Optimization of continuous TAG production by Nannochloropsis gaditana in solar-nitrogen-limited culture.

Nitrogen limitation and changing solar conditions are both known to affect triacylglycerol (TAG) production in microalgae. This study investigates the optimization of TAG production with a continuous nitrogen-limited culture of Nannochloropsis gaditana in simulated day-night cycles (DNc). The effect of DNc was first investigated in nitrogen-deprived condition (i.e., batch culture), emphasizing a significant change in mechanical resistance of the strain during the night. The concept of released TAG, which shows how much of the TAG produced is actually recovered in the downstream stages, that is, after cell disruption, was shown here of interest. For a maximum released TAG, the optimum harvesting time was suggested as being 4 h into the night period, which minimizes the losses due to a too great cell mechanical resistance. The protocol for continuous nitrogen-limited culture was then optimized, and a continuous nitrogen addition was compared to a pulsed-addition. For the latter, nitrogen was supplied in a single pulse at the beginning of the light periods, while the bulk medium was supplied separately at a slow but constant dilution rate of 0.005h-1$0.005\,{{\rm{h}}}^{-1}$ . The pulse dose was calculated after the study of nitrogen consumption and TAG production/consumption during the DNc. The estimated released TAG for the pulsed-addition of 1.4 × 10-3 kg/m2 d found significantly higher than the one achieved in batch culture (0.3 × 10-3 kg/m2 d) but lower than for continuous nitrogen addition which obtained the highest released TAG of 3×10-3kg∕m2 d$3\times 1{0}^{-3}\,\text{kg}\unicode{x02215}{{\rm{m}}}^{2}\unicode{x0200A}{\rm{d}}$ .

Enhanced bioproduction of anticancer precursor vindoline by yeast cell factories.

The pharmaceutical industry faces a growing demand and recurrent shortages in many anticancer plant drugs given their extensive use in human chemotherapy. Efficient alternative strategies of supply of these natural products such as bioproduction by microorganisms are needed to ensure stable and massive manufacturing. Here, we developed and optimized yeast cell factories efficiently converting tabersonine to vindoline, a precursor of the major anticancer alkaloids vinblastine and vincristine. First, fine-tuning of heterologous gene copies restrained side metabolites synthesis towards vindoline production. Tabersonine to vindoline bioconversion was further enhanced through a rational medium optimization (pH, composition) and a sequential feeding strategy. Finally, a vindoline titre of 266 mg l-1 (88% yield) was reached in an optimized fed-batch bioreactor. This precursor-directed synthesis of vindoline thus paves the way towards future industrial bioproduction through the valorization of abundant tabersonine resources.