Hydrocarbon production in high density Botryococcus braunii race B continuous culture

Continuous cultures of Botryococcus braunii race B were maintained at photosynthetic cell densities as high as 20 g dry weight per liter for up to 3 months. Growth associated triterpene hydrocarbon accumulation was nearly constant at 22.5% of dry weight for a range of growth rates maintained by daily replacement of 5-15% of the respective cultures. The ability to achieve high cell concentrations and oil levels of roughly 5 g triterpene oil/L resulted from a combination of high light (∼ 1/4 full sun for 15 h/day) and replenishing stoichiometrically balanced growth medium. Due to light-limited growth conditions, cell concentration dropped nearly linearly with increased dilution rate. This reduction in cell number resulted in increased productivity per cell at higher dilution rates and was accompanied by a dramatic increase in algae colony size from 0.09 to 0.343 mm at high dilution rate. This change in colony size resulted in an equally dramatic change in optical density (OD) per gram dry weight, which precluded use of simple correlations of OD and cell concentration. A trickle-film photobioreactor was also demonstrated as a scalable approach to achieving these ultra-high cell concentrations. Additional media analysis revealed a steady increase in photobioreactor conductivity suggesting an accumulation of ions may be the reason for rapid culture crash and washout observed at all dilution rates after several months of continuous operation. The volumetric productivity of 22.5 mg oil/L/photo-h reported here is more than an order of magnitude higher than previous reports for B. braunii race B, reflecting the high cell densities used in this work and substantiating a higher metabolic rate for B. braunii race B than previously surmised from its relatively long doubling times.