Studies on frost hardiness in <italic>Chlorella ellipsoidea</italic> I. Development of frost hardiness of <italic>Chlorella ellipsoidea</italic> in synchronous culture

Abstract

Cells of Chlorella ellipsoidea Gerneck (IAM C-27) were synchronously grown under a 28-hr light-14-hr dark regime at 25°C. The algal cells at different stages during the cell cycle were hardened at 3°C for 48 hr. The survival rate of hardened cells was maximum (70%) at the L2 stage(ripening phase) in the life cycle. The average cell volume of L2 cells increased during hardening, but the process of nuclear division scarcely advanced. The hardiness of L2 cells increasedwith prolongation of hardening time up to 48 hr. Their viability decreased upon increasing the rat of cooling and lowering the final freezing temperature. But the hardened cells, which had been prefrozen stepwise, showed a survival rate above 50% even at −196°C when thawed rapidlyin a bath at 25°C. Although L2 cells were somewhat hardened in the dark, illumination was the more effective when used with bubbling gas. Under illumination, bubbling of 1% CO2-air increased the hardiness more than CO2-free air, but in the dark, this relation was reversed. The hardiness was lowest with nitrogen gas bubbling under both conditions.