Assessment of bioreactors for transformed root cultures

This thesis presents a comparative study of growth and secondary metabolite production in Artemisia annua hairy root cultures in liquid-phase bubble column reactors and gas-phase nutrient mist reactors. Two reactor systems used identical culture chambers and the working volume was 1.5 L. The mist reactor was operated as a bubble column initially to distribute the roots throughout the culture chamber, and therefore initial growth in the two reactor systems was almost identical.; First, the overall biomass production was examined. When six days of bubble column mode was used in the mist reactor, the biomass density was less than 12 g FW/L (1.5 g DW/L) at the time the reactor was switched to mist mode. In that case, the mist reactor consistently produced less biomass than the bubble column reactor. However, when longer bubble column mode periods, 14 and 21 days, were used, the final biomass yield from the nutrient mist reactor was comparable to that of the bubble column reactor. The highest final biomass achieved was 14.4 g DW/L in the nutrient mist reactor and 15.3 g DW/L in the bubble column reactor with two medium exchanges and 38 days of culturing. These results suggested that a minimum amount of biomass (>40 g FW/L or 3.5 g DW/L) is needed for a successful mist reactor run. The hypothesis was investigated by using an aerosol deposition model to predict the volume of medium captured by the root bed at different packing density. The volume of medium captured was compared to the volume of medium required to maintain specified growth rates. The results suggest that under our current operating conditions, the growth rate in the mist reactor is limited to 0.08 day −1, at least in part, by mist capture efficiency.; The transient behavior of A. anuua hairy roots in different culture systems was then analyzed. From the nutrient levels in the medium, three different correlations were derived to estimate biomass in the reactors, and their accuracies were examined. The instantaneous specific growth rates in the reactors were obtained from the estimated biomass using numerical differentiation. The growth rates, together with the specific sugar uptake rates and the specific respiration rates, were used to calculate nutrient requirements and utilization rates. The actual biomass yield from sugar in the bubble column reactors was 0.493 g DW/g glucose equivalent compared to 0.437 g DW/g glucose equivalent in the mist reactors. The maintenance coefficients for sugar were 0.0423 and 0.1027 g glucose equivalent/g DW/day in the bubble column and mist reactors, respectively. In bubble column reactors, the actual biomass yield from oxygen for A. annua was 0.048 g DW/mmol O₂ and the maintenance coefficient for oxygen was 0.0676 mmol/g DW/hr.; Finally, the levels of the secondary metabolite in Artemisia annua hairy roots, artemisinin, in different culture systems were obtained. On average, the roots grown in nutrient mist reactors produced nearly three times as much artemisinin as roots grown in bubble column reactors, 2.64 μg/g DW and 0.98 μg/g DW, respectively.