining. The Berkeley Lab researchers have identified two autophosphorylation sites on the extremely large DNA-dependent protein kinase catalytic subunit (DNA-PKcs) and have generated antibodies that will enable the in vivo monitoring of DNA-PK response to DNA damage. The newly discovered autophosphorylation sites are located at Threonine 2609 and Serine 2056. A mammalian cell line that expresses a mutation at Threonine 2609, preventing DNA-PKcs autophosphorylation at the site, showed an increase in cell radiosensitivity. Ionizing radiation and cancer drugs inflict DSBs in order to kill cancer cells. The ability to intervene in autophosphorylation of T2609 or S2056 and hinder DBS repair, either through application of a drug or an antibody, would increase the radiation-induced killing of cancer cells. Therefore, identification of these critical autophosphorylation sites is a first and necessary step in advancing this line of cancer treatment. “A cell lacking this kinase will become extremely radiation sensitive,” says David Chen. A drug that renders the two identified sites inert would “reduce a patient’s radiation dose and reduce their side effects.” Dr. Chen and colleagues have generated antibodies that recognize Threonine 2609 and Serine 2056 but do not bind to the unphosphorylated DNA-PKcs protein or peptide. These antibodies identify areas where DSBs are being repaired and therefore can be used to monitor the effectiveness of treatments that target the DNA repair pathway of cancer cells. Before the Berkeley Lab phosphorylation-specific antibodies were identified, DNA-PK activity could not be monitored in vivo. Because there is an abundance of DNA-PK in a cell’s nucleus, it has been impossible to distinguish between the large DNA-PK background signal and DNA repair protein foci using immunofluorescence and currently available antibodies.