Effects of culture age on cryopreservation of marine microalgae

Abstract

The effects of culture age (3, 7 and 14 days) on viability after cryopreservation of five marine microalgae were studied. Higher viability levels were generally found in older cultures, particularly when conditions of cryoprotectant concentration and salinity were not optimal. These differences decreased or disappeared when the algae were cryopreserved under optimal conditions, except for Rhodomonas baltica. Its cryopreservation using 15% dimethylsulphoxide in medium of 20%o salinity (optimal conditions) showed 451+1−3%, 424±133% and 350±13−7% viability respectively for cells from 3−, 7− and 14−day−old cultures. For the same culture ages, the viability of Chaetoceros gracilis was 257±7'7%, 2814−9% and 360±4−6%. This diatom could be recovered from −196°C without the use of any cryoprotectant only when cells were collected from 14−day−old cultures (mean viability of 31±3−7%). The cryopreservation of Tetraselmis chuii was relatively unaffected by culture age and was very close to 100% under optimum cryoprotedant concentration and salinity. Only in the absence of cryoprotectant were cells from the exponential phase (3 days) found to be more sensitive to cryopreservation (average viability 222±5−5%)than were cells from older cultures (372±149% and 42±126% for cells from 7− and 14−day−old cultures respectively). A lower resistance to cryopreservation was found in 3−day−old cultures of Nannochloris atomus (830±110%) in comparison with older cultures (100±78% and 949−±89% for 7− and 14−day−old cultures). The main increase in the viability of this alga with age was found in the absence of cryoprotectant and using a suboptimum salinity of 20%o. Three−day−old cultures of Nannochloropsis gaditana completely failed to recover after freezing under all conditions tested. Cells from 7− and 14−day−old cultures achieved mean viabilities of c. 67%.