Production of Biofuel-Related Isoprenoids Derived from Botryococcus braunii Algae
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The colony algae Botryococcus braunii produces large amounts of C30+ triterpene hydrocarbons. Recent discovery of the associated biosynthetic genes has facilitated the metabolic engineering of these triterpene hydrocarbons in alternative hosts – where squalene has served as an analytical standard and a closely associated model hydrocarbon biosynthetic pathway. An extraction and analysis method is provided for both the native and heterologous systems. In the case of the native algae, the hydrocarbons are tightly associated with a complex wall matrix. In addition to quantification of extracted triterpenes by GC-FID, secondary ion mass spectrometry (SIMS) has also provided an assessment from “in vivo” samples at room temperature. For heterologous expression of the triterpene pathway in alternative hosts, the hydrocarbons are found both intracellularly and extracellularly. The highly hydrophobic nature of these triterpenes provides for relatively straightforward recovery by extraction into an organic phase. The methylation of the Botryococcus braunii race B hydrocarbons (which enhances its fuel precursor value) is readily resolved by GC-FID methods for routine analysis.
Keywords:Botryococcene Extraction Heterologous expression Hydrocarbon Triterpene
We acknowledge the generous efforts of Dr. Joe Chappell throughout our development of algae and heterologous production of triterpenes, including “in-house/hands-on” training on initial extraction and GC analytical methods. Amalie Tuerk and Justin Yoo are acknowledged for their extensive efforts in the culturing of Botryococcus braunii strains and various growth and hydrocarbon production studies. This collaborative work with W. R. C. was supported by US Department of Energy. Grant Number: ARPA-e Electrofuels, DE-AR0000092, and the National Science Foundation Collaborative Grant No. CBET-0828648 titled “Development of a Sustainable Production Platform for Renewable Petroleum Based Oils in Algae.” Any opinions, findings, and conclusions or recommendations expressed in this material are those of the authors and do not necessarily reflect the views of the National Science Foundation.
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