The role of micronutrients and strategies for optimized continual glycerol production from carbon dioxide by Dunaliella tertiolecta.

The microalga Dunaliella tertiolecta synthesizes intracellular glycerol as an osmoticum to counteract external osmotic pressure in high saline environments. The species has recently been found to release and accumulate extracellular glycerol, making it a suitable candidate for sustainable industrial glycerol production if a sufficiently high product titre yield can be achieved. While macronutrients such as nitrogen and phosphorus are essential and well understood, this study seeks to understand the influence of the micronutrient profile on glycerol production. The effects of metallic elements calcium, magnesium, manganese, zinc, cobalt, copper, and iron, as well as boron, on glycerol production as well as cell growth were quantified. The relationship between cell density and glycerol productivity was also determined. Statistically, manganese recorded the highest improvement in glycerol production as well as cell growth. Further experiments showed that manganese availability was associated with higher superoxide dismutase formation, thus suggesting that glycerol production is negatively affected by oxidative stress and the manganese bound form of this enzyme is required in order to counteract reactive oxygen species in the cells. A minimum concentration of 8.25 × 10(-5)  g L(-1) manganese was sufficient to overcome this problem and achieve 10 g L(-1) extracellular glycerol, compared to 4 g L(-1) without the addition of manganese. Unlike cell growth, extracellular glycerol production was found to be negatively affected by the amount of calcium present in the normal growth medium, most likely due to the lower cell permeability at high calcium concentrations. The inhibitory effects of iron also affected extracellular glycerol production more significantly than cell growth and several antagonistic interaction effects between various micronutrients were observed. This study indicates how the optimization of these small amounts of nutrients in a two-stage system can lead to a large enhancement in D. tertiolecta glycerol production and should be considered during the design of a large scale bioprocess for this alternative route to glycerol.