Effect of cultivation parameters on growth and pigment biosynthesis in flagellated cells of Haematococcus pluvialis

The volvocalean microalga Haematococcus pluvialis is used as a source of the ketocarotenoid astaxanthin for applications in aquaculture and the pharmaceutical and cosmetic industries. This green alga accumulates astaxanthin, mostly esterified, canthaxanthin and echinenone in lipid vesicles outside the chloroplast. This accumulation process normally but not exclusively accompanies formation of the resting state in the developmental cycle. With regard to increased bioavailability of the accumulated secondary carotenoids, the fragility of the extracellular matrix makes the flagellated state of H. pluvialis an interesting alternative to the thick-walled aplanospore state. A two-step batch cultivation scheme was developed that leads to accumulation of secondary carotenoids in flagellated cells of H. pluvialis (strain 192.80, Göttingen, Germany). Germination of green aplanospores during the first step of cultivation proceeded optimally under 30 μmol photon m-2 S-1 of white fluorescent light 20°C. For optimal induction and enhancement of carotenoid biosynthesis, the flagellated cells formed were then exposed to a decreased level of nitrate (0.4 mM KNO3) and to enhanced irradiance (150 μmol photon m-2 s-1). Under these conditions, which still permitted cell division and chlorophyll synthesis during the first two days of exposure, carotenoid accumulation in the flagellated cells reached 2% of dry mass at the fourth day of exposure. As a mixotrophic carbon source, addition of acetate at a concentration not higher than 10 mM increased carotenoid synthesis only slightly whereas partial or complete phosphate deficiency or salt stress (40 mM NaCl) did not