Date of Award


Degree Name

MS in Civil and Environmental Engineering


Civil and Environmental Engineering


Tryg Lundquist


Growth media recycling during algae cultivation is necessary to increase the efficiency and reduce the cost of biofuel production from algae feedstocks. Without recycling media, the cost of algae based biofuel production would be prohibitively high and large scale algae based biofuel production would not be economically viable. The ratio of media recycled to media wasted assumed for algae farms is generally calculated to maintain salt concentrations below growth inhibitory levels, ignoring the influence of secondary metabolites which might decrease productivity. Secondary metabolites, which include allelopathic or auto-inhibitory biological contaminants, might lead to the accumulation of growth-inhibiting compounds in recycled media used in algae production.

Chlorella sorokiniana (strain DOE1412) was a leading algae biofuel feedstock candidate and has not previously been evaluated for inhibitor production. To test the effects of water recycling on the growth of DOE1412, media was recycled through multiple rounds of algae cultivation. DOE1412 was grown in modified BG11 culture media until reaching the end of linear growth phase, at which point the biomass was removed, nutrients replenished to their initial concentrations, and the recycled culture media used for a subsequent round of growth. The culture media was recycled through five rounds of growth with cultures grown on recycled media compared to controls grown on freshly prepared growth media. Biomass density was monitored via optical density and the specific and productivity growth rates were used to quantify the extent of inhibition. Exploratory work was performed with the goal of identifying potential inhibitory substances produced by DOE1412 during cultivation. Samples of recycled media were analyzed for polyunsaturated fatty acids which have been demonstrated to be inhibitory. The carbohydrates content of used media was analyzed to assess the amount of organic materials shed by DOE1412 into recycled media during growth.

The log phase growth rate (day-1) of DOE1412 was inhibited by 3±2%, 8±1%, 10±2%, and 18.6±0.9% when grown in media recycled 1-4 times, respectively, with a 99% level of confidence that inhibition was observed in each round of regrowth. The productivity growth rate (OD750/day) of DOE1412 was not inhibited in media recycled 1-3 times. The productivity growth rate of DOE1412 was inhibited by 13±3% when grown in media recycled 4 times with a 99% level of confidence that inhibition was observed.

Zinc was found to accumulate in the recycled media to potentially toxic levels (>0.09 mg/L), therefore it is uncertain if the observed inhibition was due to an accumulation of inhibitory secondary metabolites or the accumulation of zinc.

Two inhibitory polyunsaturated acids, linoleic and linolenic acid, were identified in media recycled 4 times. The carbohydrate content of recycled media fluctuated between 8-10% of total fixed carbon in media recycled 1-3 times and increased to 18% in media recycled 4 times. However, changes observed in media recycled 4 times may have been due to improper storage of used media.